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Data Acquisition and Analysis Software User's Manual

1. es 6 Add Line Cut Paste Print Load Save lt Prev Next gt _ Finish Close Help The following peak events are supported e suppress peaks A time interval is entered from which peaks will be removed Chapter 10 4 8 Automatic Shoulder Peak Recognition Automatic shoulder peak detection is set up on the sixth page of the peak find dialog box Refer to chapter Chapter 10 10 7 for details Chapter 10 4 9 Normalising Peak Widths Some versions of DAx have the ability to normalise peak widths For instance in DNA analyses it may be known that peaks are normally 1 base pair wide Peak widths are normalised using page 7 of the peak find dialog box 120 5158 4 Dal IV Nomalise Peak Widths 5158 4 Dal STAND 1 Da1 STAND 1 Da1 Peak width unit BP v fa Normal left peak width 050 A 2 right peak width oso WA 3 Peak cut off signal limit f10 0 WA ale al eig lt Prev Next gt Finish Close Help Peaks are normalised using a number of parameters ls unit This is either seconds unity or the calibrated unit normally BP It is possible to express the expected peak width in BP units even when the data have time coordinates along the horizontal axis The reverse is not possible time units cannot be used to denote the expected peak width if the data have been converted to a calibrated horizontal axis left peak width is the normal expected difference between the l
2. 5 enter the component names in the name column in the peak list chapter Chapter 10 11 4 6 assign peak types to the peaks as needed 7 enter peak annotations as needed 8 enter the component concentrations in the concentration column in the peak list Component names and concentrations may also be entered by right clicking a peak top marker then entering data into the peak popup dialog box 9 if marker peaks are going to be used to recognise components enter marker peak names for those peaks for which marker peaks will be used Marker peak names can be set by right clicking a peak top marker then entering data into the peak popup dialog box or by highlighting one or more lines in the peak list window clicking the right mouse button and using the Set Marker Peak menu option to choose a marker peak name from the list of named peaks 10 invoke the Analysis Edit Database menu option discussed in detail in chapter Chapter 18 4 Chapter 18 2 Analysis Load database Analysis Save database Analysis Stop analysis menu options Identification Databases may be saved to disk using the Analysis Save database option They may be loaded from disk using the Analysis Load database option If the Separate Parameter Files item in the File Customise gt Extensions dialog box is unchecked Identification Databases can only be saved as part of an analysis procedure so the Load Save database menu options are not displayed If th
3. Total Concentration Identification Database not available available relative concentration area relative concentration area fraction fraction not available concentration not concentration relative calculated concentration total concentration concentration calibrated concentration calibrated available relative concentration relative concentration calibrated sum of calibrated calibrated total concentration Chapter 10 11 9 Only List Named Peaks Use the View Only Named Peaks menu option to hide all peaks in the peak list window that have not been recognised by the qualification module cf chapter Chapter 18 and have not been assigned a peak name manually Chapter 10 11 10 Only Show Peaks of Certain Types Use the View Select Peak Types menu option to specify which types of peaks should be displayed Peak types can be entered in the peak list window or by placing the mouse cursor over the top marker of a peak clicking the right mouse button then selecting the peak type in the dialog box that appears A peak can have multiple types Chapter 10 11 11 The Peak List window Popup Menu Peak list windows display a popup menu when the right mouse button is clicked The menu can be used to e locate or highlight peaks in the Data Set Graphic window cf chapter Chapter 10 11 3 e delete peaks cf chapter Chapter 10 11 2 e mark a reference peak cf chapter Chapter 1
4. Help 1 The Operation item determines the kind of arithmetic operation that will be performed on the selected data sets Some operations can only be performed using a constant operator power calculation time coordinate adjustments and some operations can only be performed using an operator curve MIN MAX curves dCurve dCurve 2 The Operator Curve is the data set that will be used to perform an operation on all of the data sets that have been selected in the Data Selection list box Operator curves are used in arithmetic operations and to de convolute data In both cases the operation cannot be performed on the operator data itself so the operator data set will be disabled in the data selection list box If New Curve or New Window which automatically means New Curve as well has been selected this restriction does not apply Chapter 9 11 Data Average This command invokes a dialog box that lets you select data sets to average Typically the average data set will be used as a blank to which other data sets can be compared A single overall average data set can be calculated or one average data set per enslavement group The Select all button can be used to select all data sets that contain a given string of characters in their names Enter the string or choose one of the pre defined strings listed Check deselect to deselect all data sets that contain the string If the Alt key is pressed when the Selec
5. This option is activated automatically for the calibration trace in trace files and is best limited to that specific use Chapter 17 8 Calibration Windows Calibrations can be investigated in two types of windows There is a text window listing the calibration points and the parameters for the calibration polynomial There is also a graphics window that displays the calibration points and the polynomial Use the Calibration Calibration list and Calibration Calibration curve menu options to display these windows Chapter 17 8 1 Calibration List window The Calibration List window has the usual text window functionality chapter Chapter 1 It also has the option of deleting rows to remove calibration points To remove a point click the right mouse button on the line containing the point and select the Delete entry option in the pop up menu that appears These are the columns in the Calibration List window e point index e polynomial fit parameters applicable only if the calibration uses a polynomial e usage points are either included in or excluded from the calibration 181 e calibration parameter values e calibrated values e check value applicable only for polynomial calibrations The check value is the calibrated value calculated for the calibration parameter in the row using the current polynomial e data source name of the measurement the data in the row were derived from Chapter 17 8 2 Calibration Curve wind
6. 10 5 On data with very distinctive linear or higher order trends it can be useful to temporarily remove the trend for baseline construction The optimum trend to be used is automatically determined 6 The Close button closes the dialog box parameters will be adjusted to the values entered but no baselines will be constructed Chapter 10 1 1 Using Gradients If a gradient has been defined DAx has the option of using it as a basis for the baseline to be constructed Moving Average and DerLim baselines only Check the Use HPLC Gradient box in the baseline construction dialog box to use the gradient Only the time coordinates at which the gradients occur will be used signal values will be derived from the data set For measurements where the gradient has caused a significant amount of baseline drift using the gradient can help to create much more accurate baselines Chapter 10 2 Peaks Subtract baselines This command invokes a dialog box that allows baselines to be subtracted from data sets When a baseline has been subtracted from a data set operations that normally require a baseline to have been calculated no longer require an explicit baseline Instead a signal value of zero is considered to be the baseline Because of GLP chapter Chapter 3 3 2 raw data sets cannot be overwritten This means that the new window checkbox has to be marked or a data set must already have been modified must be Modified Data chapter
7. 64 76 77 86 104 Sizing standard 183 skew 137 226 skim 131 slant 130 slope requiring threshold 118 sorting data 99 sorting data sets 91 spike 85 94 147 148 spikes 96 spline adding nodes 63 splines 24 79 111 adding spline nodes 82 cubic splines 29 definition 29 modifying splines 81 removing spline nodes 81 spline nodes 29 64 81 spreadsheet 43 45 Stacking data 105 statistical analysis 204 statistical mode 107 Statistics Lists 231 232 subtracting baselines 110 subtracting gradients 157 suppressing peak detection 120 257 system requirements 13 tacks 83 151 adding 63 temperature programme 156 157 terminology 23 text column width 44 text variables in reports 241 text windows binning sheet window 207 calibration list 22 calibration list window 41 comparison sheet window 204 configuring 43 copy 45 data value list windows 20 data value list windows 41 data value table windows 20 41 deleting lines 42 editing 41 editing contents 41 GPC calibration list 21 GPC calibration list window 41 highlighting lines See Selecting Lines Identification Database list 22 navigating 42 peak list windows 20 peak list windows 61 peak list windows 132 pop up menus 42 quantitative calibration list 22 selecting columns 43 selecting lines 41 selecting text 42 45 258 theoretical data 101 threshold 74 112 time corrected signal axes 21 173 time displacements
8. Chapter 9 15 Data Generate data If the menu option Data Generate Data is not present you need to use File Customise gt Data and check Data Generating Not all versions of DAx contain this option This command invokes a dialog box that allows pure theoretical data to be generated 11 Available data curves are a Gaussian curve of selectable width a symmetrical triangle an asymmetrical triangle a block function an exponentially distorted Gaussian curve These are used to model tailing or fronting peaks The equation for the curve is 11 The command must be invoked from the Edit menu as Edit Generate Data when no windows are open yet If a window is open use the Data menu 101 2 25 A OG T 2 rT Z x2 f t e e dx with te the location of t 7 2 a G the peak top og the peak width A the peak area t the t te _ oG OG T asymmetry factor and Z y y JT e a Fraser Suzuki skewed Gaussian curve The equation for the 2f p1 400 a curve is f t He 7 maz with ta the location of the peak top o the peak width H the peak height and A the asymmetry factor cf Fraser amp Suzuki Anal Chem 41 1969 37 You may choose to not add noise or you may add block noise or Gaussian noise Block noise has a block distribution whereas Gaussian noise has a normal distribution Chapter 9 16 Data Overlay The Data Overlay dialog box is used to move or stretc
9. Choose between user defined bins where the binning interval of each bin is determined by the user and automatically derived bins where enough bins are derived to just contain all peaks in all selected data sets Data set selection User defined or automatic bins Qualifying coordinate Trace types User defined bins Bin intervals Peak type use and selection Curve samples and peak parameters Curve sample scaling Quantifying parameter labels Peak parameter selection ee n St aie al iteme Determine if user defined bins should be turned into peaks No Display options peaks will exist outside the user defined bins and each bin will Peak thresholds correspond to precisely one peak Bin annotations Persistent data C Automatically derived bins User defined bins can only use peak top coordinate or peak top base pair count as the qualifying parameter Tl Bins equal peaks This option should be used when the shape of the curve in a bin is important rather than whether or not the curve in the bin can be considered a peak Determine if multiple peaks can be listed for each user defined bin This option cannot be used if bins equal peaks lt Prev i Multiple peaks per bin Maximum number of peaks listed per bin 2 Al Leave the maximum number of peaks listed blank to list all peaks in OK Cancel Apply the bin Checking User defined bins allows the user to define bins rather than h
10. Data Lists 231 232 data modification 18 data set 20 25 26 28 29 47 59 61 63 64 69 76 79 81 85 89 91 92 100 103 104 112 definition 24 data set name 26 27 89 definition 26 data tag 24 25 47 64 69 77 81 82 89 91 92 104 161 deconvolution 100 default data set 70 72 81 111 143 definition 25 deleting data 70 deleting lines from text windows 42 deleting peaks 133 derivative 29 74 85 94 97 112 derivatives 94 112 deriving filter characteristics 146 DerLim 108 descriptive labels 152 descriptive tacks 151 deselect check box 57 91 92 98 116 dilution 79 dilutions 199 Directory for default filenames See Measurement Window manual display Analysis Assay 229 displaying data set names 60 drag 69 dragging 41 69 82 86 89 130 drift 226 removing 247 drift factor 96 drifting baselines 107 drivers 14 16 dropping files 52 editing in text windows 41 editing reports 237 editing scale 60 effective mobility 169 172 emancipate 90 enslavement 26 EOF 246 error log 17 24 56 exponential Gaussian curve 101 exporting reports 243 extensions 15 external marker peaks 197 extra input fields 19 filter 85 94 filter characteristics 146 filtering 23 144 Fourier filter 94 moving average filter 85 94 147 Savitzky Golay filter 85 94 147 finding data 233 finding peaks 112 251 fitting Gaussians 123 fixating analysis results 124
11. When multiple peaks are listed per bin the peak with the highest value of the quantifying parameter is considered the main peak Functionality that uses a single peak per bin such as export of Nexus files uses the main peak in each bin Qualifying coordinate If user defined bins are used either peak top coordinate or calibrated value at peak top typically base pair count is used as a qualifying coordinate chosen on the Qualifying coordinate parameter page If user defined bins are not used a full range of qualifying coordinates is available A tolerance value must be entered to determine the width of automatically assigned bins Peaks must have qualifying parameters within tolerance of each other to be assigned to the same bin Trace types Bins can be limited to certain trace types so that only data from data sets matching the specified trace type will end up in the bin Each bin can have its own trace type This option can be selected on the Trace types parameter page 210 Bin intervals User defined bins can be set up on the Bin intervals parameter page as shown below They can also be defined in the graphic or colour view window see chapter Chapter 18 12 3 amp Binning Sheet Setup 2 x Setup parameters Enter the user defined bin intervals here Interval can also be added from the right mouse click popup menu in the binning sheet or from a colour view window Data set selection User defined or autom
12. data set names and curve samples 1 5 If you click to the left of the check mark or click an unchecked data tag the selection will be removed 58 Chapter 7 2 5 Edit Create metafile This command creates an Aldus placeable metafile of the contents of the graphics window This metafile can be read into drawing programs You will be given the option of including data set names and curve samples See the examples above Chapter 7 2 6 Edit Undo Edit Redo These commands cancel or redo a peak modification data set sizing spline modification or tack or label command depending on which of these commands was last executed Chapter 7 3 Data Set Graphics Window View menu The view menu contains options that determine which data sets are displayed and in what way Chapter 7 3 1 View Attributes This command invokes a dialog box that configures many aspects of the graph Refer to chapter Chapter 5 2 4 for more information Chapter 7 3 2 View Plot Select This menu option is discussed in chapter Chapter 9 2 Chapter 7 3 3 View Plot all This menu option makes all data sets in the window visible Chapter 7 3 4 View Plot previous View Plot next View Plot data sets View Plot groups The plot previous and plot next commands show only the previous or next data set or group of data sets as chosen using the View Plot data sets and View Plot groups commands 59 Press the shift key while executing the c
13. pushbutton Clicking the button will display a dialog box which will let you enter values 41 Chapter 6 1 3 Deleting Lines If the text window has the option of deleting lines click the right mouse button on the line you want to delete A pop up menu will appear Click the Delete Row item to delete a line To delete multiple lines select the lines then use the Delete Selection menu item in the pop up menu Chapter 6 1 4 Pop up Menus Some text windows will display a pop up menu when the right mouse button is clicked on a line The pop up menu may contain various commands Refer to the chapters on the specific text window you are using for further details A different popup menu will be displayed when the right mouse button is clicked on the column header Typically the popup menu will contain options to Fit columns making columns wide enough to display their content and Shrink and fit columns making columns wide enough for their contents but making them narrower where possible Sometimes there is an Equalise column widths option which will make all columns of the same type as the column clicked on have the same width Chapter 6 1 5 Navigating Text Windows Move to a new position in a text window using e Shift Tab to move left or right not for selecting lines In dialog boxes the tab key moves between controls in the dialog box e Cursor Left Right to move left or right e Cursor Up Down to move up or down e Page Up
14. the object to a new location All objects are placed on positions that are multiples of millimetres Size an object by clicking on one of the squares at the corners and dragging the mouse Objects can also be moved by using the cursor keys Select the object or multiple objects as above then use the left and right cursor keys to move objects left and right in steps of one millimetre The up and down cursor keys will move objects up and down one millimetre To select the next previous object use the shift tab key NB Objects are ordered in the order they were entered in However when a report definition is loaded from disk the objects are re ordered from left to right top to bottom and across pages Chapter 20 3 2 Changing the Type of an Object To change the type of an object click the right mouse button on the object A pop up menu appears which lets you set a new object type except for line and text objects This is very useful if multiple objects of the same size but different types are needed Simply copy the objects using the Ctrl key and the mouse then right click the new object and change its type 237 Chapter 20 3 3 Editing Multiple Objects To edit multiple objects click the first object then press the Ctrl key and click any other object to add it to the selection You may now move all selected objects simultaneously by again pressing the Ctrl key clicking any of the selected objects and dragging th
15. use the Config button to set up sorting Data can be displayed as a colour view by checking the Colour view box The data can be stacked by checking the Stacked box use the Config button to configure stacking options Chapter 7 1 2 1 Automatic Analysis By clicking the AutoAnalyse check box in the file load dialog box all files that are loaded will be marked for automatic analysis Click the Config button to enter the automatic analysis preferences set up dialog 4 Separate automatic analysis setups are used for new measurements and for files that are opened with the Automatic Analysis option checked 49 i AutoAnalysis preferences De On Filtering il Filter Overlay ll Baseline l2 i3 MV Construct Subtract amp remove Peaks v Find eas Fie M Print Only named peaks le l Close peak list windows Add to Global sheet 8 Add to Binning sheet r 0 l Repor Curv Derive calibration 7 Convert axis to calibrated Derive GPC calibration Convert axis to Mw Convert axis to mobility rc Cancel Help 1 If the Filter option is active the data will be filtered before a baseline is constructed The most recently used filter parameters will be used or you can click the blue underlined Filter button to configure filtering parameters 2 Check the Baseline construct box to automatically construct a baseline when a measurement finishes The most recently used baseline construction
16. ForeGround jo WA B WA a WA Axis Area oso pA 1 y jaro J x0 fos p4 pT Tl Apply to all graphics windows OK Cancel Apply Help 1 Pen thicknesses are in device units font sizes for screen and printer are in points 1 point 1 72 clipboard font size is in units where the clipboard has a size of 2000 1500 units Chapter 5 2 5 View Select curves This command invokes a dialog box that allows the curves that are displayed in the window to be shown or hidden Note that the term curves is used both for lines and collections of point markers Chapter 5 2 6 View Edit scale This command invokes a dialog box that allows the scale to be set Each window has one horizontal axis but may have several vertical axes The scale for all of these may be entered Scales can be entered globally or zoomed Zoomed scaling can be undone by zooming out using the View Zoom out menu command or by clicking the right mouse button Several levels of zoomed scale can be entered 39 Chapter 5 2 7 View Zoom in The zoom command zooms in on the centre one third of the graph Chapter 5 2 8 View Zoom out Zoom Out undoes one level of zoom generally ten levels are available Chapter 5 2 9 View Reset scale Reset scale undoes all zooming and optimises the scale so that all curves will be optimally visible 40 Chapter 6 DAx Text Windows DAx contains several types of text window
17. In the Command Line box type R autorun Click the OK button The set up menu will now start Select DAx Master Setup and follow the instructions on screen Run the Install USB dongle program that has been installed in the DAx program group This should be run before the USB dongle is attached Follow the instructions on screen Attach the USB dongle after installation completes This completes the installation Set up has created a Start Menu group containing the DAx icon Double click on the icon to start DAx Chapter 2 2 1 Drivers DAx uses an external driver to perform the hardware specific aspects of measurements To select a driver the menu option File Customise gt StartUp and Measurement Drivers should be used 14 Chapter 3 Customising Window Types Terminology Chapter 3 1 Customising DAx DAx can be customised to a large extent To start customising use the File Customise menu option A dialog box will be displayed that has the options listed below Chapter 3 1 1 File Customise gt Extensions Any or all of the following extensions can be included in DAx HPLC extensions GPC extensions CE extensions Calibration extensions and Qualification Quantification extensions In the case of GC HPLC CE and GPC extensions DAx has the option of making the status of the extensions depend on the measurement data type For instance GPC extensions will then only be shown and accessible if a measurement is of
18. Refer to chapter Chapter 11 for details on the waste barrel Chapter 3 3 11 Baselines There is a large category of measurements where a signal is measured in time to determine the presence and possibly the abundance of components in a sample These measurements share in common the fact that they use some way to separate components that is to make one component appear in the measurement signal before the other In order to be able to determine when a component shows up in the measurement signal we need to establish the form the signal would have taken if a particular component had not been present in the sample This form of the signal is called the baseline Only when the measured signal differs from the baseline may we conclude a component or multiple overlapping components are present DAx contains several powerful baseline construction algorithms Refer to chapter Chapter 9 27 Chapter 3 3 12 Blanks Blank measurements theoretically make for ideal baselines since they are the essence of what a baseline is the signal one would get in the absence of any measurable compounds Chapter 3 3 13 Peaks When a baseline chapter Chapter 3 3 11 has been established for a signal we can search for peaks i e intervals where the signal shows a sustained difference from the baseline Peaks ideally are either Gaussian or triangular in form Chapter 3 3 14 Normalisation Peaks Normalisation peaks are used to overlay data sets and to
19. Select one of them then click the Copy key to copy the sizes and calibration trace identity 2 These items are used to copy settings from a Genographer configuration file 185 Automatic Trace Calibration 2 xi Trace type amp fragment sizes Import sizes Restrictions Calibration Advanced J Require minimum number of calibration points 23 Al 4 I Skip initial 00 1 2 MV Require minimum peak area 0 20 pJ C Absolute Relative l3 I Use largest peaks 50 Al l 1 Require minimum number of calibration points will often be set to the number of standard sizes but can be set to a lower value if the sample may not have been analysed completely i e if the analysis was cut short 2 Skip initial can be used to disregard the early part of a measurement which speeds up the analysis but is not required 3 Require minimum peak area can be used to disregard small peaks which again speeds up the analysis but is not required Values of 0 2 or 0 4 generally work well 4 Use largest peaks is another way to disregard small peaks A good value is 2 the number of calibration peaks 186 Automatic Trace Calibration 2 xi Trace type amp fragment sizes Import sizes Restrictions Calibration Advanced Select the type of calibration curve that should be derived from the data Calibration Curve Type j Multilinear Cubic Spline Polynomial Degree 2 Al C Local Southern Global
20. Southern 1 The Calibration curve type defines what sort of curve will be drawn through the sizing calibration points Use Analysis Save ATC parameters to store ATC parameters Analysis Load ATC parameters to load ATC parameters Use Analysis Stop ATC to remove the ATC parameters from memory Analysis Perform ATC automatically can be used to turn the use of ATC parameters on and off without removing the parameters from memory If the use of ATC parameters has been switched off when the ATC dialog box is closed you will be asked if it should be switched on If ATC is not performed automatically it can still be invoked manually from the data tag peaks menu cf Chapter 8 3 NB Automatic trace calibrations take precedence over the Identification Database chapter Chapter 18 and marker peaks to recognise peaks However after an ATC has been performed the Identification Database and marker peaks will still be applied to the data 187 Chapter 18 DAx Qualification amp Quantification Identification Database Extensions If the menu option Analysis is not present you need to use File Customise gt Extensions and check Analysis Options DAx has a number of options that make it possible to set up a database of peak properties for certain components The database can be used to determine which components are present in an unknown sample The database can also be used to determine the concentrations of components
21. The polynomial and global Southern calibrations do not necessarily intersect any points Use the Calibration Calibration Curve menu option to display the calibration as a graph Refer to Calibration Windows chapter Chapter 17 8 for details on how to include or exclude individual points from the calibration Points can also be excluded by using parameter limits 3 The Parameter limits determine which points will be used in the calibration The limits are also used in both drawing Calibrated axes windows and calculating peak data Any data points outside the time limits will not be plotted or used in the calculation Use the derive from data check box to make the limits equal to the range of calibration parameter values in the calibration list 4 The Calibrated Value limits determine which values calculated when plotting Calibrated or calculating calibrated peak data will be accepted as valid Use the derive from data check box to make the calibrated value limits equal to the range of values in the calibration list for points that are within parameter bounds 5 Do not alter points usage ensures that the same points will be used in the new calibration that were used in the old calibration This option is useful when points have been manually excluded from the calibration 6 Add to existing calibration is used to add calibration points to an existing calibration If the option is not checked a new calibration will be derived from the select
22. a WA i Ea HPLC Solvent Gradient za 30 r 7 Cancel Help A WA WA Al a 1 The parameters entered in this dialog box will be stored with the data sets selected here Other data sets will retain their old gradient parameter values 2 Gradient signal values should normally be derived from the data Time limits must be set manually as must gradient percentages or programme temperatures 3 Select between HPLC gradient and GC temperature programme here Chapter 14 3 HPLC GC Load gradient HPLC GC Save gradient menu options Gradient parameters may be saved to disk using the HPLC GC Save gradient option They may be loaded from disk using the HPLC GC Load gradient option f the Separate Parameter 156 Files item in the File Customise gt Extensions dialog box is unchecked HPLC parameters can only be saved as part of an analysis procedure so the Load Save gradient menu options are not displayed If the gradient parameters have been changed but have not been saved to disk yet when DAx is about to terminate you will be asked to confirm the loss of the new parameters Saving the parameters to a floppy disk does not prevent this you must save to hard disk Chapter 14 4 HPLC GC Subtract Gradient menu option Gradients can be subtracted from data using the HPLC GC Subtract gradient option Subtracting the signal gradient does not affect the gradient percentages or programme temperatures t
23. all This command selects highlights all text in the window Certain commands work on selected text Edit Copy File Export selected Chapter 6 2 7 Edit Copy Edit Copy using Tabs These commands copy the contents of the text window to the clipboard as ASCII text The contents of the clipboard can be pasted into a word processor or spreadsheet If Copy using tabs is used data columns will be separated by TAB characters which makes it easier to import the data into spreadsheets 45 Chapter 7 Data Set Graphics Windows Data Set Graphics windows often named graphics windows for short are used to represent data sets by plotting the data versus a horizontal coordinate Each data set is represented by a data tag which can be used to manipulate the data Click on the data tag to display a pop up menu or drag the data tag to move the data Refer to chapter Chapter 7 for complete details on data tags Data set graphics windows have the normal menu commands that each graphics window in DAx has These include commands to copy the graph or print it Zooming scaling commands and set up commands Refer to chapter Chapter 5 2 for details Data set graphics windows also have the normal zooming features chapter Chapter 5 1 Additionally if you press the Shift key while zooming in on the graph window by clicking and dragging a zoom box the vertical scale will be optimised to contain all data in the zoomed horizontal ranges If
24. around any changed spline nodes to be replaced with the spline whenever a spline node is changed 4 Use the Undo and Redo menu options to cancel or redo the last spline modification operation 80 K Chapter 8 7 Modifying Splines Splines chapter Chapter 3 3 16 are used in DAx to modify curves and to create new curves notably baseline curves A spline is displayed as a series of node markers connected either by straight lines or by cubic polynomials as is shown in the example below Use the Add Spline option in the data tag spline menu to add a spline to a data set You may also use the add spline toolbar button a spline will be added to the default data set chapter Chapter 3 3 4 Example of curve with cubic spline added Nodes have been moved to make them more easily visible When a spline is added to a data set DAx makes an effort to initially place spline nodes at the most critical points Spline nodes can be modified using the mouse Move the mouse cursor over one of the nodes It will change into a drag cursor consisting of four arrows e you may drag a node to a new location e if anode is dragged beyond the following node that node will be removed If there are twenty nodes you may drag the second one all the way to the right to remove up to seventeen nodes You may also drag nodes beyond preceding nodes to remove the preceding nodes e to remove a node drag it to a point outside the graph Alternatively clic
25. averaged each N data points are replaced by one data point with the average signal value The time coordinate of the point is also the average of the time coordinates of the N points e f data are yanked one point will be kept then N 1 points will be discarded The time coordinate for the point that is kept does not change STAND 1 Dal Data points in measurements 3903 3903 3906 MEJ stanp 1 ba1 36 O aea Include data til s 0 Fa 4 5158 4 Dal Reduce points by factor 2 Al l Calculate Averages lt na gt M New Window V New Curve Cancel Help 1 The End Time item is used to set a cut off time Any data beyond this time will be discarded from the data set If using the end time would leave less than 5 data points the operation will not be performed An end time of zero means no end time will be applied 2 Check the Calculate Averages check box to average data otherwise data will be yanked 3 Resultant data point count Curves that have not been selected are listed as lt na gt for not applicable Points after reduction peak Ea raa Chapter 9 5 Data Interpolate If the menu option Data Interpolate is not present you need to use File Customise gt Data and check Data Interpolatng This command invokes a dialog box that allows interpolated values to be added between existing data points The number of new points to be interpolated between existing points can be set The type of interpol
26. axes in units of seconds and units of minutes All graphics windows are affected Peak lists and numerous dialog boxes are also set up using the unit selected here Chapter 7 3 13 View Detector units curves View First derivative curves View Second derivative curves View Inter curve derivatives View Frequency domain curves These commands determine if certain types of data are to be plotted Frequency Domain curves are representations of data in the frequency domain Frequency domain data are available for data sets that have had a Fourier bar chapter Chapter 11 1 added to them there is no type of data set that has only frequency domain data Chapter 7 3 14 View Track data set Use this menu option to switch data set tracking on or off When data set tracking is on when the mouse is positioned on a curve the corresponding data tag is depressed Press the Shift key to temporarily stop tracking the mouse Chapter 7 3 15 View Mark bins Do not mark bins Chapter 7 3 16 View Mark bins Mark above graph Chapter 7 3 17 View Mark bins Mark above and inside graph 61 Use these menu options to start or stop marking bins along the top of the graph Cf chapter Chapter 18 12 3 If user defined bins are marked inside the graph they can be edited using the mouse Click the right mouse button on a binning line to display a binning popup menu cf Chapter 18 12 2 Bins will only be marked if the binning sheet s
27. baseline drift up into the peak The width can be specified as a percentage of the length of the data set expressed in horizontal axis units or specified as a number of data points Note that the precise meaning of the width parameter is different for different types of baselines so a width value that works for one type may be too small or large for another type 2 Higher numbers of passes will tend to yield better baselines However there is often an optimum number of passes Use automatic pass count detection to optimise the number of passes Generally excellent results can be achieved with three or four baseline passes Even if automatic passes are used the number of passes indicated in the dialog box is relevant it is the maximum 3 The percentile value determines how many of the data points are used to construct the baseline The unused data points are thought to lie in peaks and are thought not to be part of a baseline Lower percentile values use less data The percentile value has a lower limit of 40 no analysis should ever contain more than 60 peaks 4 The straight under peaks option performs a very sensitive peak search after the baseline has been constructed Any part of the baseline lying under a peak is then replaced by a straight line 109 This option is best used in conjunction with a moving median baseline using automatic or a low width 1 or 2 a single pass automatic percentile and a low median percentage
28. be performed on the data e drag the tags to move data between windows or create a new window You drag a tag by clicking the mouse button on it then moving the mouse cursor while keeping the mouse button depressed then releasing the mouse button in the window that you want to move the data to Multiple selected tags can be dragged at once see below for details on selecting multiple tags When data are dragged to a graph window drop onto a data tag to insert the dragged data before that tag Drop anywhere else in the graph window to find the best insertion point automatically or press the Shift key when dropping to add the data after all existing data Keep the Ctrl key depressed while dragging the tag to copy data otherwise data is moved If data for a running measurement are copied only a snapshot of the current data in the measurement will be created The copy will not receive any further points from the measurement e A single data tag can be dragged and dropped inside its own graph window to change the order of the data sets Dropping 69 onto an earlier tag moves the data set in front of that tag dropping onto a later tag moves the data set behind that tag e Moving data to a blank part of the DAx client area will create a new graphics window e Moving data to the waste barrel chapter Chapter 11 destroys the data set e Dropping data onto a comparison sheet or binning sheet causes the data set to be added to the sheet Da
29. by area of peak width of right side divided by width of left side at 10 of height number of theoretical plates calculated as toptime widthathalf height number of theoretical net plates calculated as plates 5 54 net plates 5 54 toptime referencetime widthathalf height number of theoretical plates statistically calculated as 2 plates sev Foley Dorsey plates current plate number and maximum achievable resolution resolution at half height skew kurtosis plate number in the absence of asymmetry Refer to Anal Chem 1983 55 730 737 for details Foley Dorsey plate numbers are available only for generic and GC data types resolution between a peak and the previous peak Lrop pear 10 previous width width peak previous 2 calculated as R bop Slop R 1 177 pe Pe whh peak T third peak moment of peak times with respect to signal Leading peaks have negative skew trailing peaks have positive skew Skew is calculated as S signal me a ee fourth peak moment of peak times with respect to peak signal Sharper than Gaussian peaks are leptokurtic positive kurtosis flatter than Gaussian peaks are platykurtic negative kurtosis In between peaks are skew 137 mesokurtic Kurtosis is calculated as 5 endl cen M4 ji skew 3 gt T The following items are Molecular Weight related elution volume elution volume at peak t
30. by pressing the Ctrl key and dragging the mouse across the intended bin Click the right mouse button on a binning line to display a binning popup menu cf Chapter 18 12 2 Chapter 7 8 13 Colour View Mouse Tracking Behaviour If bins are marked and a binning sheet chapter Chapter 18 12 is displayed when you move the mouse cursor across the colour view the bin in the binning sheet that corresponds to the mouse 66 location is brought into view and highlighted unless you press the Shift key 67 Chapter 8 DAx Data Set Tags DAx uses small tag windows that look like buttons to represent data sets 5 DAXSIM4 PRN x B DAXSIM1 PRN Data tags The yellow B indicates the data are being used as a baseline The tag windows are displayed in Data Set Graphics windows chapter Chapter 7 3 for all data sets Data tags are also shown in Measurement Windows for each available measurement channel Data tags are used in a number of ways e click on the peak list button at the left side of the data tag to display a peak list window cf chapter Chapter 10 11 Right click on the peak list button to display a popup menu with peak related menu items Baselines do not have a peak list button Data tags for running measurements have a tack button instead of a peak list button Clicking the tack button will add a timed comment tacks to the data e click on any other part of the data tag to display a menu of operations that can
31. component Obviously many components may have calibrations To display them several Quantitative Calibration List objects should be created with different object indices set in the upper left corner of the Report Editor FontBar Components can also be referred to by name in a report definition Quantitative Calibration Graph This is a plot of the quantitative calibration Obviously many components may have calibrations To display them several Quantitative Calibration List objects should be created with different object indices set in the upper left corner of the Report Editor FontBar Components can also be referred to by name in a report definition Statistics This is a statistics list based on the current Data List Fourier Characteristics List This is a list of the Fourier characteristics of a data set Fourier Characteristics Plot This is a plot of the Fourier characteristics of a data set Colour View A representation the measurement data where signal values are denoted using varying colours Binning Map A representation of the binning sheet where the strength of the quantification parameter in the bins is turned into varying colours 240 Chapter 20 4 1 Spreading report objects across multiple pages Certain report objects such as peak lists comparison sheets and binning sheets can contain a long list of data that will not fit into the item as it was defined on the report In this case the report page will be
32. dal Ge TESTST dal Testib dal TATEST3e dal Ta zoefdehaas da1 File name TEST3 DAX Save as type Dax files dax da1 7 Cancel Example of File Save As dialog box DAx can save data in a number of formats selected using the Save as Type combobox e DAx files have the most complete format e WinVolt files contain little more than bare measurement data e Voltage text files are text files containing a list of data point values if multiple data sets with the same timing offset and frequency are saved DAx will create a single multi column text file e Time Voltage text files as Voltage text files but with time coordinates in the first column e Andi AIA CDF files use a standard format supported by many instrument vendors The File Save command saves all data sets to the files they were loaded from The command will not work on running measurements It will also not work on data sets that were not loaded from DAx files but for instance from text files The File Save command is disabled unless file overwriting has been enabled under the GLP settings cf Chapter Chapter 3 1 3 Chapter 7 1 4 File Close This command closes the graphics window All peak list windows chapter Chapter 10 11 data value list windows local calibration curves or lists and graphics windows employing alternate horizontal axes belonging to data sets in the window will also be closed If the window contains any data sets that have n
33. data type of a data set some coordinates may not be available for use as a qualifying parameter For instance mobility cannot be used to find peaks in HPLC measurements The qualifying parameter can be used in several ways to determine which is the marker peak e near coordinate by setting the coordinate for the peak top and a relative or absolute tolerance Set relative tolerances by following the tolerance value with a e nearest to coordinate by setting the coordinate for the peak top and searching the nearest peak at any tolerance e by defining lower and upper bounds for a search interval The largest highest first or last peak in the search interval will be made the marker peak e by defining lower and upper bounds for a search interval as well as a minimum peak size The first or last peak in the search interval that exceeds the size limit will be made the marker peak The size limit is entered in the tolerance column It can be entered as an absolute value in which case peak area is used If you enter a relative value using a percent sign the peak area fraction is used Some versions of DAx let you limit the horizontal axis types that certain markers can be recognised in The On Axes item is used for this purpose It is possible to set up identification of a single marker with the same name throughout on different axis types simultaneously DAx will use only the database entries that match the axis type bein
34. external markers in related data box in the marker peak dialog box you can force DAx to try to find marker peaks in other related data sets in the current graphics window Use external markers in any data follows the same principle but the data sets may belong to different enslavement groups 197 These options also apply to locating reference peaks used to calculate relative coordinates and capacity factors By checking Do not use internal markers any peaks in the current data set will not be used as markers This option is only enabled if the use of external markers is enabled This option does not apply to locating reference peaks Chapter 18 7 Reference Peak Allocation Dialog Box If the menu option Analysis Reference peak is not present you need to use File Customise gt Extensions and uncheck Use Marker Peaks Using marker peaks is strongly preferred however This dialog box is used to indicate if and how a reference peak should be allocated automatically It is invoked using the Analysis Reference peak menu option Refer to the on line help for details about the reference peak allocation dialog box Chapter 18 8 Quantitative Calibration Configuration Dialog Box This dialog box is used to select the type of quantitative calibration to be used for a component or a range of components The dialog box also lets you set lower and upper limits for concentrations Concentrations outside the limits will still be calc
35. gradients 158 modifying peaks 124 modus 107 molecular weight axes 21 78 162 167 Molecular weight axis 163 moment See first peak moment mouse double click 63 moving average 85 86 94 108 147 moving data 69 Moving data 90 multiple detectors 97 multiple identifications 194 multiple qualifications 194 named peaks 140 naming peaks See peaks navigating text windows 42 new curve 89 100 new window 89 noise Point to point noise 76 112 RMS noise 76 112 values 135 normalisation peak 137 definition 28 normalisation peaks 128 Normalising peak widths 120 opening a measurement window 33 opening data files 48 automatic analysis 48 255 opening groups of data 52 operator 23 operator name 24 optimum Fourier filter 146 Optimum overlay 105 optimum scale 40 61 Order of data sets 90 Overlapping data 105 overlay 28 64 102 128 Overlay optimum 105 overwriting files 24 padlock 70 parameters 31 Patterns in meta analysis 222 peak list windows See Text Windows peak qualifications 192 peak quantifications 192 peak skim 131 peak slant 130 peak types 127 135 140 peaks 73 85 102 103 107 112 de activating peaks 127 136 adding 63 adding peaks 126 annotations 134 178 area limit 116 asymmetry 137 automatic shoulder peak detection 130 automatic thresholds 117 bleedthrough peaks 121 boundaries 74 columns in peak list windows 135 componen
36. in a sample by setting up quantitative calibrations The procedure involves analysing a known sample or a number of known samples and entering the names peak types peak annotations and concentrations of the known components in the peak list windows cf chapter Chapter 10 11 Names peak types and annotations are later assigned to unknown peaks based on one of a number of possible peak properties Possible qualifying properties are e peak top time first statistical peak moment peak top time offset peak begin time peak end time capacity factor peak top volume peak top molecular weight number averaged molecular weight for the peak weight averaged molecular weight for the peak M for the peak apparent mobility for the peak top effective mobility for the peak top calibrated value for the peak top peak top time minus marker peak top time peak top time divided by marker peak top time relative position index between two markers All identification parameters have an adjustable tolerance The tolerance is either absolute or relative 189 The components in an identification database do not need to be identified using a single qualifying parameter The qualifying parameter coordinate and tolerance can be adjusted for one or more components in the Identification Database list window chapter Chapter 18 9 1 Depending on the measurement data type of a data set some coordinates may not be available for use as a quali
37. in the next section about the requirements for calculating effective mobilities Chapter 21 5 Why are there no curves in my Mobility Axis Window Chances are that you have selected display of effective mobilities Either use the View Apparent Mobilities menu option to display apparent mobilities or make sure effective mobilities can be calculated This requires that e areference peak is specified e an effective mobility for the reference peak s component be entered as the peak s annotation If no effective mobility is entered for the reference peak the default reference mobility will be used Refer to chapter Chapter 16 5 for details Chapter 21 6 How do exclude the EOF peak from area calculations By making it inactive because only active peaks are included in total area and area fraction calculations Move the mouse cursor over the EOF peak top marker Click the right mouse button A 246 pop up menu appears Use the Active menu option to make the peak inactive Chapter 21 7 Why is the value for RMS noise often half the value for PP noise in the peak list window Whenever DAx searches for peaks it needs a noise level If no noise level has previously been determined it will calculate point to point noise If no RMS noise level has been calculated the value for the point to point noise level will be divided by two to get an estimate for the RMS noise level Use the data tag peaks menu chapter Chapter 8 3 to
38. it nor do you have to load all parameters from a file when you load it Dialog boxes will be displayed that let you choose which parameters to save or load Possible parameters are e Baseline construction parameters e Peak find parameters 30 Peak plot parameters and peak list window layout Filtering parameters HPLC gradient definition GPC calibration CE parameters Calibration Identification database Standard peak identification parameters Analysis Assay parameters Automatic Trace Calibration parameters Report name Only the file name of the report definition is stored NB If certain parameters are not present in memory when an analysis procedure is saved they will appear in the Analysis Procedure Save dialog box marked as lt clear gt If you check this item these parameters will be removed from memory cleared when the Analysis Procedure is loaded Measurement sequences cf Measurement Window manual can contain lines specifying that an analysis procedure with a given name must be loaded Chapter 3 5 DAx Command Line Parameters A number of command line parameters are available for DAx To use these parameters go to the Run option in the Start Menu and enter DAx32 lt command line options gt Command line parameters can also be used when DAx is run from a batch file for instance for the unsupervised analysis of vast numbers of files For a full explanation of all available command line par
39. line through the two points preceding the current point with the time coordinate of the current point Y4 Y Y2 2Y Y2 e intersection of the line through the two points following the current point with the time coordinate of the current point Y Ya J Y 42 2Y 41 Y 42 147 The lowest and the highest of these four values are determined Call them Yimin and Ymax Ymin and Ymax are expanded by the Drift Factor as follows Ymin Ymin DriftFactor Ymax Ymin Ymax Ymax DriftFactor Ymax Ymin A point is considered a spike if it does not lie between Yimin and Ymax If this is the case it is replaced by the average of the values preceding and following it 0 5 Y Y41 Note that the algorithm assumes all points are equidistant along the time axis DAx only supports those kinds of measurements 148 Chapter 12 The Waste Barrel DAx has a small window that depicts a chemical waste barrel This barrel is used to delete data sets A number of the most recently deleted data sets can later be restored The waste barrel can be used in a number of ways Drag a data tag chapter Chapter 1 to the waste barrel to delete the data set The data will be stored in the waste barrel from where they may later be recovered If the waste barrel restore option is not available you will be asked for confirmation The waste barrel has a limited capacity and may overflow losing the oldest data it contained Chap
40. menu option to display a dialog box that lets you determine where peaks should be located relative to each other Meta analysis is performed automatically after normal analysis Peak names used in meta analysis can be peak names assigned from normal analysis or peak names assigned manually To temporarily stop performing meta analysis use the Analysis Use meta analysis menu option 221 The meta analysis species with highest likelihood is shown in the data tag info dialog invoked using the Info option in the data tag menu cf chapter Chapter 8 2 A detailed meta analysis result window is invoked using the Peaks Show meta analysis menu option in the data tag popup menu23 The meta analysis results window that is displayed is live so updating the meta analysis entries or changing peak names in the data set immediately updates the result window The results are always sorted so that the entries with the highest likelihood are at the top Meta analysis results windows contain the following columns that the meta analysis list window does not e value value of the quantifier If the value is below the limit it is displayed in red e pattern value value of the pattern quantifier If the pattern is not matched the pattern value is displayed in red e likelihood likelihood of identification of the meta analysis entry e overall likelinood likelihood of identification of the species derived by combining the likelihoods of ide
41. no noise level has been determined yet DAx will calculate and use the point to point noise level If an RMS noise level has been calculated DAx will use this instead The noise levels are displayed in the peak list window chapter Chapter 10 11 Chapter 8 4 Data Tag sizing menu DAx contains a number of techniques to change the time and voltage coordinates of a data set There are five main categories see chapter Chapter 7 7 76 The data tag sizing menu looks like this Bring to Top Info wv Plot Use as Baseline Blank Measurement Peaks gt Sizing Controls Sizing Fourier Filtering gt Sizing Dialog 7 Undo sizma 7 W G iakin Bedo Sizing Data Table Undo All Sizing Copy to New Curve Copy to New Window Delete Data 1 Use the Sizing Sizing controls menu option to display a bounding rectangle encompassing the data set The sides or corners of the bounding rectangle can be dragged to size the data set or the entire bounding rectangle can be dragged to move the data set Press the Shift key while executing this menu option to show or hide bounding rectangles for all data sets in the graphics window 2 The bounding rectangle for the default data set which has a data tag that has a slightly thicker edge may conveniently be shown and hidden by double clicking in the graph and selecting Sizing controls from the popup menu that appears 3 Use the Undo and Redo menu options to c
42. not the peaks are considered missing Chapter 18 14 Analysis Logs Analysis logs are used to keep a record of peaks found in DAx analyses The log is highly configurable using the Analysis Analysis log Configure menu option e one or more data set level parameters can be logged These parameters are e analysis date amp time data set name data set file name number of peaks in data number of logged peaks in data total peak area percentage of total peak area of logged peaks for each peak none one or more of the normal peak list parameters chapter Chapter 10 11 can be logged e all peaks all named peaks or peaks with names in a user supplied list can be logged e you can choose to log only peaks with certain peak types e analysis logs can be written at the following times e whenever peaks are located in data e whenever peaks are located as part of automatic analysis cf chapter Chapter 7 1 2 1 Automatic analysis can be performed when data files are opened when files are scanned for cf chapter Chapter 4 2 and when measurements are performed cf chapter Chapter 4 1 223 e whenever peaks are calculated e manually using the data tag peak popup menu chapter Chapter 8 3 e analysis logs can be written in the following formats e as tab separated text easily imported into spreadsheets e as HTML viewable in web browsers Chapter 18 14 1 Analysis Analysis log Keep menu option This menu option switches betwe
43. option in the Calibration Set up dialog box invoked using the Calibration Calibration menu option e f Derive Calibration is checked in the Automatic Analysis setup when a file is opened with Automatic Analysis checked In this case a Calibration is derived from the data and stored as a local calibration It is also automatically copied to be the current main calibration The data tag popup menu chapter Chapter 8 2 contains options to list the local calibration draw the local calibration as a curve and remove the local calibration Chapter 17 10 Automatic Trace Calibrations In Fragment Length Analysis experiments a sizing standard is generally included as one of the traces The sizing standard is known to contain fragments with certain lengths In order to be able to set up a fragment size calibration based on the sizing standard the peaks corresponding to the standard fragment sizes must be identified Recognising the standard fragments can be somewhat difficult to do with traditional techniques such as looking for certain peaks in certain intervals especially when one or more of the following occurs e presence of spurious actual peaks in between standard fragments e presence of spurious peaks at the end of the measurement e overloading of the sizing standard trace from one of the other traces 183 Automatic Trace Calibrations offer a heuristic technique which can be used to reliably find the standard fragments
44. or press Ctrl V to insert the objects from the clipboard 238 Chapter 20 3 6 Customising the Report Window Use the View Attributes menu option to customise the appearance of the Report Editor Specifically this menu option is used to show or hide grid lines Use the View Centimeters menu option to display coordinates in centimeters with the origin at the top left of the page Use the View Inches menu option to display coordinates in inches Chapter 20 4 Report Objects Reports can contain the following items e Logos Logos are Windows metafiles or Device Independent Bitmap files They may contain any type of graphics Many drawing programs are able to create metafiles Device Independent Bitmaps are typically created by image editing software e Text objects These print a text Text objects are literal strings of text but they may also contain variables i e predefined texts between square brackets that will be replaced by the appropriate values when the report is printed The report editor contains a full list of these variable names for easy inclusion The list is also included below e Lines e Rectangles e Data Info This object will be filled with the information displayed in the data tag information dialog box chapter Chapter 8 10 e Data Plot A plot of the measurement data One report may be used to print several data sets at once Use the object index set in the upper left corner of the Report Editor
45. parameters will be used or you can click the blue underlined Construct button to configure baseline construction parameters 3 Check Subtract Baseline to automatically subtract the baseline from the data The baseline is removed after it is subtracted Since this means there is no way to determine the quality of the baseline this option should be used with caution If this option is checked but the baseline construction option is not the data will be marked as having had a baseline subtracted without any actual modification taking place This option should be used with caution and only on data that are known to have a straight baseline with signal value 0 4 Check the Find peaks box to detect peaks in the measurement The peaks are detected using the parameters that were last used to detect peaks or you can click the blue underlined Find peaks button to configure peak find parameters 5 Check the File box to automatically create a text file with peak analysis results The file name will be the same as the measurement s data file but with the file extension changed to pks for single data sets or pal pb1 etc for series of data sets The file is created using a peak list window 6 Check the Print box to print a peaks table Check the Only Named Peaks option to only print recognised peaks 50 7 If the Close Peak List Windows option is used peak list windows that have been created in order to print or file peak analysis results
46. qualification coordinate is compatible with the horizontal coordinate of the graphic window both must be a time coordinate or both must be calibrated values Chapter 7 3 18 View Colour view The View Colour view menu option displays the contents of the graphic window as a coloured map Colour views are convenient when large numbers of data sets need to be compared Refer to Chapter 7 8 for details Not all versions of DAx contain this functionality Chapter 7 3 19 View Data set base calls View Consensus base calls View Allele base calls These commands can be used to display or hide various nucleotide strings Chapter 7 3 20 View Remove Highlight Certain actions such as locating peaks from within a peak list window chapter Chapter 10 11 3 will add a highlighted area to a graphic window Use the View Remove Highlight menu option to remove the highlighted area Chapter 7 4 Data Set Graphics Window Data Menu The Data menu is discussed in detail in chapter Chapter 9 62 Chapter 7 5 Mouse Tracking Behaviour When the mouse cursor is moved across a graphics window its location is tracked in the coordinate display to the right of the toolbar The cursor tracks the current default data set with two exceptions e if the cursor is placed on a data curve that curve is tracked and its data tag is depressed6 e if data are displayed stacked the display can be considered to consist of bands of stacked data The mouse c
47. repeated as many times as is required to output the entire list It is also possible to define a peak list item on the first page of a report and another peak list item e g a much larger one on the second page In this case the list will flow from the first into the second item If the entire list still does not fit the second page will be repeated To prevent continuation of long lists on the next page in the report editor right click the item and choose the Config table or Config peak list item as applicable A dialog box will be displayed In it check the If table runs long do not continue on next page item Chapter 20 4 2 Text Object Variables Text objects are literal strings of text but they may also contain variables i e predefined texts between square brackets that will be replaced by the appropriate values when the report is printed The report editor contains a full list of these variable names for easy inclusion Available variables are page page number print date date and time the report is printed header n n 1 5 5 predefined header strings Use File Customise gt Reports to enter these headers file path fully qualified disk file name of the data file name disk file name of the data disk file date date and time the disk file was last updated data set name name of the data set trace type trace type of the data set operator name operator name of the operator who measured the data measurement
48. run e g after a sequence was interrupted to run a priority sample no new data list will be started You may choose to include Extra Field names in the data list in addition to the extra field values This is useful if your measurements do not all have the same extra fields NB Once a data list entry has been created the extra field name has either been included or not To add or remove extra field names at a later time use Data List Assert list Chapter 3 2 Window Types in DAx The main types of windows that are used in DAx are listed below e Measurement window There can be only one measurement window This is the window that is used to perform data acquisition by letting you start and stop measurements and by letting you set up measurement sequences The Measurement Window is described in detail in the Measurement Window manual e Data Set Graphics windows chapter Chapter 7 2 Data sets are presented in these windows The graphics window is the owner of its data various alternative representations may be created in different windows but when the graphics window is closed all windows containing alternative representations of the data will also be closed e Data Value List window These windows contain a list of data values for a data set Each data set has its own data value list window A number of common text window operations are available for data value list windows such as saving as text files or copying to the clipb
49. sheet Data can also be added to the global 205 comparison sheet when they are analysed when they are loaded from files by checking the Add to Global Sheet box in the Automatic Analysis set up dialog box The Comparison Sheet has the usual text window menu options Additionally when the right mouse button is clicked in the sheet a popup menu appears This popup menu offers the ability to locate or highlight a peak in a graphic window press the Shift key to hide all other data sets or to highlight all selected peaks The Delete peak menu option deletes the peak that was clicked on The peak is removed from the source data set not just from the comparison sheet For global comparison sheets there is also a menu option Sort data sets used to sort the data sets in the comparison sheet using one or more criteria Chapter 18 11 1 Method for Grouping Peaks Together When a qualifying parameter is used to group peaks together the following method is used e in each data set the first peak that is not in a group yet is found e of these first peaks the peak with the lowest value for the qualifying parameter is found e anew group is started with that peak as the first member e peaks from other data sets are added to the group Peaks are added only if the qualifying parameter is within the specified tolerance If more than one peak is within tolerance the peaks with the closest matching qualifying parameters will be matched e when
50. text If there are three lines the middle line is always vertical An arrow point at the end of the last line can be displayed or hidden using a button in the label editing dialog box item 9 above To move the arrow click and drag either one of the ends of any of the arrow s lines This will also display the label editing dialog box Whenever the label editing dialog box is visible the start and end of the arrow s lines will be marked by small rectangles except the arrow point if it is used 152 c test5 label dal SSS Example of the use of labels Labels with one two and three lines were used 153 Chapter 14 DAx HPLC GC HPLC Gradient amp GC Temperature Programme Extensions If the menu option HPLC GC is not present you need to use File Customise gt Extensions and check HPLC Options or GC Options Not all versions of DAx contain these options The HPLC GC version of DAx has a number of extra options that make it possible to correct measurement data for signal gradients To be able to do this a number of parameters about the applied gradient need to be established Other than correcting for signal gradients DAx also has the capability to plot a curve depicting either HPLC gradient percentages or GC temperature programmes Gradient percentages or programming temperatures must be entered as parameters to plot these curves The type of gradient either HPLC Solvent composition or GC Temperature p
51. the Peaks Construct baselines and Peaks Find peaks menu options Baseline construction and peak find parameter may also be read from an Analysis Procedure file using the File Load analysis procedure menu option Chapter 10 6 Peaks Fit Gaussians This command attempts to fit Gaussian curves to all peaks in the data The number of Gaussian curves is equal to the number of peaks present in the data no new peaks will be detected amp Fit peaks with Gaussians i 2 x El C win SETUPYDAx test3 dax J Limit fitting to zoomed window 4 STAND 1 Da1 D STAND 1 Da1 Mipi n Fit Gaussians 2l Fit asymmetric Gaussians Fit Fraser Suzuki skewed Gauss Fit options Separate by baseline Do not move peak tops Treat shoulders as regular peaks m Peak marking upon completion Select all fka Deselect pee a Cancel Help V Plot total curve 1 The fit can be limited to the visible part of the horizontal axis by checking Limit fitting to zoomed window 2 Various curve types can be fitted to the peaks Refer to chapter Chapter 9 15 for details on the curve types 3 Various options can be activated to help the fitting process Separate by baseline causes the fitting process to split the peaks into groups that are baseline separated if this option is not checked groups of peaks that are 123 close together are fitted If Do not move peak tops is checked only the peaks height and w
52. the first data set that is overlaid as target area The absolute value of peak areas is used Scaling will also be applied in such a way that the smallest signal value encountered will be 0 Movements made using the Data Overlay dialog box can be undone like all other sizing operations chapter Chapter 7 7 Use the Undo Size menu option in the data tag size menu chapter Chapter 8 4 Baseline data sets denoted with a yellow 8 cannot be selected This is because they are automatically sized along with the data set that owns them in order to keep the relationship between peak data set and baseline data set intact 104 Chapter 9 17 Data Stack data The Stack Data dialog box is used to move data sets that overlap each other in such a way that they no longer overlap This can make comparing the data sets much easier particularly if many peaks are present The Percentage of vertical axis used for data stack item determines how much room is left open between the data sets If a value of 50 is entered half the vertical axis will be covered by data points Lower values will leave more room open between data sets Checking the First Data Set at Top box causes the data sets to be arranged so that the first data set will be highest with consecutive data sets being displayed lower down First Data Set at Bottom is the reverse Data sets can be stacked individually or by enslavement group DAx 7 1 PP 17 0
53. the peak begin or end coordinate can be adjusted to coincide with the end or begin of the previous or next peak To do this move the mouse cursor over the peak begin or end marker and click the right mouse button A pop up menu appears Select Connect to previous or Connect to next as appropriate Peaks can also be combined i e two peaks can be combined into a single new peak Click the right mouse button on the peak begin or end marker again and select Combine with previous or Combine with next as appropriate Chapter 10 11 Peak List Windows When peaks have been detected for a data set a text window may be created listing the peaks Such a text window is called a peak list window Peak list windows are created using the Peak list menu option in the data tag peak menu There is also a peak list button at the left side of the data tag for data sets that contain peaks click the button to create a peak list window Peak list windows have the normal functionality of text windows see chapter Chapter 1 132 Additional functionality is listed below Chapter 10 11 1 Peak Highlighting When the mouse cursor is moved over a peak marker begin top or end in a Data Set Graphics window the corresponding peak is highlighted in the peak list window When part of a graphics window is zoomed in on the peaks that lie within the window will be highlighted in all peak list windows associated with the graphics window The total area of t
54. to investigate crosstalk between traces in DNA analyses Chapter 9 13 Data Sort If the menu option Data Sort is not present you need to use File Customise gt Data and check Data Sorting This command invokes a dialog box that allows data sets to be sorted by their signal value or their absolute signal value Sorting data can be useful to get an idea of the distribution of signal values Typically a baseline will be subtracted from the data first 99 Chapter 9 14 Data De Convolute data If the menu option Data De Convolute Data is not present you need to use File Customise gt Data and check Date de convolution This command invokes a dialog box that allows data sets to be deconvoluted or convoluted by either a theoretical curve Gaussian or triangular or by another data set De convolution is achieved using a Fast Fourier Transform 9 technique Note that deconvolution is very sensitive to signal noise so your data should be carefully filtered before attempting to deconvolute it The available theoretical curves are the same as the ones used to generate data described in the next chapter Each of these curves can have its top at a selectable time coordinate The curves are scaled in such a way that the area below them equals unity If you do not use a theoretical curve but use a data set to perform the de convolution this data set will be disabled in the Data Selection list box at left in other words yo
55. type GPC or of type Generic To use this option for a certain extension grey check its check box lf Local Calibrations are enabled each data set can store and use its own calibration This makes it possible to re analyse data each time using the calibration that pertains to that specific data set Qualification i e peak recognition can be customised to either automatically allocate a full set of marker peaks or just a single reference peak Refer to chapters Chapter 18 7 and Chapter 18 6 for details A preferred horizontal axis unit can be chosen for use when a specific unit is not available Choose the unit that most of your data will have If the Store parameters with data box is checked filtering baseline construction peak finding identification and assay parameters are stored in each data set they are applied to They 15 can be retrieved later on this ensures that all analyses can be fully reproduced Each of Base call amp allele match sheets reports and data lists can be activated or deactivated DAx uses an Analysis Procedure file to save filter parameters baseline construction and peak find parameters GC HPLC CE GPC parameters as well as qualification and quantification parameters The enable separate parameter files item allows you to also load and save GC HPLC CE GPC and qualification and quantification parameters separately Chapter 3 1 2 File Customise gt StartUp and Measurement d
56. type of printer and paper orientation selected NB it is possible to use a report definition created for one type of printer to print a report on a different type of printer Chapter 20 1 The Report Menu The Report menu contains items to start a New Report Load an existing report definition and Save the current report definition Report Close report removes the report definition from memory Report Edit report opens the report editor to let you change the report See the next sections for details Report Preview report previews a report for the current data Report Print report prints a report for the current data 235 Chapter 20 2 Editing Reports The FontBar The Report Editor displays a FontBar toolbar The FontBar is used to 1 set an object index There will often a ee be more than one object of the same 2 6 type in a report definition For w al instance there may be two data plot xl 2 objects By setting the object index of the first of these to 1 and the object index of the second to 2 two different Ale Af data plots will be placed in the report B ul If only one data plot is available the a 2 data plot object with index 2 will be 5 left blank Al 7 0 2 enter text for text objects Click the down arrow in the text edit combo isisisi i box to see a list of predefined texts wi SiFigigia Texts between square brackets will be replaced with information when the report
57. window menu is a standard window menu If you execute the Window Close all menu option you will be asked to confirm closing all windows that contain unsaved information Data will not be sent to the waste barrel chapter Chapter 11 If the Measurement window is open it will not be closed Similarly Window Minimise all will minimise all windows except the measurement window e mouse double click commands e data set sizing techniques Chapter 7 1 Data Set Graphics Window File menu The file menu contains commands to start a new measurement open and save files print graphics close windows and close DAx Chapter 7 1 1 File New This command opens a Measurement window Chapter 7 1 2 File Open This command reads data sets from a file Several file formats are supported Older versions of DAx saved data sets that belong together see chapter Chapter 3 3 9 using the same file name with different extensions For this reason when DAx opens a data file it will always open all files that have the same name together The File Open menu commands enforce this by always automatically 48 selecting all files that have the same name in the directory list as soon as one of them is selected Files can either be read into a new graphics window or be added to the current graphics window by checking the Add to Window box in the File Open dialog After loading the data sets can immediately be sorted by checking the Sorted box
58. window will be created containing a scaled preview of the printed report The previews are live that is if the data change the preview changes Chapter 20 6 1 File Write report From the preview window reports can be written to disk in two formats e as an RIF file which can be read by programs such as Microsoft Word and Microsoft PowerPoint e as a report view which is a file that contains all the information needed to reproduce the report the data as well as the report definition Use the File Write report menu item to write reports Select which type of file to create using the List Files of Type item in the dialog box Not all versions of DAx have the ability to write reports Chapter 20 6 2 Report Optimise scale When this menu option is checked all data plot items will have an optimised scale Normally data plot items use the same zoom setting that the original graphics window uses Optimise Scale negates this It also means that if there are several data plot items each with their own data the scales will be individually optimised Chapter 20 6 3 Report Single graph Separate graphs Graph per set Single graph causes all data sets from a graphic window to be plotted in a single data plot item on the report Separate graphs causes each data set possibly including its baseline to have a separate data plot item If the report definition does not contain 243 enough data plot items the enti
59. you press the Alt key while zooming a popup menu will be displayed that offers the option of selecting all data sets whose curves are present anywhere in the zoomed box or of deleting all peaks whose tops are contained in the zooming rectangle You can also add a user defined bin with limits equal to the zoom box using this technique Note that if peaks have been detected for any data set in the graph and the peaks are being displayed in a peak list window chapter Chapter 10 11 when you zoom in on part of the graph the peaks that lie within the zoomed graph window will be highlighted in the peak list window unless you press the Shift key while zooming in Peaks are marked using triangular markers When the mouse cursor is placed on a peak marker if a peak list window is open 3 If data are stacked only curves within the zoomed window are included in the scaling process unless the zoomed window is entirely within a single stacked range in that case all curves in that range are included whether they are contained in the zoomed window or not 47 the peak will be brought into view and highlighted unless you press the Shift key The same is true for comparison and binning sheets The following subjects will be discussed in this chapter File menu Edit menu View menu The Data menu is discussed in chapter Chapter 8 The data menu contains the options that change the data such as filtering and baseline construction e The
60. 0 10 4 e mark a normalisation peak cf chapter Chapter 10 10 3 140 e set the marker peaks for all highlighted peaks cf chapter Chapter 3 3 16 e activate or deactivate peaks cf chapter Chapter 10 10 2 141 Chapter 11 Filtering Techniques Chapter 11 1 Fourier Filtering To be allowed to use Fourier techniques you need to use File Customise gt Data and check Fourier Transforms Fourier transforms are used to convert data from the time domain to the frequency domain Any N data points that constitute a data set in the time domain can be converted to N data points that represent the same data set in the frequency domain without losing any information Since noise is by definition a change in the signal that occurs faster than any real change in the signal converting to the frequency domain may give an optimum way of removing noise simply by discarding the frequency data at high frequencies 4 this is called a low pass filter By analogy signal drift occurs at frequencies lower than the real signal it may be removed by discarding the frequency data at low frequencies high pass filter DAx has the ability to calculate frequency domain data for each data set using the tag menu see chapter Chapter 8 2 When this has been done two arrow controls are placed above the graph These controls can be dragged to change the interval of frequency data that is considered part of the real data While the controls are d
61. 1 01 16 18 59PM DAx 7 1 PP 17 01 01 16 19 11PM Y Y 000 ToO da aa XL mhs Asi 4 sta 4 on SOUE os an 000 0 VERE sks EE aka EE SE YE eV T I T T T I T T I 0 50 100 150 0 50 100 Time min Time min Overlapping data sets before and after stacking Chapter 9 18 Data Best overlay If the menu option Data Best overlay is not present you need to use File Customise gt Data and check Best overlays Not all versions of DAx contain this option The Best Overlay option is deprecated Refer to the help file for a full explanation of its functionality 105 106 Chapter 10 Baselines and Peaks Data Set Graphics Window Peak Menu This chapter discusses the commands in DAx that construct baselines subtract baselines from signals and detect and calculate peaks Editing capabilities to manually adjust both baselines and peaks will also be discussed Chapter 10 1 Peaks Construct baselines This command invokes a dialog box that is used to construct baselines for one or more data sets The algorithms DAx uses to construct baselines either construct a horizontal constant value baseline where the baseline value does not change in time or a drifting baseline DAx has two horizontal baseline algorithms e Modus or mode baselines The baseline is thought to lie at the signal value that is most likely to occur in the data set e Average below average baselines are constructed by determining the average sign
62. 12 3 User defined bins 217 Chapter 18 12 4 Displaying curve samples 218 Chapter 18 12 5 Binning maps 218 Chapter 18 12 6 Binning sheet menu options 219 Chapter 18 12 7 Method for Grouping Peaks Together 220 Chapter 18 13 Meta analysis 220 Chapter 18 13 1 Meta analysis patterns 222 Chapter 18 14 Analysis Logs 223 Chapter 18 14 1 Analysis Analysis log Keep menu option 224 Chapter 18 14 2 Analysis Analysis log Configure menu option 224 Chapter 18 14 3 Analysis Analysis log View menu option 224 Chapter 18 15 Analysis Assays 224 Chapter 18 15 1 Assay Weighting 225 Chapter 18 15 2 Baseline Parameters 226 Chapter 18 15 3 Peak Parameters 226 Chapter 18 15 4 Required peaks 227 Chapter 18 15 5 Unwanted peaks 227 Chapter 18 15 6 Calibration Quality 228 Chapter 18 15 7 Analysis Edit assay parameters 228 Chapter 18 15 8 Analysis Load assay parameters Analysis Save assay parameters Analysis Stop quality assays 229 Chapter 18 15 9 Displaying an Analysis Assay 229 Chapter 19 Data Lists 231 Chapter 19 1 Data List New list Data List Load list Data List Save list Data List Close list menu options 232 Chapter 19 2 Data List Add data menu option 232 Chapter 19 3 Data List Assert data menu option 232 Chapter 19 4 Data List Display list Data List Display statistics232 Chapter 19 5 Data List Find data 233 Chapter 19 6 Edit Sort list menu option 233 Chapter 19 7 Edit Find Edit Find next Edit Select menu opti
63. 16 1 Calculating Mobilities step by step To be able to calculate apparent mobilities DAx needs to have a number of parameters on the capillary being used Use the CE Calibrate menu option to enter these parameters or use the CE Load calibration menu option to load parameters from disk To be able to calculate effective mobilities DAx also needs to be able to calculate the electro osmotic flow This means that the effective mobility for one peak needs to be known This peak is always the reference peak Use the following steps e construct a baseline for your electropherogram using the Peaks Construct baseline menu option chapter Chapter 10 1 e find the peaks in your electropherogram using the Peaks Find peaks menu option chapter Chapter 10 4 e mark the reference peak by placing the mouse cursor over its peak top then clicking the right mouse button The peak popup dialog box appears Check the Reference peak box You may also enter the effective mobility for the reference peak in the annotation field in this dialog box Alternatively e create a peak list window using the data tag peak menu chapter Chapter 8 3 169 e enter the effective mobility for the reference peak in the annotations column in the peak list window chapter Chapter 10 11 4 If the annotation item is not being displayed in the peak list use the View Configure menu option to add it to the displayed columns If no effective mobility is entered for the
64. 17 8 3 When the calibration changes 182 Chapter 17 9 Local Calibrations 182 Chapter 17 10 Automatic Trace Calibrations 183 Chapter 17 10 1 Setting up the Automatic Trace Calibration parameters 184 Chapter 18 DAx Qualification amp Quantification Identification Database Extensions 189 Chapter 18 1 Creating an Identification Database 190 Chapter 18 2 Analysis Load database Analysis Save database Analysis Stop analysis menu options 191 Chapter 18 3 Analysis Qualify peaks 192 Chapter 18 4 Identification Database Edit Dialog Box 192 Chapter 18 5 Multiplicity 194 Chapter 18 6 Marker amp Reference Peak Allocation Dialog Box _ 194 Chapter 18 6 1 Using Existing Peaks 197 Chapter 18 6 2 Using external marker peaks 197 Chapter 18 7 Reference Peak Allocation Dialog Box 198 Chapter 18 8 Quantitative Calibration Configuration Dialog Box 198 Chapter 18 8 1 Dilutions 199 Chapter 18 8 2 Injection Volumes 199 Chapter 18 9 Identification Database Windows 200 Chapter 18 9 1 Identification Database List Window 200 Chapter 18 9 2 Quantitative Calibration List Window 201 Chapter 18 9 3 Quantitative Calibration Graph Window 202 Chapter 18 9 4 When the database changes 203 Chapter 18 10 Using Internal Standards 203 Chapter 18 11 Comparison Sheets 204 Chapter 18 11 1 Method for Grouping Peaks Together 206 Chapter 18 12 Binning Sheets 207 Chapter 18 12 1 The Binning Sheet dialog box 208 Chapter 18 12 2 Binning sheet popup menu options 215 Chapter 18
65. 2 1 File Print Chapter 5 2 2 Edit Copy graph Chapter 5 2 3 Edit Create metafile Chapter 5 2 4 View Attributes Chapter 5 2 5 View Select curves Chapter 5 2 6 View Edit scale Chapter 5 2 7 View Zoom in Chapter 5 2 8 View Zoom out Chapter 5 2 9 View Reset scale Chapter 6 DAx Text Windows Chapter 6 1 1 Selecting Lines Chapter 6 1 2 Editing Text Window Contents Chapter 6 1 3 Deleting Lines Chapter 6 1 4 Pop up Menus Chapter 6 1 5 Navigating Text Windows Chapter 6 2 Text Window Menu Commands Chapter 6 2 1 File Export File Export selected Chapter 6 2 2 File Print Chapter 6 2 3 View Configure Chapter 6 2 4 View Fit columns View Shrink amp fit columns Chapter 6 2 5 View Text size Chapter 6 2 6 Edit Select all Chapter 6 2 7 Edit Copy Edit Copy using Tabs Chapter 7 Data Set Graphics Windows Chapter 7 1 Data Set Graphics Window File menu Chapter 7 1 1 File New Chapter 7 1 2 File Open Chapter 7 1 2 1 Automatic Analysis Chapter 7 1 2 2 Dropping Files Chapter 7 1 2 3 Opening Groups Chapter 7 1 3 File Save File Save as Chapter 7 1 4 File Close Chapter 7 1 5 File Scanning Chapter 7 1 6 File Preview report Chapter 7 1 7 File Print report 54 Chapter 7 1 8 File Print 54 Chapter 7 1 9 File Printer setup 55 Chapter 7 1 10 File Load analysis procedure File Save analysis procedure 56 Chapter 7 1 11 File View message log 56 Chapter 7 1 12 File Operator 56 Chapter 7 1 13 Fil
66. 4 138 198 definition 28 reference peaks 128 relationships 89 relative origin 63 remark labels 152 remark tacks 83 151 removing analysis results 124 Removing baselines and peaks 75 removing peaks 133 removing peaks in a zoomed window 47 report preview 54 print 54 report views 243 reports 51 235 changing object type 237 copying objects 238 customising 239 editing multiple objects 238 headers 19 logos 239 moving objects 237 multipage lists 241 objects 239 removing objects 238 sizing objects 237 text variables 241 required peaks 227 reset scale 40 61 resolution 137 RMS noise See noise RTF files 243 saving data 52 Savitzky Golay 85 94 100 147 scale edit scale 39 60 reset scale 40 61 Scanning for files 33 SCF files 30 SEG 15 161 seconds 61 select all button 57 91 92 98 116 calibration graph selecting a measurement driver 16 selecting curves 39 selecting curves in a zoomed window 47 selecting data sets 57 58 selecting data sets in a zoomed window 47 selecting extensions 15 selecting lines in text windows 41 selecting text 42 45 selecting text window columns 43 setting marker peaks 127 135 setting scale 60 setting up a printer 55 setting up automatic analysis 49 shape overlay 103 shoulder peak area calculations 131 shoulder peaks 126 129 130 131 show boundaries 113 simplifying DAx 15 Size Exclusion Chromatography 161 sizing 24 59
67. 4 Data Set Graphics Window Data Menu 62 Chapter 7 5 Mouse Tracking Behaviour 63 Chapter 7 6 Mouse Double Click Commands 63 Chapter 7 7 Data Set Sizing Techniques 64 Chapter 7 8 Colour View Windows 65 Chapter 7 8 1 View Extra space Chapter 7 8 2 View Use trace colours Chapter 7 8 3 View Darker Chapter 7 8 4 View Halftones Chapter 7 8 5 View Subtract baselines Chapter 7 8 6 View Separate lanes Chapter 7 8 7 View Group per lane Chapter 7 8 8 View Calibrated axis Chapter 7 8 9 View Mark peaks Chapter 7 8 10 View Mark bins Do not mark bins Chapter 7 8 11 View Mark bins Mark above Chapter 7 8 12 View Mark bins Mark above and inside Chapter 7 8 13 Colour View Mouse Tracking Behaviour Chapter 8 DAx Data Set Tags Chapter 8 1 Selecting Data Tags Chapter 8 2 Data Tag menu Chapter 8 3 Data Tag peaks menu Chapter 8 3 1 Showing Boundary Conditions Chapter 8 3 2 Fixating Peak Names and Concentrations Chapter 8 3 3 Recalculating Peak Data Chapter 8 3 4 Clearing Analysis Results Chapter 8 3 5 Calculating RMS Noise Levels Chapter 8 4 Data Tag sizing menu Chapter 8 5 The Data Set Sizing Dialog Box Chapter 8 6 Data Tag splines menu Chapter 8 7 Modifying Splines Chapter 8 8 Data Tag Gradient Menu Chapter 8 9 Data Tag Tack Menu Chapter 8 10 Data Set Information Dialog Chapter 8 11 The Data Set Action List Dialog Box Chapter 8 12 The Add Remarks to Log File Dialog Chapter 9 Data Set Graphics Window Data men
68. 46 72 106 9 285 7 5 2 107 8 107 8 755 72 107 8 1418 6 6 2 109 7 109 8 95 834 109 5 172 53 7 2 116 7 116 7 236 5 116 7 373 1 8 2 117 6 117 6 91 203 117 6 71 429 9 1 119 7 a 119 7 598 54 10 1 123 1 123 1 890 35 11 1 125 0 125 0 98 35 12 1 126 3 a 126 3 96 257 13 1 130 6 130 6 108 46 14 1 138 4 eo 53 138 4 517 73 15 1 139 4 139 4 191 75 16 1 141 0 E 141 0 300 04 17 1 141 9 a 141 9 273 81 18 2 142 9 143 0 73 933 142 9 829 84 19 1 144 0 5 5 144 0 294 11 20 2 148 1 148 1 66 814 148 1 1571 5 Data can be added to the binning sheet when they are analysed as they are loaded from files by checking the Add to Binning Sheet 207 box in the Automatic Analysis set up dialog box In this way binning sheets containing data from hundreds of analyses can be created Chapter 18 12 1 The Binning Sheet dialog box The binning sheet dialog box invoked using the Analysis Binning sheet menu option has a parameter selection area at the left side A binning sheet uses over 10 sets of parameters which can be easily navigated here either by using the Next and Prev buttons or by clicking on any of the parameter set names Data set selection The first set of parameters is concerned with selecting the data sets that should be included in the binning sheet E Binning Sheet Setup 21x Setup parameters Select the data sets that are to be included in the binning sheet Data set selection User def
69. 6 File Save as 52 File Scanning Scan for files 33 File Scanning Scan setup 33 File Target printer 235 File View message log 56 File Write report 243 GPC Axis conversion 163 GPC Calibrate 164 GPC Calibration Curve 166 GPC Calibration List 166 GPC Load calibration 162 GPC Mw axis 162 GPC Save calibration 162 GPC Stop calibration 162 GPC Track Mw 162 GPC Volume axis 162 HPLC GC Gradient Corrected Axis 158 HPLC GC Load gradient 156 HPLC GC Save gradient 156 HPLC GC Show Gradient Nodes 157 HPLC Subtract Gradient 157 Peaks Baselines amp peaks 123 Peaks Clear analyses 124 Peaks Construct baselines 107 Peaks Find peaks 112 Peaks Fit Gaussians 123 Peaks Fixate analyses 124 Peaks Recalculate 124 Peaks Subtract baselines 110 Peaks Unfixate analyses 124 Report Close Report 235 Report Edit Report 235 Report Graph per set 243 Report Load Report 235 Report New Report 235 Report Optimise Scale 243 Report Preview 243 Report Preview Report 235 Report Print Report 235 Report Save Report 235 Report Separate graphs 243 Report Single graph 243 View Allele base calls 62 View Attributes 38 59 239 242 244 View Binning map 218 View Calibrated axis 66 View Centimeters 239 244 View Colour view 62 65 View Consensus base calls 62 View Cur
70. 64 total concentration 79 Trace Analysis 120 Trace files 183 tracking 61 tracking calibrated values 177 tracking mobility 171 tracking molecular weight 162 trend removing 247 undo 82 unfixating analysis results 124 unrecognised peaks 227 unwanted peaks 227 user defined bins 47 62 66 197 209 210 211 214 216 217 220 221 volume axes 21 78 162 waste barrel 27 48 54 70 149 destroying contents 150 emptying 150 moving 150 popup menu 149 restoring items 149 waste barrel cursor 149 Weight to concentration conversion 163 writing reports 243 yanking 93 yellow B 70 zoom 37 40 60
71. ARVARD__1_OT_ FAM HARVARD__1_OT_ FAM HARVARD__1_OT_ IC HARVARD__1_OT_ IC a in0008BoooooOm hear Rox J I Deselect OK Cancel Help When the dialog box is closed the selected data sets have a green check mark on the right side of their data tag Click on one of these check marks to invoke the data tag popup menu Some of the items in the menu can be applied to all selected data tags when such a menu item is highlighted all selected data tags will be depressed It is possible to move or copy all selected data sets To do this click on a check mark on a data tag and drag the selected data 57 tags to a different window pressing the Ctrl key to copy instead of move 9 Chapter 7 2 3 Edit Invert selection This command selects all unselected data sets and deselects all selected sets Chapter 7 2 4 Edit Copy graph This command copies the contents of the graph window to the clipboard as a metafile picture This picture may then be imported into a drawing program or word processor using the Edit Paste command in that program You will be given the option of including data set names and curve samples See the examples below DAx 1 0 pp 9 1 96 16 48 21 DAx 1 0 pp 9 1 96 16 48 29 VOORK 9 TXT a VOORK 9 1XT 4 J VOORK 9 TXT T T T T T T i i i 400 600 800 is a a time s time s Examples of metafile pictures without left and with right
72. Chapter 3 3 7 in order for baseline subtraction to be possible on a given data set It is possible to subtract a baseline from a data set even if no baseline is present The data will be marked as having had a baseline subtracted without any actual modification taking place This option should be used with extreme caution and only on data that are known to have a straight baseline with signal value 0 110 Note that when a new baseline is constructed for a data set that previously had a baseline subtracted the baseline subtracted flag is cleared because otherwise the newly constructed baseline would be ignored All signals 0 except peaks removes all fluctuations in the data that have not been recognised as peaks by making all data that do not lie inside a peak have zero signal value This can be a very powerful feature but it also removes any small fluctuations in the data that might have been recognised as peaks if the peak find parameters were adapted slightly Chapter 10 3 Modifying Baselines Once a baseline has been constructed chapter Chapter 10 1 it may be necessary to adjust it manually This is done using splines chapters Chapter 3 3 16 Chapter 8 7 The following steps should be taken 1 add a spline to the baseline curve using either the data tag spline menu chapter Chapter 8 6 for the baseline s data set or by clicking the add spline button on the toolbar The add spline button on the toolbar adds a s
73. DAX Data Acquisition and Analysis Software 2014 PP van Mierlo User s Manual ts DAx Data Acquisition amp Analysis PP 1 B laix File Edit wiew Data Peaks HPLC GC GPC CE Calibration Analysis Report Datalist Window Help plelmlal Efa a a al A pea eas aa D al 285 83 0 2369 V Mw lt na gt AppMob 338 6 CAL lt na gt E c win setup dax test3 dax C win setup dax test3 dax eS Ea STAND 1 Da1 B STAND 1 Da1 4 heptanal 0 39 216 20 221 70 227 40 261 60 2 0 methanal 0 275 0 100 0 400 297 80 ethanal 0 365 0 000 0 400 300 50 propanal 0 568 H 0 000 0 400 303 20 E A butanal 0 360 0 000 0 400 306 10 f pentanal 0 246 0 000 0 400 308 20 f hexanal 0 189 0 000 0 400 311 20 y K heptanal BATT 335 40 E k methanole 5 289 H 0 300 0 510 337 90 h k ethanole 4512 340 80 A i propanole 5 028 342 90 1 butanole 6 764 345 60 i j bentanole 6 353 www dax nl Contents Chapter 1 Introduction 11 Chapter 2 Installation 13 Chapter 2 1 System Requirements 13 Chapter 2 2 Installing the Software 13 Chapter 2 2 1 Drivers 14 Chapter 3 Customising Window Types Terminology15 Chapter 3 1 Customising DAx 15 Chapter 3 1 1 File Customise gt Extensions 15 Chapter 3 1 2 File Customise gt StartUp and Measurement driver options16 Chapter 3 1 3 File Customise gt GLP 16 Chapter 3 1 4 File Customise gt Baselines 17 Chapter 3 1 5 File Customise gt Finding
74. DAx contain this option The GPC version of DAx has a number of extra options that make it possible to use the program to determine molecular weights from chromatograms To be able to do this a calibration needs to be established The calibration consists of a list of Elution Volume Molecular Weight data pairs Chapter 15 1 Creating a GPC calibration To create a calibration 1 measure one or more samples with known molecular weight components Make sure the flow rate is correctly entered using either the measurement tag menu cf Measurement Window manual before or during the measurement or the data tag menu chapter Chapter 8 2 when the measurement is displayed in a graphics window 2 construct a baseline refer to chapter Chapter 10 1 for details 3 detect peaks refer to the chapter Chapter 10 4 for details 4 use the data tag peak menu to display a peak list chapter Chapter 10 11 for each of the measurements you want to include in the calibration 5 enter the known molecular weights in the annotation column in the peak list chapter Chapter 10 11 4 If the annotation item is not being displayed in the peak list use the View Configure menu option to add it to the displayed columns Annotations may also be entered by right clicking peak top markers to invoke the peak popup dialog box 6 invoke the GPC Calibration menu option discussed in detail in chapter Chapter 15 5 As an alternative to entering molecular we
75. Deselect I Tack removed peaks 5 lt Prev Next gt Close Help 1 Some bleed through peaks can be recognised by the presence of a much higher entirely encompassing peak in alternate data that have been measured simultaneously with the data being inspected Set a lower limit for the ratio of signal and signal of suspected bleed through peak to make sure that only peaks for which a sufficiently high encompassing peak exists are removed 2 Some bleed through peaks can be recognised by being highly asymmetric Use Right width Left width to recognise peaks with a sharp front end Peak widths are measured at 10 of peak height 3 Use Left width Right width to recognise peaks with a sharp front end Peak widths are measured at 10 of peak height 4 Asymmetric bleed through peaks often show high signal values in alternate data that have been measured simultaneously with the data being inspected If baselines have been constructed for the alternate data you may require that the alternate data have a value that is at least a number of times as high as the peak top height of the suspected bleed through peak 5 Peaks that were removed as bleed through peaks can be marked using tacks 122 Chapter 10 5 Peaks Baselines amp peaks This command constructs baselines for all data sets in the window then finds peaks in them Baseline construction and peak find parameters are used as they were last entered in
76. Down to move an entire page up or down e Ctrl Home to go to the top of the data e Ctrl End to go to the bottom of the data e F2 to use cursor keys inside the edit control or combo box Hit the F2 key again to once again be able to use the cursor keys to navigate between cells If the Shift key is pressed with the Cursor Up Down Page Up Down or Ctrl Home or Ctrl End keys lines will be selected 42 Chapter 6 2 Text Window Menu Commands The menu of a text window contains a number of standard options Chapter 6 2 1 File Export File Export selected This command is used to export data as ASCII text Tabbed ASCII text as Rich Text Format or as HTML The files can be read into a word processor or spreadsheet The exported file type is selected using the Files of type item in the File Export dialog box File Export selected limits the exported data to highlighted lines in the text window Chapter 6 2 2 File Print This command prints the contents of the text window Chapter 6 2 3 View Configure This command invokes a dialog box that lists all columns of data that can be displayed in the text window Columns can be moved between the list of displayed columns and the list of invisible columns Columns will be displayed in the order they appear in the list This order may differ from the default order that is used in the list of invisible available columns The order may be changed by selecting one or more lines in th
77. Eee 4 Grouping Peak types Selection Display HARVARD __1_ON_ ROX HARVARD__1_ON_ ROX List peaks ME HarvarD_1_on_ Fam all matching missing JE HaRVARD__1_ON_ FAM MMM Harvard _1_on_ vic I List all data sets in each group E HARVARD__1_ON_ vIC MM HarvarD__1_ON_ NED HARVARD__1_ON_ NED a HARVARD__1_OT_ ROX HARVARD__1_OT_ ROX MIB HarvarD_1_oT_Fam E HarvarD_1_oT_Fam v gt Select all car x J Deselect Cancel Help 1 The comparison sheet can either list all the peaks in all the data sets or only those peaks that occur in more than one data set i e peaks that have matching peaks in at least 2 data sets DAx also has the option of only showing missing peaks peaks that do not occur in all data sets This is a very powerful way to find differences between analyses T Require presence of peak from 204 2 List all data sets in each group causes each group of peaks to include a line for each data set even if a data set does not contain a peak in that case a mostly empty line is shown 3 The require presence of peaks from option is used to list only those peaks in the comparison sheet that have corresponding peaks in the selected data set Typically that data set contains a standard set of peaks and only peaks matching those standard peaks are of interest Select Data Sets for Comparison Sheet 2 xi Grouping Peak types Selection Display MM HAR
78. FontBar to indicate which data plot object will receive which data set e Peak List e Peak Item A peak item reports a certain peak value about a component Items that can be reported are peak name peak types peak concentration peak area or peak height The name of the component also has to be specified or you can specify a named peak index to print data about the first second named peak and so on e Analysis Assay This displays the Analysis Assay of a data set 239 Comparison Sheet This is a comparison sheet Global Comparison Sheet Binning Sheet This is a binning sheet GPC Calibration List This is a list of the GPC calibration points GPC Calibration Graph This is a plot of the GPC calibration Mw Plot This is a plot of the data using an Mw axis instead of a time axis Like the Data Plot item all data sets can be plotted in a single Mw Plot item or a separate Mw Plot item can be generated for each data set e Calibration List This is a list of the calibration points Calibration Graph This is a plot of the current calibration Calibrated Axis Plot This is a plot of the data using a calibrated axis instead of a time axis Like the Data Plot item all data sets can be plotted in a single item or a separate item can be generated for each data set e Database This is a list of the identification database Quantitative Calibration List This is a list of the calibration points for the quantification of a
79. This is used in plotting concentration versus plotting weight NB If you have checked Local GPC Calibrations in the File Customise gt Extensions dialog box the dialog box will look more like the one discussed in chapter Chapter 17 6 Chapter 15 6 GPC Windows GPC calibrations can be investigated in two types of windows There is a text window listing the calibration points and the parameters for the calibration polynomial There is also a graphics window that displays the calibration points and the polynomial Use the GPC Calibration list and GPC Calibration curve menu options to display these windows Chapter 15 6 1 Calibration List window The Calibration List window has the usual text window functionality chapter Chapter 1 It also has the option of deleting rows to remove calibration points To remove a point click the right mouse button on the line containing the point and select the Delete entry option in the pop up menu that appears These are the columns in the Calibration List window e point index 166 e polynomial fit parameters applicable only if the calibration uses a polynomial e usage points are either included in or excluded from the calibration e elution volume e molecular weight at elution volume e check value applicable only for polynomial calibrations The check value is the molecular weight calculated for the elution volume in the row using the current polynomial e data source name of the m
80. VARD 1 0N Fam J List only peaks of types JE HaRvARD__1_ON_ Fam MIB Hararo ion vIe JJ HaRVARD__1_ON_ VIC MIB HarvarD_1_0N_ NED HARVARD__1_ON_ NED cS ai HARVARD__1_OT_ ROX HARVARD__1_OT_ ROX MIM Harvarp_1_oT_ Fam E HarvarD_1_oT FAaM v Uia 4 Select all far x I Deselect Cancel Help 4 It s possible to include only peaks of certain peak types in the comparison sheet One possible use of this is to create a list of all peaks that require attention i e that have peak type attention HARVARD_1_ON_ FAM HARVARD__1_ON_ WIC JE HaRvaRD__1_oN_ vIc MIB HarvarD_1_0N_ NED HARVARD__1_ON_ NED B HARYARD_1_0T_ ROX HARVARD__1_OT_ ROX J7 No data set names at top of window 5 MIM Harvarp_1_oT_ Fam E HarvarD_1oTsFam v rt Select all lt all gt v Deselect Cancel Help 4 Using the Do not show statistics option forces the comparison sheet to list only peaks Averages and standard deviations are not displayed If you do show statistics DAx lets you select which ones to show 5 Comparison sheets normally list the names of all data sets included in the sheet at the top of the sheet including noise levels This list can be removed by checking the No data set names at top of window box Some versions of DAx can display Global Comparison Sheets which contain data from multiple graphic windows Use the Analysis Global sheet menu option to set up the global comparison
81. Windows Chapter 15 6 1 Calibration List window Chapter 15 6 2 Calibration Curve window Chapter 15 6 3 When the calibration changes Chapter 15 7 Local Calibrations Chapter 16 DAx CE Capillary Electrophoresis Extensions 169 Chapter 16 1 Calculating Mobilities step by step Chapter 16 2 CE Capillary Parameters Dialog Box Chapter 16 3 CE Load calibration CE Save calibration menu options 170 Chapter 16 4 CE Apparent mobility CE Effective mobility CE Track mobility menu options Chapter 16 5 Mobility Axes Chapter 16 5 1 Converting to a Mobility Axis Chapter 16 5 2 Mobility Axis Options Chapter 16 6 Time Corrected Signal Axes Chapter 17 DAx Calibrations 157 157 158 158 161 161 162 162 162 163 163 163 164 166 166 167 167 168 169 170 171 171 172 172 173 175 Chapter 17 1 Setting Up the Calibration Type 175 Chapter 17 2 Creating a calibration 176 Chapter 17 3 Calibration Load calibration Calibration Save calibration Calibration Stop calibration menu options 177 Chapter 17 4 Calibration Track value menu option 177 Chapter 17 5 Calibrated Axes 177 Chapter 17 5 1 Calibrated Axis Options 178 Chapter 17 6 Calibration Dialog Box 178 Chapter 17 6 1 Local and Global Southern Calibrations 180 Chapter 17 7 Automatic Calibration Derivation 181 Chapter 17 8 Calibration Windows 181 Chapter 17 8 1 Calibration List window 181 Chapter 17 8 2 Calibration Curve window 182 Chapter
82. a sets If many data sets are included in the binning sheet this can cause very wide lines Lines can be wrapped so that each line contains only as many data sets as fit and additional lines are used to accommodate all data sets Popup menu Display single bin View Previous bin View Next bin View All bins DAx has the option to display a single bin at a time This is started using the popup menu in the binning sheet which is invoked by clicking the right mouse button lines are automatically wrapped if a single bin is displayed Use the Previous bin and Next bin menu options to navigate to different bins you can also use the Ctrl PgUp and Ctrl PgDn keyboard shortcuts for this 22 if multiple peaks per bin are listed a number of alternative quantities can be used to quantify the binning map selected on the Quantifying parameter page of the binning sheet setup dialog box 219 Analysis Convert to user defined bins Use this option to convert automatically assigned bins to user defined bins Chapter 18 12 7 Method for Grouping Peaks Together The method to group peaks is the same as for comparison sheets cf chapter Chapter 18 11 1 Chapter 18 13 Meta analysis Meta analysis uses the presence of one or more peaks in one or more data sets to determine the likelihood that the data sets contain certain species For instance in DNA fragment analysis a set of analysis traces may be known to potentially contain a seri
83. able are not supported there is no extrapolation in that model e setting upper and lower limits for valid calibrated values This may be used to limit the extent to which the calibration can be extrapolated beyond the scope of the calibration points 178 e storing local copies of calibrations in data sets retrieving locally stored calibrations and making them the main calibration Edit Main Calibration A Set Calibration Type Derive Main Calibration from m Calibration Curve Type Copy Main Calibration from B erineda Store Main Calibration in Change Configuration for Multilinear 2 Delete Calibration from Cubic Spline a is C win SETUP DAx test3 dax C Polynomial Degree I WA E STAND 1 Da1 C Local Southern STAND 1 Da1 E 5158 4 Da1 C Global Southern 5158 4 Da1 m Peak Coordinate Offset Limits Low id Al High 1E 20 Al I Derive from data 3 m Base pairs Limits L Do got aller guint usage 5 Low fo pJ High fiE 20 pJ Add to existing calibration Cancel Help 1 Edit Main Calibration is used to change an existing calibration for instance to set a new polynomial degree or new parameter limits This is also used to pre set parameters when no calibration is present yet the pre set parameters will be used when a calibration is derived from automatic analysis of data Derive Main Calibration is used to derive a calibration from the selected data sets If local calibrati
84. al concentration is used to calculate relative concentrations All total concentrations for a set of enslaved data sets are always the same Finally a dilution may be entered for the sample All dilutions for a set of enslaved data sets are always the same Chapter 8 6 Data Tag splines menu DAx has the option of defining a spline for a data curve A spline is defined here as an array of nodes through which either a collection of straight lines or a series of cubic polynomials a cubic spline is drawn The spline may be turned into a new data set or it may supersede the data set See also chapter Chapter 3 3 16 See chapter Chapter 8 7 for details on modifying splines Splines are especially useful to create or modify baselines Use one of the built in baseline construction techniques chapter Chapter 10 1 to create an initial baseline data set Then a spline may be added to this data set The spline can be modified and may finally supersede the data set The new data set can then be used as a baseline It is not recommended to modify any data other than baselines this way 79 Bring to top Info v Plot Attention Colour gt Use as baseline Blank measurement Peaks gt Sizing gt Fourier filtering gt Splines v Add spline Cubic spline Tacks gt A Add label Replace curve with spline gt Entire curve l Turn spline into curve window only Gradient gt a Turn spline i
85. al value then determining the average of the signal values that lie below the previous average This process can be repeated several times Three passes generally yield excellent results DAx uses a slight modification to allow for negative peaks if more points lie above than below the first average the one where all the signal points were used peaks are thought to drop down from the baseline instead of rise above it The algorithm then turns into average above average The average below average algorithm is not suited to data sets with positive and negative peaks Instead use the modus algorithm DAx has several drifting baseline construction algorithms 107 e Moving average baselines are constructed by repeatedly calculating a moving average over the data From the second repeat onwards the data points that lie furthest from the previous pass are left out Parameters are width and number of passes Values of 10 for width with 3 passes may give good results Narrower peaks require narrower widths The number of passes may be automatically adjusted If automatic passes are selected the algorithm will stop as soon as further passes yield worse results than the previous pass e DerLim baselines Parameters are width and passes see above Somewhat narrower widths should be used here than for moving average baselines An added parameter is the percentile value which determines which percentage of the data set is used to construct th
86. always use the same units Fraction units range from 0 for the lowest signal value in the data set to 1 for the highest signal value Vertical axis units are typically volts Ois C win SETUP DAX test3en4 dal Indicate if peak search should be STAND 1 Da1 limited STAND 1 Da1 5158 4 Da1 Limit peak search to zoomed window 5158 4 Da1 Skip initial d Z 2 on horizontal axis type lt all gt x Select all lt a gt I Deselect lt Prev Next gt Finish Close Help Fourth page of the peak find wizard Limits the area in which the peak search is performed 1 Limit to zoomed window limits the peak search to that part of the data set that is currently visible in the data set graphics window Any peaks already present in the window will be removed but peaks outside of the window will not be affected This makes it possible for instance to search for peaks in two small time intervals and disregard the rest of the data Use these controls to enter an initial horizontal axis interval where no peaks will be detected expressed in the horizontal axis units in effect at the time of analysis It is possible to skip an interval only on certain types of axis 115 oO C win SETUP DAX test3 da Indicate if shoulder peaks should STAND 1 Da1 be automatically allocated STAND 1 Da1 5 5158 4 Dal Detect V Leading Shoulders 5158 4 Dal I Trailing Shoulder
87. ameters as well as the use of parameter definition files please refer to the DAx help file 31 Chapter 4 Measurement window Scanning for files Chapter 4 1 DAx measurement window The DAx measurement window is the window that is used to perform data acquisition by letting you start and stop measurements and by letting you set up measurement sequences Measurement windows are opened using the File New command found in most menus There is also a toolbar button to do this Please refer to the Measurement Window manual for further details Chapter 4 2 Scanning for files DAx has the ability to automatically scan for any new data files that appear in a specified location on the hard disk or on the network These files will be opened and possibly added to the data list and analysed Afterwards they can be moved to a new location This feature is intended to be used with data acquisition hardware that is not supported in the DAx measurement window Use the File Scanning Scan for files menu option to start scanning for files or to suspend scanning Use the File Scanning Scan setup menu option to specify where to scan for files and how to handle new files see below for details Turn on the Scroll Lock to make DAx stop scanning after a single file has been handled DAx will scan for new files every 60 seconds or every 5 seconds if the scroll lock is on 33 Chapter 4 2 1 File Scanning Scan setup menu o
88. ancel or redo the last sizing operation Sizing undo is reset every time the bounding rectangle is hidden i e once the bounding rectangle is hidden no undo is possible anymore 4 Use the undo all sizing menu option to restore sizing parameters time offset frequency voltage offset and factor to their original values i e the values they had when the measurement was first performed Press the Shift key while executing this menu option to restore sizing parameters for all data sets in the graphics window Chapter 8 5 The Data Set Sizing Dialog Box This dialog box is displayed when the Sizing Dialog item in the data tag sizing menu is invoked Changed values will be displayed in blue Changed values are applied to all data sets checked in the list at the left side of the 17 dialog Some changes automatically check select an entire enslavement group because they must be applied to the entire group Data sizing S158 4 Dal 1 2 x EX C win SETUPIDAxitest4 dax C Data Generic z lt none gt gt el oe x Horizontal Axis Unit 5158 4 Dal COM stano 1 ba1 l Freq Hz 10 WA Name Volt M DE stanp 1 ba1 Offset s 0 00 Offset jo J Inett sk joo GA Factor 1 35842 DA 3 Unit Time s z Move first point to 0 0 Flow Rate ml min ft Z New window New Curve Total Concentration fo Al Cancel Help Dilution fo Al 1 Time coordinates will be calculated as time time of
89. and warnings as well as program start and stop and operator name changes are logged in a message log The log may be turned off Chapter 3 1 4 File Customise gt Baselines DAx can construct baselines using several algorithms You may hide any of these algorithms from users by customising baseline construction Some of the algorithms use parameters width number of passes percentile value The baseline construction dialog offers a choice of deriving the parameter values from the data the Auto setting or entering a fixed value Using baseline customisation you may hide any or all of these parameters from the user Hidden parameters will use the auto setting i e will be derived from the data The DerLim baseline construction algorithm uses a Savitzky Golay technique to determine derivative signal values The filter width used may be entered in the baseline customisation dialog box Chapter 3 1 5 File Customise gt Finding peaks You may elect not to display the complex peak find dialog box to the user but use a simplified form instead The simplified dialog box causes automatic thresholds as well as slope limits to be used Peaks must be at least 6 times as high as the noise level To find peaks using signal slope a Savitzky Golay algorithm is used to determine derivative signal values The filter width used may be entered in the peak customisation dialog box Several options are available that determine how shoulder pea
90. apter 21 7 Why is the value for RMS noise often half the value for PP noise in the peak list window 247 Chapter 21 8 Can manually remove a drift or trend from a signal 247 Chapter 21 9 How do I account for different injection volumes 248 Chapter 22 Index 250 10 Chapter 1 Introduction DAx is a program for data acquisition and data presentation and analysis This User s Manual is intended as a complete reference to DAx It is recommended that you read the Quick Start Guide first Chapter 2 Installation Chapter 2 1 System Requirements To be able to run DAx your computer must have the following A PC running MS Windows version 9x or better A minimum of 128 megabytes of RAM A VGA or better graphics adapter Graphics adapters capable of displaying 800 600 resolution recommended A mouse or similar pointing device CD ROM drive for installation e A hard disk with a minimum of 30 megabytes of free disk space You must have permission to make changes to the system registry and permission to create the DAx directory If you are unsure whether or not you have these permissions please ask your systems administrator DAx must have write access to the directory it is installed in even during normal program use Chapter 2 2 Installing the Software _ Ol D 7 Insert the DAx distribution CD into the CD ROM drive We will assume drive R is being used From the Start menu execute the Run option
91. ar marker denotes the presence of a peak Manually added peaks are marked with an open triangle normal peaks are marked with a filled in triangle Double click on the curve sample to add a peak if none existed or remove a peak Chapter 18 12 5 Binning maps Binning maps are graphical representation of binning sheets in which each bin is represented by a coloured dot The colour of the 218 dot corresponds to the value of the binning sheet s quantifying parameter22 A binning map is displayed using the binning sheet s View Binning map menu option In the binning map use the View Use trace colours menu option to use the trace curve colour of each data set When trace colours are not used all values are mapped using the colours set using the View Attributes menu option When you move the mouse over a binning map the corresponding area of the binning sheet is brought into view unless you press the Shift key Right clicking displays a popup menu which can be used to display the pertinent area in the source trace data Chapter 18 12 6 Binning sheet menu options Binning sheets can be saved to disk using the Analysis Save binning sheet menu option and loaded from disk using Analysis Load binning sheet To remove all retained data and close the binning sheet use the Analysis Clear binning sheet menu option View Wrap lines Each line in the binning sheet normally contains information from a single bin for all dat
92. arkers for the shouldered peak enlarged in insert DAx will try to establish if shoulder peaks exist based on a number of criteria See below To mark additional shouldered peaks press the Shift key while adjusting either peak boundary of the shouldered peak in the normal way by dragging the peak begin or end marker Shouldered peaks are marked in the graph their begin and end markers have an extra Smaller begin or end marker Shouldered peaks display an S behind their index number in peak list windows chapter Chapter 10 11 Shoulder peaks display an S Note that the Shift key needs to be pressed each time the begin or end marker of the shouldered peak is moved or the peak will no longer be a shouldered peak It is recommended to do any manual peak additions before manually defining shoulder peaks Chapter 10 10 7 Automatic Shoulder Peak Detection DAx has a number of ways it can use to try and establish if peaks are shoulder peaks Peaks can be considered shoulder peaks for four possible reasons e High slant i e the signal value of the peak end is much closer to the signal value of the peak top than to the signal value of the peak begin or vice versa In other words a line drawn 130 connecting peak begin and end coordinates has a high slant The degree of slant is the ratio of the signal difference between peak begin and end and the signal difference between peak top and either peak begin or peak end e Hig
93. as baseline plotting and hiding the graph for the data set Blank measurement Peaks as requiring further attention The data tag will be as displayed with an exclamation point on the right side _ Saing Dialog boxes such as the plot select dialog box have the Fourier filtering ability to select only data sets that require attention oF plines set the baseline Other data sets that have been marked Tacks as baselines will be unmarked as each non baseline Add label data set can have only one baseline However as an i Gradient example a data set and its filtered version can each have its own baseline GPC calibration Calibration the data set is a blank measurement Beekse eta For operations requiring a baseline if no baseline is Data table found DAx will check for the presence of a blank Copy measurement and use it as the baseline Blank _ Delete data measurements are indicated with a blue B at the left of their data tag Use the Fourier Filtering Add Fourier bar menu option to generate a frequency domain data set to be used in Fourier filtering chapter Chapter 10 Refer to chapter Chapter 11 2 for details on the Derive Fourier characteristics menu option Press the Shift key while clicking the menu option to generate frequency domain data sets for all data sets in the graphics window The GPC Calibration and Calibration menu options are used to list plot or delete the data set s
94. atic bins Qualifying coordinate Trace types Bin intervals Bin name Low Base High Base In Trace Peak type use and selection Curve samples and peak parameters 205 00 Curve sample scaling 2 Second 235 00 238 00 lt all gt Quantifying parameter labels Peak parameter selection 3 Third 285 00 292 00 lt all gt Statistical items New Display options Peak thresholds Bin annotations Persistent data lt Prev P P Allow overlapping bins requires bins equal peaks Sort by size Sort by trace type Import OK Cancel Apply If the Allow overlapping bins option is in effect the program does not check if user defined binning intervals overlap This option requires that Bins equal peaks is also in effect The option is useful when expected peak intervals for consecutive peaks are fairly wide and therefore overlap Intervals can be sorted by size only or by trace type then by size The import button can be used to import interval definitions from an appropriately formatted text file refer to the on line help for a specification of the file format Peak type use and selection On the Peak type use and selection parameter page you can choose to limit the binning sheet to peaks of certain types This 211 could for instance be used to create a binning sheet containing only peaks that have been marked as needing attention Curve samples and peak parameters Binning sheets can displ
95. ation can be straight line where extra points are added along a straight line between existing points or cubic spline where points are added along a cubic spline through existing points Interpolating on a cubic spline yields a smoother curve 93 Chapter 9 6 Data Slice If the menu option Data Slice is not present you need to use File Customise gt Data and check Data Slicing This command invokes a dialog box that allows parts of a measurement to be removed or replaced by a straight line The dialog box uses these parameters e begin and end of interval to be sliced By default this interval is equal to the current graph window e slicing mode Part of the measurement can be removed or replaced by a straight line or zero values Peaks and data tacks in the sliced interval are removed Chapter 9 7 Data Remove trend If the menu option Data Remove trend is not present you need to use File Customise gt Data and check Removing trends This command invokes a dialog box that allows polynomial trends to be removed from data The trend can also be turned into a new curve Chapter 9 8 Filtering Data Filter derivative These commands invoke a dialog box that allows data sets to be filtered using a Savitzky Golay moving average filter or Fourier filter It is also possible to calculate first or second order derivatives Savitzky Golay only Additionally spikes can be removed To remove spikes without a
96. aving bins automatically derived Bins are entered manually see below or defined in the graphic window or colour view window see chapter Chapter 18 12 3 The bins equal peaks option available when user defined bins are used causes all existing peaks to be removed from data sets that are added to the binning sheet New peaks with coordinates exactly equal to all user defined bins are then created in the data sets This option is used when the shape of the curve in the predefined bins is more important than whether or not a peak can be found there The peaks that are created this way cannot be manually edited As long as a data set is in the binning sheet it is excluded from the Peaks Find peaks menu option The multiple peaks per bin option allows more than one peak to be listed for each bin This option requires the use of user defined bins A maximum number of peaks listed per bin can be set Up 209 to the number of specified peaks are listed for each bin if they are present in the data The current quantifying parameter is used to determine which peaks are listed the peaks with the highest values of the quantifying parameter are listed preferentially Leaving the field blank causes all peaks in the bin to be listed When the number of peaks in a bin is larger than the maximum number listed the remaining peaks define a residual relative area which can be used as a quantifying parameter when drawing binning maps see below
97. ay either a quantifying parameter or a curve sample of each peak Displaying curve samples makes it possible to quickly compare large numbers of peaks See chapter Chapter 18 12 4 for more details on curve samples In addition to the quantifying parameter or curve sample a qualifying coordinate or list of peak data can be displayed You can prevent this by checking Do not show qualification coordinate when a quantifying parameter is displayed or Do not show peak parameters when curve samples are drawn If multiple peaks per bin are listed peak parameters must always be displayed Curve sample scaling When curve samples are drawn the Curve sample scaling parameter page is used to choose how curve samples should be scaled e with bin scaling peaks are scaled relative to the tallest peak on each line e with sheet scaling peaks are scaled relative to the tallest peak in the entire binning sheet e with peak scaling peaks are scaled individually so each curve sample will be equally tall Quantifying parameter labels When a quantifying parameter is displayed it is chosen on the Quantifying parameter labels parameter page Choices include peak height and peak area Instead of a quantifying parameter you can choose to display just a label whenever a peak is present e g Check Only list presence absence to display labels There is also a label that is displayed whenever no peak is present e g 212 If multipl
98. be lost from the signal by definition some information is always lost when filtering is applied The filter width has to be an odd number in order to make the filtering process symmetrical across points preceding and following the data point being smoothed This restriction does not apply to Fourier filtering because in Fourier filtering the width parameter is used in a different way 8 In Fourier filtering the filter width has a different meaning Refer to chapter Chapter 11 1 95 amp Savitzky Golay Moving Average Fourier filters 2 x Sie C winSETUPDAx test4 dax C win s No filter f 5158 4 Dal Moving avaraga 1 OI stano 1 pa1 C Savitzky Golay DE stanp 1 ba1 C 1st Derivative 2nd Derivative Fourier Filter width 25 Al V Remove Spikes Drift Factor jas A I New Window New Curve Close Help 1 Check the Savitzky Golay radio button to perform Savitzky Golay filtering on the selected data sets You may also elect to determine first or second derivatives Only data sets that are not already a derivative of some kind can be derived Check the Moving Average radio button to perform moving average filtering If the filter width is N each point will be replaced by the average of N points the N 1 2 points preceding it the point itself and the N 1 2 points following it 2 Check the Remove Spikes check box to invoke the spike removal algorithm Spikes will be removed f
99. calculate an explicit RMS noise level Chapter 21 8 Can manually remove a drift or trend from a signal Yes you can Let s suppose this is your original signal DAx 5 0 PP 21 02 96 18 41 25 AU 0 20 40 time min The signal is horizontal at first then has a slope then ends up horizontal again To remove this trend e use the tag menu chapter Chapter 8 2 to create a second copy of the data e use the tag splines menu chapter Chapter 8 6 for the second copy to add a spline e move the nodes for the spline to form three straight lines 247 e replace the curve with the spline again using the tag splines menu to get something like this DAx 5 0 PP 21 02 96 AU Example of copy of curve being used to define a trend The curves have been slightly offset for clarity e now use the Data Arithmetic option to subtract the second curve from the first The resulting curve will look like this DAx 5 0 PP 21 02 96 18 54 02 AU T T T 0 20 40 time min Chapter 21 9 How do account for different injection volumes Suppose that normally measurements are performed with an injection volume A Suppose that a measurement with dilution D is performed using an injection volume B To account for the injection volume enter D A B as the dilution 248 249 Chapter 22 Index de activating peaks See peaks ABI files 30 action list 85 activating peaks See p
100. cified value the analysis is flagged as either suspect or bad Chapter 18 15 5 Unwanted peaks A list of names of peaks that may not be present or may not be present in more than a specified amount can be entered A quantification parameter is selected height area concentration area fraction If a peak with the specified name is present and its quantification parameter is higher than the specified value the analysis is flagged as either suspect or bad 22 Chapter 18 15 6 Calibration Quality These parameters inspect the calibration that is being used if the data set being analysed has a locally stored calibration it is used otherwise the main calibration is used The following parameters can be inspected average distance between calibration points and calibration curve The Root Mean Square is taken If relative limits are used the average difference between calibration points and calibration curve is taken relative to the range of calibrated values This parameter is only available for polynomial calibrations point to point and spline calibrations go through all calibration points low high limit of calibration curve derivative For polynomials the first order polynomial factor is used For splines the average derivative is determined at all the calibration points high limit on calibration curve curvature For polynomials and splines the lowest and highest derivatives at all calibration points are determined T
101. clusion flag If this column contains a check mark the component will by default be excluded from area calculations To display the quantitative calibration for a component click the right mouse button on its line then execute either the List quantitative or Draw quantitative option in the pop up menu To set a new qualifying parameter set a marker name or set a new tolerance value for a number of components select the lines containing the components Now click the right mouse button and execute the Set up qualification menu option To set a new quantifying parameter change the type of quantitative calibration or enter new concentration limits for a number of components select the lines containing the components Now click the right mouse button and execute the Set up quantification menu option Chapter 18 9 2 Quantitative Calibration List Window The Quantitative Calibration List window has the usual text window menu options chapter Chapter 6 2 It also has the option of deleting rows to remove calibration points To remove a point click the right mouse button on the line containing the point and select the Delete entry option in the pop up menu that appears Please 19 Not all versions of DAx support this 201 refer to chapter Chapter 18 9 4 to find out what happens when a quantitative calibration is changed These are the columns in the Calibration List window e point index e polynomial fit parameters applicable only
102. concentration abundance or weight plots Chapter 15 5 GPC Calibration Dialog Box This dialog box can be used to derive a GPC calibration from a number of data sets or to switch between calibration models The dialog is invoked using the GPC Calibrate menu option If a calibration is being derived from data sets the data sets must have peaks with the appropriate Molecular Weight as their annotations Refer to chapter Chapter 15 1 for step by step instructions on the steps that should be taken before this dialog box is invoked It is also possible to have molecular weights as part of peak names The GPC calibration dialog box is used for a number of purposes e creating the initial GPC calibration list e setting the calibration model straight lines cubic spline or polynomial All of these models act either on logarithmic or linear molecular weights as indicated in the dialog box A calibration should be based on logarithmic molecular weights if there is an approximate linear relationship between elution volume and the logarithm of the molecular weights of the components e setting upper and lower limits for elution volumes that can be converted to a molecular weight value If the calibration model is 164 spline molecular weights outside the molecular weight range of the calibration table are not supported there is no extrapolation in that model e setting upper and lower limits for valid molecular weights This may be
103. cular weight values are listed Additional columns list the source for each point in the form of a measurement name as well as the point s usage status points may be included or excluded from the calibration Mobility axis windows chapter Chapter 16 5 These windows display the same signal values as graphics windows but they use mobilities for the horizontal axis Time Corrected Signal Axis windows chapter Chapter 16 6 These windows plot signal values divided by migration coordinate instead of just signal values along the vertical axis Calibrated axis windows chapter Chapter 17 4 These windows use the current calibration to calculate a calibrated value for each data point The calibrated value is then used as the horizontal coordinate 21 Calibration Curve chapter Chapter 17 8 2 The current main calibration or a local calibration is plotted in this window Calibration list chapter Chapter 17 8 1 The current main calibration or a local calibration is listed in this window For each calibration point the calibration parameter and calibrated value are listed Additional columns list the source for each point in the form of a measurement name as well as the point s usage status points may be included or excluded from the calibration Identification Database list chapter Chapter 18 9 1 The current Identification Database is listed in this window For each entry the component name and qualifying parameter values ar
104. d and the current peak is negative or vice versa 117 e or if the current peak has already gone through a maximum top and gone through a minimum valley Chapter 10 4 5 Why use slope requires threshold In noisy measurements using slope limits to detect peaks may cause too many peaks to be found However not using slope limits would mean that two overlapping peaks in cases where the signal value between the peaks never drops below the threshold value would not be detected as separate peaks The option slope requires threshold solves this problem by using the slope limit to start a new peak only in places where the signal value is at least the threshold value The graph gives some examples of peaks that can be found using thresholds or slope limits threshold 1 2 3 4 5 6 7 noise level e peak 1 is below the noise level so it will not be detected using threshold or slope limits e peak 2 is below the threshold but might be detected using slope limits e peak 3 is above the threshold so it would be detected using thresholds It might also be detected using slope limits e peaks 4 and 5 are threshold separated i e the signal drops below the threshold between peaks They would be detected as separate peaks using threshold detection 12 A noise level is indicated in the graph The signal obviously contains almost no noise but a non zero noise level was plotted for clarity 118 e peaks 6 and 7 are not threshold sepa
105. d from each other for instance you might want to subtract a blank measurement from a data set e data sets can be multiplied or divided e MIN MAX a data set can be constructed that at each time coordinate has a data point equal to the highest or lowest value of operand and operator data sets at that time coordinate This technique might be employed when two detectors are used to create a data set that shows peaks when either detector detects a component e dC1 dC2 a data set that equals the derivative of one curve with respect to a second curve can be constructed For instance if the first curve contains a total energy reading and the second curve a total mass reading the derivative would give the amount of energy per added mass Curve to curve differentiation dC dC is only possible on voltage data not on derivative curves 9 Curve arithmetic signal values for the operand and operator curves at each time coordinate are added multiplied etc Signal values for the operator curve may be interpolated if time coordinates do not overlap 97 Data Arithmetics 2 xi m Operator m Operand EPs C DAxDTest45 TEST3 Dax CADA gt a E o PAR stanp 1 ba1 Cei al C constant Al DE stanp 1 ba1 oe curve a sa ae x c Eie C ADAxDTest4S TEST3 DAX C DA PAER H O stanp 1 ba1 2 malaji OE stano 1 a1 Time Ol 5158 4 pa1 C MAX OH 5158 4 Da1 C MIN M New Window V New Curve dC1 dC2 Cancel
106. d to drop to the threshold level for peaks to be detected as separate If derivatives are also used to detect baselines no threshold separation is required e the derivative of signal minus baseline versus time traverses a certain limit Once again separate limits are available for positive and negative peaks Slope limits may be calculated automatically from the data or entered manually If the derivative tops out for positive peaks then goes through a minimum then once again traverses the start slope another peak will be started even if the peaks are not baseline or threshold separated This occurs when there is a dip between two signal maximums Only signal values more than the noise level above or below the baseline can start a peak DAx calculates the point to point noise level for this purpose The point to point noise level and possibly calculated RMS noise level are displayed in the peak list window If the slope requires threshold option is in effect the signal value needs to reach at least the threshold value above the baseline for a peak to be recognised Refer to chapter Chapter 10 4 4 for details on using slope limits DAx can use one or both of the above techniques 112 Use the Show Boundaries option in the data tag peak menu chapter Chapter 8 3 to make DAx display the boundary conditions that were most recently used to detect peaks for a data set The peak detection algorithm determines the signal noise level
107. disabled Fourier filtering may also be customised Refer to chapter Chapter 11 1 for details 18 Chapter 3 1 8 File Customise gt Measurement window Refer to the Measurement Window manual for a detailed discussion of Measurement window customisation Chapter 3 1 9 File Customise gt Measurement data directory This dialog box lets the user select where measurement files that have not been explicitly named should be stored You can also choose various forms of file naming conventions Refer to the Measurement Window manual for details This location is also used to store temporary files Chapter 3 1 10 File Customise gt Fields setup A dialog box is shown that allows the number and names of the extra input fields stored with new measurements to be set Chapter 3 1 11 File Customise gt Report headers A dialog box is shown that allows up to five header lines to be entered Report definitions can then use the variables header 1 through header 5 in text objects When the report is printed the headers will be inserted Chapter 3 1 12 File Customise gt Data lists This option lets you specify when to start a new data list A new list may be started every day every week on a certain day or whenever there are a set number of entries in the list A new list will also be started whenever the list is full It is also possible to start a new data list for each measurement sequence If part of a sequence list is
108. e Customise 56 Chapter 7 1 14 File Exit 56 Chapter 7 2 Data Set Graphics Window Edit menu 57 Chapter 7 2 1 Edit Select all 57 Chapter 7 2 2 Edit Select 57 Chapter 7 2 3 Edit Invert selection 58 Chapter 7 2 4 Edit Copy graph 58 Chapter 7 2 5 Edit Create metafile 59 Chapter 7 2 6 Edit Undo Edit Redo 59 Chapter 7 3 Data Set Graphics Window View menu 59 Chapter 7 3 1 View Attributes 59 Chapter 7 3 2 View Plot Select 59 Chapter 7 3 3 View Plot all 59 Chapter 7 3 4 View Plot previous View Plot next View Plot data sets View Plot groups 59 Chapter 7 3 5 View Set colours 60 Chapter 7 3 6 View Data set names 60 Chapter 7 3 7 View Tags setup 60 Chapter 7 3 8 View Edit scale 60 Chapter 7 3 9 View Zoom in 60 Chapter 7 3 10 View Zoom out 60 Chapter 7 3 11 View Reset scale 61 Chapter 7 3 12 View Seconds axes View Minutes axes 61 Chapter 7 3 13 View Detector units curves View First derivative curves View Second derivative curves View Inter curve derivatives View Frequency domain curves 61 Chapter 7 3 14 View Track data set 61 Chapter 7 3 15 View Mark bins Do not mark bins 61 Chapter 7 3 16 View Mark bins Mark above graph 61 Chapter 7 3 17 View Mark bins Mark above and inside graph 61 Chapter 7 3 18 View Colour view 62 Chapter 7 3 19 View Data set base calls View Consensus base calls View Allele base calls 62 Chapter 7 3 20 View Remove Highlight 62 Chapter 7
109. e listed Additional columns list the source for each entry in the form of a measurement name as well as the entry s usage status entries may be included or excluded from the active database Quantitative Calibration list chapter Chapter 18 9 2 The window displays a table of concentrations and corresponding quantifying parameter values Additional columns list the source for each entry in the form of a measurement name as well as the entry s usage status entries may be included or excluded from the active calibration Quantitative Calibration graph chapter Chapter 18 9 3 The window displays the quantifying parameter as a function of component concentration Comparison Sheet chapter Chapter 18 11 A window displaying a table comparing peaks in various measurements Peaks are matched according to a qualifying parameter selected by the user or by their names Peak values are listed as well as optionally averages and standard deviations Binning Sheet chapter Chapter 18 12 A window displaying a table comparing peaks in various measurements Peaks are matched according to a qualifying parameter selected by the user Whereas a comparison sheet lists all peak parameters with successive peaks being listed on successive lines a binning sheet only lists the qualifying coordinate and a quantifying 22 parameter of corresponding peaks on a single line Far more complex binning sheets can be created displaying curve sample
110. e mouse to a new location Multiple objects may be copied see below but multiple objects cannot be sized Multiple objects can also be selected by pressing the Shift Key then dragging the mouse cursor across the graph A rectangle will be drawn When the mouse button is released all objects that fully lie within the rectangle will be selected Any previously selected objects will be deselected unless the Ctrl key is also pressed Chapter 20 3 4 Removing Objects To remove objects select the objects you want to be removed then use the Edit Cut menu option or press Ctrl X The removed objects are placed on the clipboard and may be pasted later Chapter 20 3 5 Copying Objects To copy an object on a single report page click the mouse on it to select it Keep the mouse button depressed then press the Ctrl key and drag the mouse to the location where you want the new object to appear The mouse cursor will have a plus sign to indicate an object is being added rather than moved Multiple objects may be copied at once Select the objects press the Ctrl key click one of the objects and drag to a new location Typically you will now want to set the object index of the copied objects to a higher value An other way to copy objects is to select them then use the Edit Copy menu option or press Ctrl C The objects will be placed on the clipboard Now go to a different page using the FontBar and use the Edit Paste menu option
111. e Identification Database has been changed but has not been saved to disk yet when DAx is about to terminate you will be asked to confirm the loss of the new database Saving the database to a floppy disk does not prevent this you must save to hard disk Loading a new database may cause peak names and concentrations to be changed This can be prevented by saving data to disk or using the data tag peak menu fixate option cf chapter Chapter 8 3 191 Analysis Stop analysis stops using the Identification Database and removes it from memory Chapter 18 3 Analysis Qualify peaks Whenever the Identification Database is changed e A new database will be calculated e Any graphics windows will have their peaks re qualified and re quantified However component names entered by the user will remain unchanged Fixating also prevents re qualification Peaks are fixated by saving to disk or using the Fixate option in the data tag peak menu Chapter 18 4 Identification Database Edit Dialog Box This dialog box is invoked using the Analysis Edit Database menu option If an Identification Database is being derived from data sets the data sets must have peaks with the appropriate names See chapter Chapter 18 1 for a step by step instruction on creating an Identification Database up till invoking the edit dialog box The Identification Database dialog box is used for a number of purposes e creating the initial Identification Databa
112. e List window also has the option of deleting rows to remove database entries To remove an entry click the right mouse button on the line containing the entry and select the Delete entry option in the pop up menu that appears These are the columns in the Calibration List window e entry index e component name e component type The type can be used to limit display to certain peak types in peak list windows using the View Select Peak Types menu option e annotation e usage lines are used to qualify amp quantify just to quantify or not at all e On Axes determines the types of horizontal axis a component can be identified ont8 It is possible to set up identification and quantification of a single component with the same name throughout on different axis types simultaneously DAx will use 18 Not all versions of DAx support this 200 only the database entries that match the axis type being analysed e In Trace determines the trace types a component can be identified in19 e qualifying parameter e marker name If the qualifying parameter uses a marker the name of the marker to be used is listed here If two markers are used both marker names are displayed here e qualifying parameter values e qualifying parameter tolerance To set a relative tolerance follow the tolerance value with a e number of quantitative calibration entries e data source name of the measurement the data in the row were derived from e ex
113. e View Linear and View Logarithmic menu options may be used to switch between linear and logarithmic axes Use the Analysis Config quantification see chapter Chapter 18 8 menu option to change the type of calibration curve for the component being investigated Chapter 18 9 4 When the database changes Whenever the Identification Database is changed by removing lines from the database list the following steps are taken e A new database will be calculated e Any graphics windows will have their peaks _ re qualified However component names entered by the user will remain unchanged Fixating also prevents re qualification Peaks are fixated by saving to disk or using the Fixate option in the data tag peak menu Chapter 18 10 Using Internal Standards Frequently internal standards are used to quantify components Since the amount of internal standard that has been added is known it is possible to quantify the unknown components by their peak size relative to the size of the internal standard peak The procedure to follow in DAx is as follows e make sure the internal standard is recognised as the reference peak cf chapters Chapter 18 7 and Chapter 18 6 e set up quantitative analysis using normalised peak areas as the quantifying parameter cf chapter Chapter 18 1 Since there will not be a normalisation peak present DAx will use the reference peak to calculate normalised areas e to account for different response fact
114. e baseline and is thought not to be part of a peak 75 often gives good results here Percentiles may also be automatically adjusted For the DerLim algorithm automatic width adjustment is also available e Fourier baselines are constructed by removing high frequency components from the data set What is left are the low frequency components that are thought to make up baseline drift The parameter used for a Fourier baseline is the width in percents of the number of points measured Values of 25 and up may give good results Refer to Modifying Baselines chapter Chapter 10 3 for details on how to manually change a baseline 108 r E gt Baselines 7 Baseline type Parameters 3 0 is C A win SETUP DAX TEST1 DAX ote vil DAXSIML PRN a Drifting ve V Auto Width 11 00 vill voork 9 txt G Derim Asied f C Fourier s oO C Moving Average C Just below trace V Auto Max Passes 3 A l3 C Moving Median C Moving Median 1 V Auto Percentile 40 4 M l Straight under peaks 5 Trend removal Horizontal l Remove polynomial trend 6l C Ave Below Ave M Use HPLC Gradient Modus 4 uw p wow Help 1 Select the data sets for which a baseline needs to be constructed here The width is an important parameter in baseline constructions Narrow peaks typically require narrow widths More importantly wide peaks require wide filter widths since a narrow filter will tend to let the
115. e calibration the concentrations of the components in the measurement are divided by the dilution factor before points are added to the quantitative calibration This means that the concentrations for the components should be set to the value before dilution e conversely when a concentration is calculated based on a calibration the concentration that is calculated is multiplied by the dilution factor of the sample before being displayed Chapter 18 8 2 Injection Volumes DAx does not keep track of injection volumes It is assumed that the same unspecified injection volume is used for all 199 measurements However different injection volumes can be accounted for using dilutions Suppose that normally measurements are performed with an injection volume A Suppose that now a measurement with dilution D is performed using an injection volume B To account for the injection volume enter D A B as the dilution As an example if the injection volume used for a sample is half the normal volume the dilution should be entered as double the actual dilution Chapter 18 9 Identification Database Windows Identification Databases can be investigated in a text window listing the database entries Chapter 18 9 1 Identification Database List Window The Identification Database List window has the usual text window menu options chapter Chapter 6 2 Items in the database can be edited from within the window The Identification Databas
116. e fit failed for a certain peak a Gaussian is plotted based on the peak s top width and area Total curve causes the sum of all Gaussians to be plotted Use the View Attributes gt Fitted curves menu option to select the type of line used to mark the Gaussians and Total curve The above three options can be applied to all data sets by pressing the Shift key when clicking on the menu option Turn total curve into data set creates a new data set from the sum of the values of all Gaussians This new data set will contain peaks in the same locations as the original data set for easy comparison 5 RMS noise calculation is discussed in chapter Chapter 8 3 5 6 Fixated peaks will not have their names or concentrations overwritten by the Identification Database Saving data to disk also fixates the peaks For more information see section Chapter 8 3 2 7 Use the List Peaks menu option to create a peak list window chapter Chapter 10 11 listing the peaks in the data set The contents of the text window can be printed copied to the clipboard or saved in a text file It is possible to configure the peak list window to include or exclude various data columns use the View Configure menu option 8 The data set can be added to or removed from the binning sheet Binning sheets are discussed in chapter Chapter 18 12 9 Peak data can be added to the analysis log using the Log in analysis log menu 10 Clear analysis removes the peaks and base
117. e list of displayed columns and then pressing the To Top Up Down or To Bottom button 43 TextWin Column Selection x Available Columns Displayed Columns Concentration Dilution Relative Concentration 2 Peak Type Signal to noise ratio Moment Peak Coordinate Offset Relative Peak Coordinate Capacity factor Select All gt gt Offset to marker Coordinate over marker lt lt Deselect All Peak Width d __ lt lt Deselectal_ In some text windows columns can also be re arranged by clicking in the column header and dragging it to a new location Only the columns that are displayed will be written to file chapter Chapter 6 2 1 printed chapter Chapter 6 2 2 or copied to the clipboard chapter Chapter 6 2 7 Chapter 6 2 4 View Fit columns View Shrink amp fit columns This command if available adjusts column widths so that all column contents will fit If the shrink option is chosen column widths will be adjusted to make all data just fit otherwise columns are only made wider as needed These options are also part of a pop up menu that appears when the right mouse button is clicked in the columns header Chapter 6 2 5 View Text size This menu option is used to change the size of the text in the text window If your mouse has a scroll wheel you can also change text size by pressing the Ctrl key then rolling the scroll wheel 44 Chapter 6 2 6 Edit Select
118. e peaks per bin are listed a quantity to be used to draw binning maps cf chapter Chapter 18 12 5 other than the quantifying parameter can be selected The choices are e area of peaks in bin relative area of peaks in bin migration time corrected area of peaks in bin relative migration time corrected area of peaks in bin residual area of peaks not listed in the bin due to the maximum number of peaks listed per bin being exceeded e residual relative area of peaks not listed in the bin due to the maximum number of peaks listed per bin being exceeded e area of the main peak in the bin relative to the total area of all peaks in the bin Peak parameter selection When curve samples are drawn or multiple peaks per bin are listed the Peak parameter selection parameter page is used to determine which peak parameters to display All peak parameters that can be listed in peak list windows chapter Chapter 10 11 can be included In addition to all the normal peak parameters the following items can be displayed e data set name e qualifying coordinate e quantifying parameter If multiple peaks per bin are listed these items are also available e area of peaks in bin relative area of peaks in bin migration time corrected area of peaks in bin relative migration time corrected area of peaks in bin residual area of peaks not listed in the bin due to the maximum number of peaks listed per bin being exceeded e residual relat
119. e removed using Remove bin or Remove selected bins A new bin can be defined using the Add bin option Chapter 18 12 3 User defined bins DAx has the ability to use user defined bins where the user sets the lower and upper limit of each bin as opposed to automatic derivation of the limits by DAx Bins can be defined in a number of ways e by entering an interval in the Binning sheet dialog e by importing binning intervals from a text file also done in the Binning sheet dialog e using a colour view window by double clicking on a peak marker Use the colour view s View menu to make sure each data set is displayed in its own lane and that peak markers are displayed e using a colour view window by pressing the Ctrl key and then clicking and dragging the mouse to define the location of the bin e in a data graph window by pressing the Alt key then dragging a box A popup menu is displayed containing the option to add a user defined bin equal to the dragged box e by right clicking the mouse in the binning sheet or on the binning map see below or in a colour view window and choosing the Add user defined bin menu option The bin s lower and upper limit must be explicitly entered when this method is used When user defined bins are used the binning sheet will contain a Bin name column where a name for the bin can be freely entered 21 Note that the preceding data set may not be in the preceding binning cell in particu
120. e used There are two ways in which a data set can obtain a local GPC calibration e Using the Store Main Calibration in option in the Calibration Set up dialog box invoked using the GPC Calibration menu option e f Derive GPC Calibration is checked in the Automatic Analysis setup when a file is opened with Automatic Analysis checked In this case a GPC calibration is derived from the data and stored as a local GPC calibration It is also automatically copied to be the current main GPC calibration The data tag popup menu chapter Chapter 8 2 contains options to list the local GPC calibration draw the local GPC calibration as a curve and remove the local calibration 168 el E Chapter 16 DAx CE Capillary Electrophoresis Extensions If the menu option CE is not present you need to use File Customise gt Extensions and check CE Options Not all versions of DAx contain this option The CE version of DAx has a number of extra options that make it possible to use the program to determine mobilities from electropherograms To be able to do this a number of parameters about the capillary need to be established Important DAx will store capillary parameters with data if capillary parameters have explicitly been attached to the data using the CE Calibrate menu option If no capillary parameters have explicitly been stored for a data set DAx will use the parameters that were last entered for any other data Chapter
121. eaks adding data to a window 48 adjoining peaks 18 adjusting text column width 44 AIA files 30 analyse 123 analysis creating an Database 190 Analysis Assay 224 Assay weighting 225 Baseline parameters 226 calibration quality 228 Peak parameters 226 required peaks 227 unwanted peaks 227 analysis logs 223 analysis procedure 23 loading 30 saving 30 analysis quality 224 Andi files 30 annotations 134 apparent mobility 169 172 area limit See peaks arguments 31 arithmetic 64 86 97 assay 224 asymmetry 137 attention data sets requiring 72 attributes 38 59 automatic analysis 49 84 loaded files 48 automatic calibration derivation 181 automatic peak recalculations 124 automatic shoulder peak detection 130 automatic thresholds 117 Automatic Trace Calibrations 183 average below average 107 averaging 92 averaging data 98 Identification 250 axis conversion 51 52 163 171 172 178 baseline 70 74 79 81 85 103 107 110 111 112 definition 27 subtracting baselines 110 baseline amp peaks command 123 baselines 27 average below average 107 customising 17 DerLim 108 Fourier 108 modifying 111 modus 107 moving average 108 batch analysis 31 Best overlay 105 binning maps 218 binning sheet 70 207 blank measurement 28 70 72 86 Bleedthrough peaks 121 blue B 70 boundary conditions 74 75 113 bounding rectangle 63 calculating concentrations 139 calcula
122. eaks for which you want to define the marker click the right mouse button and select one of the Set marker peak menu options in the popup window Chapter 10 11 6 Labelling Peaks in the Graph The annotation or any other column in the peak list window can be displayed as a label in the graphics window To do this select the peak data column to be used as label using the File Customise gt Plotting peaks menu option in the Data Set Graphics window Chapter 10 11 7 Columns in Peak List Windows Peak list windows have a number of header lines that list some details on the data as well as noise levels The columns of data that can be displayed in the peak list window are listed below index peak index An S is appended for shouldered peaks chapter Chapter 10 10 4 an s is appended for shoulder peaks norm is appended for normalisation peaks chapter Chapter 10 10 3 ref is appended for reference peaks chapter Chapter 10 10 4 Inactive peaks do not display an index number Inactive peaks are not included in total area and area fraction calculations begin time coordinate at start of peak top time coordinate at highest point or lowest point for negative peaks end time coordinate at end of peak height vertical coordinate at peak top minus baseline value annotation short text describing the peak cf chapter Chapter 10 11 4 name name of the peak component Used in calibrations Names that have been entered b
123. easured 00 00 2000 00 00 00 Flow Rate Adjustment Curve Vertical Moving a Curve Vertical soa a Description Gradient sae Points 3000 jaooo Freq Hz po le Offset s 10 joo Ordinate otw Total Conc fo Inert s joo Unit Offset pooo Dilution fo Flow min 3 Unit Factor 6 97384 Copy DAx 7 1 24 08 2002 14 56 50 PP Print Read data from disk file E DAxSeq TEST1 DAX DAx 7 1 24 08 2002 14 57 25 PP DAXSIM1 PRN v Help 1 The measuring item is checked if the measurement is still running In this case the points number of points item in the info dialog will display 2 The AutoAnalysis item is checked if the measurement is waiting to be AutoAnalysed 3 The modified item is checked if the file has been modified since it was last saved In this case the disk file that is denoted in the disk file item will not be up to date If the user tries to close the data window he will need to confirm that data will be lost Data Set Action List see chapter Chapter 8 11 The Description button lets you change the description Number of points is for running measurements Click the Add button to add remarks to the log file The Copy and Print buttons will copy the information dialog contents to the clipboard write them to file or print them Only the information is written not the actual data When one of these buttons is pressed you will first be presented with a small dialog box that le
124. easurement the data in the row were derived from Chapter 15 6 2 Calibration Curve window The Calibration Curve window has the usual graphics window functionality chapter Chapter 1 When the mouse cursor is moved over a calibration point in the graphics window the corresponding line in the text window will be highlighted if a GPC list window is open Double click the mouse on a GPC calibration point to include or remove the point from the GPC calibration The View Linear and View Logarithmic menu options may be used to switch between linear and logarithmic molecular weight axes Chapter 15 6 3 When the calibration changes Whenever the GPC calibration is changed either by removing lines from the GPC calibration list or by double clicking points in the GPC calibration curve the following steps are taken e A new calibration will be calculated e The molecular weight data in peak list windows chapter Chapter 10 11 will be recalculated e Any graphics windows with Molecular Weight Axes chapter Chapter 15 3 will be re plotted 167 Chapter 15 7 Local Calibrations f Local GPC Calibrations are enabled using the File Customise gt Extensions menu option each data set can store and use its own GPC calibration This makes it possible to re analyse data each time using the calibration that pertains to that specific data set lf a data set does not have a local GPC calibration the current main GPC calibration will b
125. ectable vertical coordinates Chapter 13 1 Remark Tacks Tacks can be added moved and deleted e To add a tack double click at the point in the graph where you want to add it then select Add tack from the popup menu For running measurements you can also click on the tack symbol at the left side of the data tag This will add a tack at the current measurement time e To move a tack place the mouse cursor over it the mouse cursor will change to a tack shape click the mouse and drag the tack to a new position e To delete a tack place the mouse cursor over the tack and click the right mouse button A dialog box appears Use the Delete button to delete the tack e To change the text for a tack place the mouse cursor over the tack and click the right mouse button A dialog box appears Now enter the new tack description Tack changes can be undone using the Tacks menu option in the data tag menu chapter Chapter 8 Tacks are listed in peak list windows chapter Chapter 10 11 as well as in data list windows The peak annotation needs to be shown for tack texts to be displayed in a peak list window Use the View Configure menu option to make the peak annotation column display Tack texts can be edited in the peak list window and tacks can also be removed there by deleting lines 151 Chapter 13 2 Labels DAx has the ability to add labels to a data set Labels are used to add texts to data Unlike tacks labels are not dis
126. ed LEKO4 TR819 ROX data sets OBB LEKO4 TRS19 ROX i 4 LEKO4 TR819 6FAM z _ i CBs LEKO4 TR819 6FAM CR LEK04 TRe09 ROx OFJ veko4 TRe09 Rox i OE LEKO4 TR809 6FAM DE LEK04 TRe09 6FAM Leko4 TRe10 R0x LEKO4 TR810 6FAM LEKO4 TR810 6FAM Y b Custom 2l MV Make baselines lighter custom colours only O oe LEKO4 TR810 ROX O Select all fka Deselect ue 1 Click one of the 12 colour buttons to assign a predefined colour to all selected checked data sets Colours are predefined using the View Attributes menu option 2 Click the Custom button to choose any colour to assign to the selected data sets Check the Make baselines lighter box to assign lighter whiter colours to baselines and blank measurements The Select all button can be used to select all data sets that contain a given string of characters in their names Enter the string or choose one of the pre defined strings listed Check deselect to deselect all data sets that contain the string If the Alt key is pressed when the Select all button is clicked all previous selection will be removed before the new selection is applied Chapter 9 4 Data Reduce This command invokes a dialog box that allows the number of points in a measurement to be reduced by averaging them or by yanking them An end time may also be set any data points beyond this time are removed from the data set 92 e f data are
127. ed data sets Chapter 17 6 1 Local and Global Southern Calibrations The local and global Southern calibration curve types use this relationship between parameter value m and calibrated value L17 PSs 41 m Mpo C Mo and Lo are constants In a global Southern calibration these three constants are derived by least squares fitting all calibration points at once In local Southern calibrations each set of three points is fitted assigning the constants to the middle point For a parameter value lying between points p and q the calibrated value is then calculated as the average of the values based on the calibration around point p and the calibration around point q 17 Typically m is a mobility and L a DNA fragment length 180 Furthermore for parameter values smaller than the lowest calibration point a straight line is drawn between 0 0 and the first calibration point For parameter values smaller than the second calibration point or larger than the penultimate calibration point only the calibration around the second or around the penultimate point is used Chapter 17 7 Automatic Calibration Derivation By checking the Derive automatically menu option in the data tag menu chapter Chapter 8 whenever a peak is changed in the data set a new calibration will be derived The new calibration will be copied to other data sets in the same enslavement group At most one data set in an enslavement group can have this option active
128. een begin and end of the new peak double click while holding down both the Shift key and the Ctrl key to add a spline node chapter Chapter 8 7 Splines must have already been added to the data set Chapter 7 7 Data Set Sizing Techniques DAx contains a number of techniques to change the time and voltage coordinates of a data set There are five techniques time displacements can be entered in the measurement set up dialog see Measurement Window manual This is used mainly to allow for a slight time difference before a sample reaches a detector a data graph may be dragged or sized in a graphics window A bounding rectangle can be activated around a data set by means of the data tag menu chapter Chapter 8 2 or by double clicking the left mouse button on the graph and selecting Sizing Controls from the popup menu The sides or corners of the bounding rectangle can then be dragged to size the data set or the entire bounding rectangle can be dragged to move the data set a sizing dialog box may be invoked from the data tag sizing menu chapter Chapter 8 4 This dialog box is also used to enter flow rates and inert times i e retention times of inert components used to calculate capacity factors A total concentration can be entered which is used to calculate relative concentrations Finally a dilution can be entered data sets may be overlaid and adjusted using the Data Overlay menu option chapter Chapter 9 16 in the graph
129. eft peak border and the peak top In the example of peaks being 1 base pair wide this would be 0 5 BP unit right peak width is the normal distance between the right peak border and the peak top In the example of peaks being 1 base pair wide this is also 0 5 BP unit height limit expressed as a percentage of the signal difference between a peak border being adjusted and the peak top The signal value at a proposed adjusted peak border must not be higher than this limit This limit is used to prevent a peak that is wider than expected but that has a consistently high signal value from being made narrower You should normally use a value of around 10 here A value of 0 prevents the peak from being adjusted A value of 100 forces every single peak to be adjusted regardless of its shape Chapter 10 4 10 Removing Bleedthrough Cross over Peaks DAx has the ability to remove bleed through peaks also known as cross over peaks 121 P Find Peaks lx Ois CAWN SETUP DAX test3en4 dal Remove bleed through peaks STAND 1 Da1 Remove contained peaks STAND 1 Da1 IV Entirely contained peaks for which 5158 4 Da1 related data set must 5158 4 Da1 have signal bleed through signal ratio gt 2 00 Al Remove asymmetric peaks for which Right left width gt 4 00 Al Left right width gt 4 00 Al JV Related data set must have signal bleed through signal ratio gt 0 75 Select all fka
130. eing used aseline data sets are not displayed Chapter 7 8 6 View Separate lanes Chapter 7 8 7 View Group per lane 65 These options choose between displaying each data set in itsown lane and displaying an entire enslavement group in a single lane Chapter 7 8 8 View Calibrated axis This otion causes the colour view to be displayed with a calibrated horizontal axis rather than the same horizontal axis as the source graphic window This can be used to display a colour view with a calibrated value axis instead of a time axis without having to convert the horizontal axis of the source data Chapter 7 8 9 View Mark peaks Causes peaks to be marked with triangles If user defined bins are used double clicking on a peak marker that is not inside an existing binning sheet bin causes a bin to be added Double clicking inside a binning sheet bin adds or removes a peak Chapter 7 8 10 View Mark bins Do not mark bins Chapter 7 8 11 View Mark bins Mark above Chapter 7 8 12 View Mark bins Mark above and inside Causes binning sheet bins to be marked above the color view or above the colour view and inside it using dotted lines Bins will only be marked if the binning sheet s qualification coordinate is compatible with the horizontal coordinate of the colour view both must be a time coordinate or both must be calibrated values If user defined bins are used and bins are marked inside the colour view a bin can be added
131. en keeping and not keeping the analysis log Chapter 18 14 2 Analysis Analysis log Configure menu option This menu option is used to choose the file name for the analysis log select the parameters to log for each data set choose which peaks to log choose when to log and in what format Chapter 18 14 3 Analysis Analysis log View menu option This menu option is used to view the current analysis log Chapter 18 15 Analysis Assays It is possible to define a number of conditions that an analysis must meet to be of sufficient quality This ability is particularly useful when large amounts of data are being analysed automatically Rather than having to look at each measurement individually the analysis assay can be used to find analyses that are of bad or suspect quality There are four categories of parameters used to determine the quality of a measurement Each is listed below with a more detailed explanation of the individual parameters Some versions of DAx have a fifth category 224 All parameters have values for suspect and bad analyses If any individual analysis parameter is found to be bad the entire analysis is bad If no parameters are bad but one or more are suspect the entire analysis is suspect The contributions for all parameters can be weighted Refer to chapter Chapter 18 15 1 for details All parameters can be used or disabled To set up the analysis assay use the Analysis Edit assay paramete
132. ent CE capillary parameters and the electro osmotic flow derived from the reference peak Based on the mobilities of OH and H DAx imposes scale limits on mobility axes These limits are e for effective mobilities 250 and 400 10 m7 Vs 171 e for apparent mobilities 0 and 700 10 m Vs Chapter 16 5 1 Converting to a Mobility Axis Some versions of DAx have the option of converting the horizontal axis of a measurement to apparent mobility values instead of time coordinates using the CE Axis conversion menu option You will be asked to specify the lower and upper limits of the converted axis as well as the frequency number of points per unit mobility to be used If the mobilities present in the data do not extend far enough to cover either limit the limits will be adjusted Chapter 16 5 2 Mobility Axis Options There are several ways in which data can be plotted with a mobility weight axis All of these are selected using View menu options e Either apparent or effective mobility can be plotted In order for effective mobilities to be plotted a reference peak chapter Chapter 10 10 3 must have been defined and an effective mobility for the reference peak s component must have been entered as an annotation for the reference peak e Either the concentration as detector signal or the migration time corrected concentration as detector signal divided by migration time of each component can be plotted along the ver
133. ent has a skew that exceeds this parameter the analysis becomes suspect or bad The absolute value of the skew is used 226 e maximum allowable area of unrecognised peaks If an identification database is used to recognise name peaks in the measurement this parameter can be used to detect measurements where unrecognised peaks have an area that is too large This parameter can be entered as an absolute or relative value The relative value takes the total area of unrecognised peaks and divides it by the total area of all peaks e low high limit of ratio of areas of two named peaks Typically these limits will be used in analyses where there is one clear substrate peak and one product peak If the area of the product peak is too small compared to the area of the substrate peak the analysis is flagged Another application would be to make sure that the area of a known impurity is not too high compared to the area of the product peak e maximum allowable number of unrecognised peaks If an identification database is used to recognise name peaks in the measurement this parameter can be used to detect measurements that have too many unrecognised peaks Chapter 18 15 4 Required peaks A list of names of peaks that must be present can be entered A quantification parameter is selected height area concentration area fraction If a peak with the specified name is not present or if its quantification parameter is lower than the spe
134. enu by clicking on the data tag for the data set you want to process cf chapter Chapter 8 2 The data will be analysed and a graph such as the one displayed below will be displayed You can now use the left slider control discussed above in chapter Chapter 11 1 to set the Fourier filter width to an optimum value Y s n 40 20 log Fourier width Fourier filter characteristics plot The signal to noise ratio is a dotted line The solid lines are the peak height and noise level The filter width along the horizontal axis is plotted logarithmically 146 Average Shift 0 000 0 005 4 0 010 T T T T T T T T T T T T T 0 1 2 log Fourier Period Another Fourier filter characteristics plot The shift in the average signal value as a function of the Fourier filter width is plotted Initially the average signal value barely changes To remove noise a filter width where there is no significant change in average signal value should be used Chapter 11 3 Savitzky Golay and Moving Average Filters Refer to chapter Chapter 9 5 for details on Savitzky Golay and Moving Average filters Chapter 11 4 The Spike Removal Algorithm The spike removal algorithm considers each point in the data set to determine if it is a spike To do this it calculates the following values e data value of point preceding the current point Y1 e data value of point following the current point Y e intersection of the
135. er limits of the converted axis aS well as the frequency the number of data points per calibrated unit to be used If the calibration does not extend far enough to cover either limit the limits will be adjusted Chapter 17 5 1 Calibrated Axis Options There are several ways in which data can be plotted with a calibrated axis They are selected using View menu options e Either a linear or logarithmic scale can be used along the horizontal axis Chapter 17 6 Calibration Dialog Box This dialog box can be used to derive a calibration from a number of data sets or to set calibration parameters The dialog is invoked using the Calibration Calibrate menu option If a calibration is being derived from data sets the data sets must have peaks with the appropriate calibrated values as their annotations Refer to chapter Chapter 17 2 for step by step instructions on the steps that should be taken before this dialog box is invoked The calibration dialog box is used for a number of purposes e creating the initial calibration list e setting the calibration model straight lines cubic spline or polynomial All of these models can act either on linear or logarithmic scales as indicated in the dialog box e setting upper and lower limits for calibration parameters that can be converted to a calibrated value If the calibration model is spline calibration parameter values outside the calibration parameter range of the calibration t
136. erting to a Molecular Weight axis DAx has the option of converting the horizontal axis of a measurement to molecular weights instead of time coordinates Use the GPC Axis conversion menu option for this You will be asked to specify the lower and upper limits of the converted axis as well as the frequency that is the number of data points per unit Mw to be used If the calibration does not extend far enough to cover either limit the limits will be adjusted Chapter 15 4 3 Converting between Concentration and Weight Detectors measure component concentration or component weight As a resul measurement data express either 163 concentration or weight The menu option GPC Concentration weight conversion allows data to be converted This applies to data with a molecular weight axis For these data the vertical coordinates will change For other data the concentration weight flag will be changed in the GPC calibration As an example consider two samples being measured The first sample is 0 001m in component A which has a molecular weight of 1000 The second sample is 0 001m in component B which has a molecular weight of 10000 A detector that measures concentration will give the same signal strength for samples A and B A detector that measures weight will give a signal that is ten times higher for component B The distinction between detector types is important because it determines how the data will be converted to
137. es 30 183 Menu Analysis Analysis log Configure 224 Analysis Analysis log Keep 224 Analysis Analysis log View 224 Analysis Binning sheet 207 Analysis Clear binning sheet 219 Analysis Comparison sheet 204 Analysis Configure meta analysis 221 Analysis Edit assay parameters 228 Analysis Edit ATC 184 Analysis Edit meta analysis 221 Analysis Global sheet 204 Analysis Load assay parameters 229 Analysis Load ATC parameters 187 Analysis Load binning sheet 219 Analysis Load database 191 Analysis Marker peaks 194 Analysis Perform ATC automatically 187 Analysis Qualify peaks 192 Analysis Reference peak 194 198 Analysis Save assay parameters 229 Analysis Save ATC parameters 187 Analysis Save binning sheet 219 Analysis Save database 191 Analysis Stop analysis 191 Analysis Stop ATC 187 Analysis Stop quality assays 229 Analysis Use 221 Calibration Axis 178 Calibration Calibrated axes 177 Calibration Calibration curve 181 Calibration Calibration list 181 meta analysis conversion Calibration Load calibration 177 Calibration Save calibration 177 Calibration Stop calibration 177 Calibration Track value 177 CE Apparent mobility 171 CE Axis conversion 172 CE Effective mobility 171 CE Load calibration 170 CE Mobility axis 171 CE Save calibration 170 CE Time cor
138. es of species of bacteria Depending on the presence of certain peaks in one or more of the traces a likelihood of presence of each of the bacteria species can be determined Meta analysis entries consist of the following items e identity of species multiple meta analysis items are likely to have the same identity so that multiple peaks can be used to determine if the species is present e peak name data sets are checked for the presence absence of at least one peak with the specified name e wanted unwanted If a wanted peak is absent the likelihood of the presence of the species decreases if it is present the likelihood increases Conversely the presence of unwanted peaks decreases the likelihood of the species being present e absolute relative if a wanted absolute peak is missing identification fails If it is present likelihood is put to 100 If a wanted relative peak is present identification gains likelihood by the amount specified in the weight column If an unwanted absolute peak is present identification fails If an unwanted relative peak is present identification loses likelihood by weight Relative peaks that are not present do not influence likelihood 220 e weight used to adjust the overall likelihood of the presence of the species for relative peaks e quantifier limit peaks that have a value of the selected quantifier below the specified limit are considered absent This is used to filter out
139. es the reference peak of the data set Marker peaks are allocated in a data set before normal peak identification is used to recognise components Marker peaks can then be used to recognise components using their location relative to the marker peaks Marker peaks will typically be internal or external standards Index Marker Name Refpeak Peak type A OnAxes InTraces Qualify by Using marker Search Coor Start End Coor Tolera i ja Yes lt no type gt lt all gt ROX Peak Top Coordinate Highest in interval 1500 0 4500 0 2 100bp No lt no type gt lt all gt ROX Offset to marker 20 bp Highest in interval 400 0 1060 0 3 159 bp No lt no type gt lt all gt ROX Offset to marker 100 bp First in interval 350 0 800 0 4 221 bp No lt no type gt lt all gt ROX Offset to marker 100 bp Highest in interval 850 0 1750 0 5 288 bp No lt no type gt lt all gt ROX Offset to marker 221 bp First in interval 550 0 1300 0 6 318bp No lt no type gt lt all gt ROX Offset to marker 288 bp Highest in interval 250 0 700 0 7 350 bp No lt no type gt lt all gt ROX Offset to marker 318 bp Highest in interval 250 0 750 0 8 403 bp No lt no type gt lt all gt ROX Offset to marker 350 bp First in interval 500 0 1500 0 9 459 bp No lt no type gt lt all gt ROX Offset to marker 403 bp First in interval 500 0 1500 0 10 498 bp No lt no type gt lt all gt ROX Offset to marker 459 bp First in interval 300 0 1000 0 11 551 bp No lt no ty
140. etected they will be marked in the graph Check Print Info amp Logfile to print data set information and log files If the Close Data Windows after Analysis option is checked data sets will be removed after automatic analysis has been performed Data will be saved first Data sets will only be removed when all data sets in an enslavement group have been analysed If no data sets remain in the window the window will be closed Automatic analysis can derive a new GPC calibration from the analysed data Molecular weight values will be taken from peak names so the Identification Database must contain component names that contain Mw values The analysis will be set up to have the same form as the currently present analysis which defaults to multi linear i e lines drawn between calibration points using logarithms of Molecular Weight After the GPC calibration has been derived the horizontal axis can be converted to Mw values Click on Convert axis to Mw to configure the conversion process Automatic analysis can derive a new Calibration from the analysed data Calibrated values will be taken from peak names 51 so the Identification Database must contain component names that contain calibrated values The analysis will be set up to have the same form as the currently present analysis which defaults to multi linear i e lines drawn between calibration points not using logarithms After the calibration has been derived the horizon
141. eter page each bin can be assigned an individual lower or higher threshold by entering a value in the threshold column in the binning sheet When the mouse is double clicked on an empty bin cell and user defined bins are used a peak will be manually added29 You can indicate that manually added peaks should always be included in the bin even if they do not exceed the threshold 20 If user defined bins are used you can also manually add a peak by double clicking on a colour view 214 Bin annotations On the Bin annotations parameter page bin annotations can be defined consisting of a peak annotation a peak type or both The bin annotations are added to the popup menu that appears when the right mouse button is clicked on a binning sheet When a bin annotation is applied to a peak the peak s curve sample in the binning sheet will be displayed with the selected background colour Press the shift key when adding a bin annotation to a peak to also mark the peak and data set for attention Persistent data The Persistent data parameter page contains a Preserve data when closing windows checkbox Checking this box causes DAx to retain the peak data of data sets included in the binning sheet when a data graph window is closed If present baselines are subtracted from the data first the baselines themselves are not preserved The Restore selected to window button moves selected preserved data back into a new graph window The Re
142. even when problems such as the ones described above occur Not all versions of DAx support Automatic Trace Calibrations Chapter 17 10 1 Setting up the Automatic Trace Calibration parameters The ATC parameters are entered into the ATC parameter dialog box which is invoked using the Analysis Edit ATC menu option Automatic Trace Calibration 2 xi Trace type amp fragment sizes sizes Restrictions Calibration Advanced Calibration trace figi J f1 Horizontal axis type m DNA BFP 2 Calibration sizes a eee f i zi Cut Paste Add line 1 the Calibration trace defines the trace that contains the sizing standard in this instance LIZ Whenever a LIZ trace is analysed the ATC settings will now be used to look for the sizing standard 2 The Horizontal axis type limits the types of data the ATC algorithm will be applied to Since ATC is generally used to convert to a DNA axis it makes sense to exclude DNA axes from ATC 3 The Calibration sizes are simply a list of the sizes of the fragments in the sizing standard Annotations can also be entered which will be assigned to peaks 184 Automatic Trace Calibration GeneScan 120 LIZ GeneScan 350 ROX GeneScan 350 ROX no 250bp GeneScan 400HD ROX GeneScan 500 ROX GeneScan 500 ROX no 250 340bp GeneScan 500 LIZ GeneScan 500 LIZ no 250 340bp 1 A number of often used calibration standards are listed here
143. fixating peaks 70 75 flagging low and high concentrations 193 198 flow rate 24 78 86 161 Foley Dorsey 137 Fourier 61 85 86 100 108 143 144 Fourier filter 94 Fourier menu 72 frequency 24 61 108 Gaussian curve 101 Gaussians 123 GC 15 155 158 Gel Permeation Chromatography See GPC gel like display 62 65 generating data 101 Genescan files 30 183 global Southern method 180 GLP 16 17 24 56 86 89 110 definition 23 Good Laboratory Practice 16 Good Laboratory Practise 23 GPC 15 37 41 134 161 calibration curve window 167 calibration list window 166 creating calibrations 161 GPC windows 166 local GPC calibrations 168 GPC calibration 51 GPC Calibration 164 GPC calibration window 21 37 gradient 86 gradient corrected signal axes 158 gradient corrections See HPLC gradient nodes 157 gradient percentages 156 157 159 gradients constructing baselines 110 graphics windows calibrated axis windows 37 mobility axis windows 37 molecular weight axis windows 37 time corrected signal axis windows 37 volume axis windows 37 252 graphics windows calibrated axis windows 21 data set graphics windows 20 37 GPC calibration curve 21 gradient corrected windows 21 mobility axis windows 21 molecular weight axis windows 21 time corrected signal axis windows 21 volume axis windows 21 graphics windows data set graphics windows 47 g
144. fset point index frequency 2 Measurement values are calculated as value voltage factor voltage voltage offset 3 The unit that is used along the horizontal axis can be changed here The horizontal axis unit is always the same for a set of enslaved data sets cf chapter Chapter 3 3 9 Changing the unit will change the unit for all these data sets Timing settings frequency and time offset will be kept identical for enslaved data sets when the horizontal axis unit is changed The dialog box is also used to enter inert times i e retention times of inert components used to calculate capacity factors All inert times for a set of enslaved data cf chapter Chapter 3 3 9 are always the same If the inert time is 0 0 it will not be used The dialog box also allows the flow rate to be entered The flow rate is used to calculate elution volumes in chromatography as elution volume time coordinate flow rate All flow rates for a set of enslaved data sets are always the same If the flow rate is changed any graphics windows with Volume Axes chapter Chapter 15 3 will be re plotted Since Molecular Weights are calculated from volume data as well any graphics windows with Molecular Weight Axes chapter Chapter 15 3 will also be re plotted GPC extensions need to be enabled using the File Customise gt Extensions menu option to be able to enter flow rates 78 A total concentration may be entered for the sample The tot
145. fying parameter For instance mobility cannot be used to find peaks in HPLC measurements Concentrations can be assigned based on a number of possible quantifying peak parameters Possible quantifying properties are e peak top value peak height e peak area e normalised peak area using a normalisation peak cf chapter Chapter 10 10 3 e migration time corrected peak area Summation of signal values divided by time coordinates e normalised migration time corrected peak area Depending on the measurement data type of a data set some properties may not be available for use as a quantifying parameter For instance migration time corrected areas are available only for CE and generic measurements Note that if no normalisation peak is present DAx will use the reference peak to calculate normalised areas If no reference peak is present the largest peak is used Chapter 18 1 Creating an Identification Database If the menu option Analysis is not present you need to use File Customise gt Extensions and check Analysis Options Standard analysis should be unchecked To create a calibration 1 measure one or more samples with known components 2 construct a baseline refer to chapter Chapter 10 1 for details 3 detect peaks refer to the chapter Chapter 10 4 for details 190 4 use the data tag peak menu to display a peak list chapter Chapter 10 11 for each of the measurements you want to include in the calibration
146. g analysed 196 Some versions of DAx let you limit the trace types that certain markers can be recognised in The In Trace item is used for this purpose It is possible to automatically exclude marker peaks from area calculations by checking the Exclude column check box This is intended to be used to automatically exclude the EOF electro osmotic flow peak from area calculations in CE or generally to exclude external standards from area and concentration calculations Chapter 18 6 1 Using Existing Peaks The Copy Peak Data button is used to copy data from an existing peak into the marker peaks list Using this button filling the marker peaks list becomes much easier It also copies values for all or most qualifying coordinates If you later change the coordinate being used values will be present The procedure is as follows e load a data set e analyse the data set e name the marker peaks in the data set e use the Analysis Marker Peaks menu option to invoke the marker peaks setup dialog box use the Add Line button to add an empty line e use the Copy Peak Data button to copy name and coordinates of an existing peak onto the new line User defined bins chapter Chapter 18 12 3 can be copied and pasted into the marker peaks list Chapter 18 6 2 Using external marker peaks Normally when peaks are allocated relative to marker peaks the marker peaks need to be present in the current data set By checking the Use
147. h Close Help Second page of the peak find wizard Sets up finding peaks using threshold values 1 These controls select the unit to be used for peak detection Values for thresholds and slopes are either in fraction units or in vertical axis units thresholds and slopes always use the same units Fraction units range from 0 for the lowest signal value in the data set to 1 for the highest signal value Vertical axis units are typically volts Ois C win SETUP DAX test3 dal I Detect using Slopes STAND 1 Da1 E STAND 1 Da1 I Automatic slope ra m 5156 4 Da1 IV Slope requires threshold 4 5158 4 Da1 Positive slope limit fo 05 J I Link Positive amp Negative Negative slope limit 0 05 Z Units Threshold amp Slope C Ordinate Fraction l2 4 Select all fka I Deselect lt Prev Next gt Finish Close Help Third page of the peak find wizard Sets up finding peaks using slopes 114 1 Slope requires threshold requires that the signal reaches above the threshold value for a peak to be recognised when the slope exceeds the slope limit Use this option to be able to separate peak clusters where the signal value does not drop below the threshold value in between peaks These controls select the unit to be used for peak detection Values for thresholds and slopes are either in fraction units or in vertical axis units thresholds and slopes
148. h data along the time axis and move and stretch data along the vertical axis in such a way that they may be more readily compared The data must be in the same graphics window if you want to overlay data from different windows first drag the data sets chapter Chapter 8 to a single graphics window Possible horizontal coordinate adjustments e No adjustment e Coordinates will be adjusted in such a way that all reference peaks chapter Chapter 3 3 15 for the various data sets will have the same top coordinate as the reference peak in the first selected data set e Coordinates for all data sets will be adjusted in such a way that all reference peaks will have a specified target reference peak coordinate It is possible to use the top begin or end of the reference peaks This is useful in applications such as ion chromatography 102 In the above two options data can be moved or stretched in horizontal direction Moving alters the coordinate of the first point but leaves the frequency unaltered Stretching alters the frequency but not the coordinate of the first point If no reference peak is present the normalisation peak will be used If there is only a single peak that peak will be used regardless of whether or not it has been defined as a reference and or normalisation peak The peak used will be marked as a user defined reference peak that way it will not be replaced by an automatically assigned reference peak cf chap
149. h skim value i e the ratio of the total peak area to the area of the part of the peak that lies above a line connecting peak begin and peak end is high e Low width If the width of the widest peak in a cluster of peaks divided by the width of a certain peak exceeds a given limit the narrow peak is possibly a shoulder peak e Low area If the area of the largest peak in a cluster of peaks divided by the area of a certain peak exceeds a given limit the small peak is possibly a shoulder peak Border values for the degree of slant skim value and width and area ratios can be entered to determine which peaks DAx will mark as shoulder peaks cf chapter Chapter 10 4 1 Chapter 10 10 8 Shoulder Peak Area Calculations Normally shoulder peaks are skimmed cf the example graph in the previous section peak begin and end of the shoulder peaks are connected to form a local baseline for the shoulder peak However in the Peak Find item in the File Customise menu option you can indicate that shoulder peaks should be sliced as demonstrated below It is also possible to indicate that the area of the shoulder peak should be included in the area of the shouldered peak 131 DAx 1 0 PP 19 01 96 Volt 0 20 0 15 0 10 K time s Example of shoulder peaks lying on shouldered peak The shoulder peaks have been sliced instead of skimmed Chapter 10 10 9 Connecting and Combining Peaks Peaks can be connected i e
150. hapter 12 The Waste Barrel 149 Chapter 12 1 The Waste Barrel Pop up Menu 149 Chapter 12 1 1 Waste barrel cursor 149 Chapter 12 1 2 Restore items 149 Chapter 12 1 3 Destroy contents 150 Chapter 12 2 Moving the Waste Barrel 150 Chapter 13 Remark Tacks and Labels 151 Chapter 13 1 Remark Tacks 151 Chapter 13 2 Labels 152 Chapter 14 DAx HPLC GC HPLC Gradient amp GC Temperature Programme Extensions 155 Chapter 14 1 Defining Gradients step by step 155 Chapter 14 2 HPLC GC Gradient Parameters Dialog Box 155 Chapter 14 3 HPLC GC Load gradient HPLC GC Save gradient menu options 156 Chapter 14 4 HPLC GC Subtract Gradient menu option Chapter 14 5 View Gradient percentages View Temperature programme 157 Chapter 14 6 HPLC GC Show Gradient Nodes Chapter 14 7 Gradient Corrected Signal Axes Chapter 14 8 Modifying Gradients Chapter 14 8 1 Modifying Gradient Percentages Programme Temperatures 159 Chapter 15 DAx GPC Gel Permeation Chromatography Extensions Chapter 15 1 Creating a GPC calibration Chapter 15 2 GPC Load calibration GPC Save calibration GPC Stop calibration menu options Chapter 15 3 GPC Track Mw menu option Chapter 15 4 Elution Volume Axes Molecular Weight Axes __ Chapter 15 4 1 Molecular Weight Axis Options Chapter 15 4 2 Converting to a Molecular Weight axis Chapter 15 4 3 Converting between Concentration and Weight Chapter 15 5 GPC Calibration Dialog Box Chapter 15 6 GPC
151. hat have been entered Chapter 14 5 View Gradient percentages View Temperature programme These commands determine if HPLC gradient percentages or GC programme temperatures will be plotted They are either shown for all curves or hidden for all curves they cannot be shown and hidden individually Use the HPLC GC Gradient Parameters dialog box chapter Chapter 14 2 to enter percentage values for the gradient steps Chapter 14 6 HPLC GC Show Gradient Nodes This command determines if HPLC gradient percentage nodes or GC programme temperature nodes will be plotted By clicking and dragging these nodes the gradient or temperature programmes can be altered 157 Chapter 14 7 Gradient Corrected Signal Axes The menu option HPLC GC Gradient corrected axis displays graphs where the signal gradient has been subtracted from the signal values Gradient percentages or programme temperatures may also be displayed using the HPLC GC Show percentages or HPLC GC Show temperature programme menu options Chapter 14 8 Modifying Gradients The data tag menu contains several options that pertain to gradients cf chapter Chapter 8 2 Use the Show percentages and Show temperature programme menu options to switch between displaying gradients and hiding them NB This option does not affect the presence of the gradient in memory it merely determines whether or not the gradient is drawn Gradients are marked by little marker
152. he highlighted peaks is output in the status bar at the bottom of the DAx window whenever the highlighting changes Chapter 10 11 2 Deleting Peaks from the Peak List Window It is possible to remove peaks from the peak list window Click the right mouse button on the line listing the peak that should be removed and click the Delete peak menu option To remove several peaks at once highlight them chapter Chapter 6 1 1 then use the Delete selected menu option in the pop up menu Chapter 10 11 3 Locating or Highlighting Peaks To locate or highlight a peak in the graphics window click the right mouse button in the peak list window then select Locate peak or Highlight peak in the pop up menu that appears Press the Shift key to hide all other data sets To highlight several peaks at once use the Highlight selected menu option Use the View Remove highlight menu option in the graphics window to remove the highlight 133 Chapter 10 11 4 Peak Annotations Each peak can have an annotation that is a short text that describes the peak In CE the annotation for the reference peak should contain the effective mobility for the peak in 10 m Vs This will then be used to calculate effective mobilities from the apparent mobilities of all other peaks To set up GPC calibrations chapter Chapter 15 1 the annotation needs to be a numeric value indicating the molecular weight For Calibrations chapter Chapter 17 2 the annotation need
153. he range between these two is used either as an absolute measure or relative to the first derivative determined as above minimum required number of calibration points If fewer than the set limit calibration points are available the quality of the calibration curve becomes suspect low high limit on available calibration points If the curve starts at an unexpectedly high coordinate or ends at an unexpectedly low coordinate the quality is flagged as suspect or bad Chapter 18 15 7 Analysis Edit assay parameters Analysis Edit assay parameters is used to enter or modify a set of assay parameters 228 Chapter 18 15 8 Analysis Load assay parameters Analysis Save assay parameters Analysis Stop quality assays Assay parameters may be saved to disk using the Analysis Save assay parameters option They may be loaded from disk using the Analysis Load assay parameters option If the assay parameters have been changed but have not been saved to disk yet when DAx is about to terminate you will be asked to confirm the loss of the parameters Saving the parameters to a floppy disk does not prevent this you must save to hard disk Loading a new set of assay parameters may cause an existing analysis assay to be changed whenever peaks are recalculated This can be prevented by using the data tag peak menu fixate option Data that have been loaded from disk will also automatically have their analysis assay fixated Analy
154. hidden 3 Use the Show gradient menu option to show or hide gradient lines These lines depict signal values not gradient percentages or temperatures 82 Press the Shift key while executing this menu option to show or hide gradient lines for all data sets in a graphics window 4 Use the Mark gradient menu option to show or hide gradient markers at the start and end of each gradient step Press the Shift key while executing this menu option to show or hide gradient markers for all data sets in a graphics window 5 Use Copy gradient to main gradient to copy the gradient settings from the current data set to the main gradient and selected other data sets 6 Use Undo and Redo to undo and redo changes that were applied to the gradient Chapter 8 9 Data Tag Tack Menu The Add Tack menu option adds a remark tack at the end of the data in the data set Double clicking at the desired location for the remark tack is the preferred method to add a tack Undo Tack and Redo Tack are used to undo and redo the last modifications to remark tacks For more information about data tacks refer to chapter Chapter 13 Chapter 8 10 Data Set Information Dialog This dialog box is displayed when the Info menu option in a data tag is executed 83 Data Information 2 x Data Set ERSS maik B l Measuring File JE DAxSeq TEST1 DAX l2 AutoAnalpsis File Date Time 27 03 1996 22 46 28 lv Modified Operator Generic Measurement 4 M
155. ics window arithmetic operations can be performed on either the time or voltage coordinates using the Data Arithmetic menu option chapter Chapter 9 9 in the graphics window 64 Chapter 7 8 Colour View Windows The View Colour view menu option displays the contents of the graphic window as a coloured map Colour views are convenient when large numbers of data sets need to be compared Colour view windows include the following menu options Chapter 7 8 1 View Extra space This option causes space to be left blank between lanes which can increase readability Chapter 7 8 2 View Use trace colours When this option is active each data set is depicted using its trace colour in the graph window The colour will range from the background colour of the colour view to the trace colour of the data set When trace colours are not used all values are mapped using the colours set using the View Attributes menu option Chapter 7 8 3 View Darker Causes darker colours to be used This is useful when most signal values are low with only occasional high valus The option causes low intensity values to be displayed three times darker Chapter 7 8 4 View Halftones Causes smooth colour transitions to be used Chapter 7 8 5 View Subtract baselines This option causes baseline to be sutracted before converting to colours This can make comparing signal strengths data with different baseline signal levels easier When this option is b
156. idth are fitted not the location of the peaks tops 4 When the fit is complete fitted Gaussians and the sum of the fitted Gaussians can be marked cf Chapter Chapter 8 3 When the fit is complete the fitted parameters for each peak can be displayed in the peak list windows of the data sets using the View Configure menu option The parameters are Fitted coordinate Fitted standard deviation Fitted height Fitted area and Fitted asymmetry factor Note that the fit can sometimes fail for certain peaks especially if many peaks overlap Not all versions of DAx contain this functionality Chapter 10 7 Peaks Clear analyses This command removes peaks and baselines for all data sets in the window Chapter 10 8 Peaks Un Fixate analyses These commands fixate or unfixate all data sets in the window see chapter Chapter 8 3 2 Chapter 10 9 Peaks Recalculate peaks Use this menu option to force peak parameters to be recalculated DAx normally recalculates all parameters as needed this menu option can be used to make doubly sure all values are up to date Chapter 10 10 Modifying Peaks Peaks are marked in graphics windows using small triangular markers aS can be seen in the example graph below Use the Show peaks menu option in the data tag peaks menu chapter Chapter 8 3 to stop displaying markers Use the 124 File Customise gt Plotting peaks option to stop displaying markers altogether This option will also
157. ied to all components They can later be changed for individual components Refer to chapter Chapter 18 8 for details amp Peak Identification amp Quantification 2 x Fetch components from Concentration limits ya Remarks STAND 1 Da1 Qualification Attributes Quantification STAND 1 Da1 NERE 5158 4 Da1 Qualification coordinate 5158 4 Da1 Peak Top Coordinate Moment Peak Coordinate Offset 4 Peak Begin Coordinate Peak End Coordinate Capacity factor m Tolerance fil A Absolute 2 IV Intermediate qualification pass C Relative Add to existing calibration OK Cancel Hep 1 Select the qualification parameter here The database will be sorted in ascending value of the qualification parameter 2 The Tolerance determines by how much a qualifying parameter for a peak may differ from the value in the Identification Database for the peak to be identified Tolerances may be absolute or relative 3 For detailed information about the multiplicity tab see the next chapter Chapter 18 5 193 Chapter 18 5 Multiplicity Normally DAx assigns at most a single identification to each peak and each item in the identification database is assigned to at most one peak the peak that best matches Some versions of DAx have the option to allow multiple identifications per peak If multiple identification database items best match a single peak the peak will receive a
158. if no noise level is Known yet Peaks must exceed the noise level in order to be detected f the threshold value that is being used is lower than the noise level peaks need only exceed the threshold level Chapter 10 4 1 The Peak Find Wizard To enter parameters telling DAx how to find peaks a wizard is used Ois C win SETUP DAx test3 dal Indicate if positive and or negative STAND 1 Da1 peaks should be found STAND 1 Da1 5 5158 4 Da1 v Positive Peaks j j i 5158 4 Dal IV Negative Peaks Ai A Indicate the minimum area for valid peaks Minimum area o o FA C Absolute Vs Percentage 4 Select all lt a I Deselect lt Prey Next gt Close Help First page of the peak find wizard The user can disable negative or positive peaks A minimum area for each peak can be entered This is a very powerful way to reduce the number of peaks found If it is known that an analysis procedure has an accuracy of 2 it is a good option to enter a minimum peak area of 2 or 1 for a margin of error 113 Ots C win SETUP DAx test3 dal JV Detect using threshold STAND 1 Da1 STAND 1 Da1 5158 4 Dal 5158 4 Dal1 fa 0 noise level Positive threshold fo Link Positive amp Negative Negative threshold 0 1 Al Units Threshold amp Slope e Ordinate Frcton 4 4 Select all fka I Deselect lt Prev Next gt Finis
159. if the calibration uses a polynomial e usage points are either included in or excluded from the calibration e concentration e quantifying parameter value one of the following e peak height e peak area e normalised peak area e migration time corrected peak area e normalised migration time corrected peak area e check value applicable only for polynomial calibrations The check value is the quantitative parameter value calculated for the concentration in the row using the current polynomial e data source name of the measurement the data in the row were derived from NB This may be unequal to the name displayed in the Identification Database window itself The data source displayed there indicates what data were used to set up qualitative calibration whereas the data source displayed in the quantitative calibration list indicates what the quantitative calibration is based on Use the Analysis Config quantification see chapter Chapter 18 8 menu option to change the type of calibration curve for the component being investigated Chapter 18 9 3 Quantitative Calibration Graph Window The Quantitative Calibration Curve window has the usual graphics window options When the mouse cursor is moved over a calibration point in the graphics window the corresponding line in the quantitative calibration list window will be highlighted Double click the mouse on a Calibration point to include or remove the point from the calibration 202 Th
160. ights as peak annotations molecular weights may also be entered as part of peak names This makes it possible to identify peaks using an 161 Identification Database and then set up a GPC calibration using automatically assigned peak names Chapter 15 2 GPC Load calibration GPC Save calibration GPC Stop calibration menu options GPC calibrations may be saved to disk using the GPC Save calibration option They may be loaded from disk using the GPC Load calibration option f the Separate Parameter Files item in the File Customise gt Extensions dialog box is unchecked GPC parameters can only be saved as part of an analysis procedure so the Load Save calibration menu options are not displayed If the GPC calibration has been changed but has not been saved to disk yet when DAx is about to terminate you will be asked to confirm the loss of the new calibration Saving the calibration to a floppy disk does not prevent this you must save to hard disk GPC Stop calibration stops using the calibration and removes it from memory Chapter 15 3 GPC Track Mw menu option Use the GPC Track Mw menu option to determine if molecular weight coordinates are displayed in the coordinate tracking bar when you move the mouse cursor across a graphics window Chapter 15 4 Elution Volume Axes Molecular Weight Axes The Data Set Graphics window menu options GPC Volume axis and GPC Mw axis display graphics where the time axi
161. in peak list windows Normalisation peaks are used in a number of ways e Data sets can be overlaid on their normalisation peaks e Normalised peak areas in peak list windows are calculated as peak area relative to the area of the normalisation peak If no normalisation peak is present the reference peak will be used If no reference peak is present the largest peak is used Chapter 10 10 4 Reference Peaks One peak in each measurement can be marked as the reference peak To mark a peak as a reference peak in the data set graphics window chapter Chapter 7 2 move the mouse cursor over the peak top then click the right mouse button The peak popup dialog box appears Use the Reference Peak check box to mark the peak as the reference peak this will unmark any existing reference peak Repeat the procedure to unmark the peak Peaks may also be marked as a reference peak in the peak list window chapter Chapter 10 11 Click the right mouse button on the peak you want to make reference peak and select Set reference peak from the pop up menu Reference peaks are listed with ref following the peak number in peak list windows 128 To learn how to automatically allocate Reference Peaks refer to chapter Chapter 18 7 Reference peaks are used in two ways e In chromatography the capacity factor k is calculated as k Tio with t the retention time of the component and t the 0 retention time of an inert component Peak to
162. ined or automatic bins OWNLOAD Utah dG01 19 15Lots2 da1 py Qualifying coordinate dG01 19_15 ROx Trace types dG01 19_15 6F4m a Wadia Jadia dG01 19_15 HEX eak type use and selection 3 Curve samples and peak parameters dao em Curve sample scaling 5 u dG01 23_16 ROX Quantifying parameter labels d amp 01 23_16 6FAM Peak parameter selection dG01 23_16 HEX Statistical items dG01 23 16 NED Display options d ma Peak thresholds um G02 08_15 ROX Bin annotations dG02 08_15 6FAM Persistent data dG amp 02 08_15 HEX dG02 08_15 NED MABE dc02 21_16 Rox Enter part of a data set name or enter a trace type then click Frey Next gt Select all to select all data sets that contain the search text or are of the specified trace type Select all fka Deselect 2 Check Deselect to deselect rather than select data sets Press the Cancel Apply Alt key when clicking Select all to remove all previous selections 1 Select which data sets to include in the binning sheet 2 The Select all item allows the quick selection of all data sets that meet certain criteria across all graphic windows For instance all data sets with ROX labels can be selected or deselected if the Deselect box is checked 208 Press the Alt key when clicking the Select All button to remove all previous selection User defined or automatic bins amp Binning Sheet Setup eix Setup parameters
163. irst To remove only spikes and perform no filtering select a Savitzky Golay filter with width 3 For details on the spike removal algorithm refer to chapter Chapter 11 4 The Drift Factor item determines which data points will be considered spikes A higher drift factor will make fewer points be considered spikes NB Clicking the Close button will cause the filter parameters to be updated for use in AutoAnalysis but no data will be filtered Chapter 9 9 Data Integrate If the menu option Data Integrate is not present you need to use File Customise gt Data and check Data integration This command invokes a dialog box that allows data to be integrated 96 Chapter 9 10 Data Arithmetic This command invokes a dialog box that allows data sets to be modified by performing either curve9 or constant arithmetic on them Constant arithmetic can be performed on both the vertical and horizontal axis values e a constant value can be added to or subtracted from all data points e data points can be multiplied by a constant or divided by a constant for additional ways to adjust signal values see chapter Chapter 7 7 e data points can be raised to a power e g they can be squared or the square root can be taken from them e time coordinates can be multiplied by a constant or have a constant added to them for additional ways to adjust time coordinates see chapter Chapter 7 7 e data sets can be added together or subtracte
164. is The Select menu option works much the same way as the Find but here all measurements that satisfy the search properties will be selected in the data list They may subsequently be loaded or removed 233 Chapter 19 8 View Selected lines This menu option limits the data being displayed in the Statistics List to those lines that are selected highlighted in the Data List Use the Edit Select menu option to easily select a number of lines 234 Chapter 20 Reports and the Report Editor If the menu option Report is not present you need to use File Customise gt Extensions and check Reports The Report Editor is used to create report definitions A report definition looks like a vector graph with many objects that will be filled with specific information when the report is printed Text objects for instance may have the content name when they are edited in the report editor When they are printed however name will be replaced with an appropriate name Similarly a data plot object looks like an empty rectangle in the report editor but will contain a data plot when the report is printed The various objects that can be used in a report are listed in chapter Chapter 20 4 The Report Editor displays a printer sheet as its work area The current default printer is used but a new type of printer may be selected using the File Target printer menu option The work area will change according to the
165. is printed 3 select a font and a font size Buttons are used to indicate bold italic and underlined text 4 set a colour for the object 5 set a border width for the object for lines this is the line thickness If the border width is zero border will be printed 6 go to the previous or next page in the report Reports may have as many pages as you like 7 add new objects An array of buttons is displayed in the lower left corner of the FontBar Click on one of these buttons to start adding a new object of a certain type After the button has been clicked go to the report sheet and drag a rectangle where you want to add the new object If multiple objects are selected see below any properties they share e g when they all have the same font will be displayed but properties that differ will not be displayed Changing any property will affect all selected objects 236 Chapter 20 3 Editing Report Definitions Chapter 20 3 1 Changing Position and Size of objects To change the position or size of an object first select the object by clicking on its surface If multiple objects overlap the object whose centre is closest to the position the mouse was clicked in will be selected Once an object has been selected a dotted line will appear around it as well as 8 small squares at the corners except for ines which only get two squares one at each end Move the object by clicking the mouse button on it and dragging
166. ithmetic operation involving a constant operator data were modified using an arithmetic operation that involved another data set number of data points was reduced by yanking i e only the first out of every N points was kept or by bunching replacing every N points by their average data points may also have been removed at the end of the measurement data points have been sorted by signal value or absolute signal value Sorting can be used to study noise for instance time coordinates were changed by dragging the curve see sizing chapter Chapter 8 4 time coordinates were changed by sizing the curve see sizing chapter Chapter 8 4 flow rate has been altered curve data points were changed by dragging the curve see sizing chapter Chapter 8 4 curve data points were changed by sizing the curve see sizing chapter Chapter 8 4 a gradient has been defined for the curve a gradient has been subtracted from the curve the data set is a blank measurement the data set has been sliced either removing data points or replacing them with a straight line if data points were removed the Reduced Data attribute will also be set Chapter 8 12 The Add Remarks to Log File Dialog This dialog box is used to manually add remarks to a log file for a data set The log file is kept by DAx to indicate all operations that are performed on a data set that may change the data set or 7 In the case of bunching time coordinates of the new points
167. ive area of peaks not listed in the bin due to the maximum number of peaks listed per bin being exceeded The above items can be listed only once since they pertain to the entire bin The following item would be listed for each peak e area of the peak relative to the total area of all peaks in the bin 213 Statistical items The Statistical items parameter page determines which statistics about each bin to display The average qualification coordinate value for each bin can be displayed as well as its standard deviation across the peaks found in the bin The range of the qualification coordinate can also be displayed When user defined bins are used the range is equal to the bin limits set by the user Display options The Display options parameter page is used to determine if bin lines should be wrapped displaying multiple horizontal lines per bin rather than one very wide line You can also indicate that empty bin cells should be hidden When this option is used all cells containing peaks are displayed first on each bin line not interspersed with cells that do not contain peaks Peak thresholds The Peak thresholds parameter page is used to indicate if peak thresholds should be used to determine if a bin cell contains a peak Even if a peak exists within the coordinate range of the cell if its height does not exceed the threshold the cell will be considered empty In addition to the global threshold value set on this param
168. k the right mouse button when the mouse is over a node and select the Delete node menu option from the popup menu 81 e press the Ctrl key while dragging the node to add a new node the mouse cursor will get a plus sign e pressing the Shift key and the Ctrl key then double clicking the left mouse button also adds a spline node All of these spline edits may be undone Type Alt BackSpace to do this Typing Shift Alt Backspace redoes the edit Undo redo is also available from the data tag spline menu Chapter 8 8 Data Tag Gradient Menu For more information about using gradients refer to chapter Chapter 14 Bring to Top Info w Plot Use as Baseline Blank Measurement Peaks Sizing gt Fourier Filtering gt F p Tacks gt Gradient Show Percentages Show Temperature Programmes Data Table Copy to New Curve v Show Gradient Copy to New Window v Mark Gradient Delete Data Copy Gradient to Other Data Sets Undo Gradient Redo Gradient 1 Use the Show percentages menu option to draw an extra curve in a graphics window depicting gradient percentages Percentages are either shown or hidden for all data sets they cannot be individually shown or hidden 2 Use the Show temperature programmes menu option to draw an extra curve in a graphics window depicting the temperature programme Temperatures are either shown or hidden for all data sets they cannot be individually shown or
169. ks are treated Refer to chapter Chapter 10 10 8 for details You have the option of not using the baseline to calculate peak areas but instead using a line connecting peak begin to peak end Note that when peaks overlap this will have undesirable results Finally the Make all peaks adjoin option can be checked When the option is active the end of one peak is always the start of the next peak This option should be used with caution Chapter 3 1 6 File Customise gt Plotting peaks Peaks are marked using triangular symbols Width and height of these symbols may be entered Furthermore you can select one or two peak labels and you can choose between placing peak labels horizontally and vertically Use the Draw perpendiculars option to start or stop dropping dotted perpendiculars from peak start and stop coordinates onto the baseline Component concentrations can be displayed in peak lists with or without their range The number of decimals that will be used to list concentrations and relative concentrations can be set as well Relative peak areas can be displayed as fractions or as percentages Chapter 3 1 7 File Customise gt Data Some techniques to modify data may be shown or hidden These techniques are Fourier filtering sorting slicing de convolution data set correlation interpolation integration converting data to 2Log removing trends and best overlays Generating theoretical artificial data may be enabled or
170. l standard peak as the reference peak cf chapter Chapter 10 10 4 in each of the measurements you want to include in the calibration use the data tag peak menu to display a peak list chapter Chapter 10 11 for each of the measurements you want to include in the calibration enter the known calibrated values in the annotation column in the peak list chapter Chapter 10 11 4 If the annotation item is not being displayed in the peak list use the View Configure menu option to add it to the displayed columns The annotations can also be entered by right clicking peak top markers and entering the annotation in the peak popup dialog box invoke the Calibration Calibration menu option discussed in detail in chapter Chapter 17 6 As an alternative to entering calibrated values as peak annotations they may also be entered as part of peak names This makes it possible to identify peaks using an Identification Database and then set up a calibration using automatically assigned peak names 176 Chapter 17 3 Calibration Load calibration Calibration Save calibration Calibration Stop calibration menu options Calibrations may be saved to disk using the Calibration Save calibration option They may be loaded from disk using the Calibration Load calibration option f the Separate Parameter Files item in the File Customise gt Extensions dialog box is unchecked CE parameters can only be saved as part of an analysis
171. l Config dialog box for new measurements invoked using the Config Acquisition menu option For existing measurements the data type can be changed either in the data sizing dialog box or in the data info dialog box both invoked from the data tag menu cf chapter Chapter 8 Chapter 3 3 6 Horizontal Axis Units Traditionally chromatographic measurements use time coordinates along the horizontal axis In addition to this DAx supports e point indices The first data point has coordinate 1 additional points are at coordinates 2 3 4 etc e calibrated values Typically DNA base pair counts fragment sizes Refer to chapter Chapter 17 for details e molecular weight axes e mobility axes 25 The terminology in this manual assumes that a time coordinate is used Chapter 3 3 7 Modified Data Modified Data is the opposite of raw data i e data that have not been modified since they were measured DAx does not allow any irreversible operations to be performed on raw data so those operations require Modified Data as their input Irreversible operations always have the option of creating a copy of the raw data before performing the actual operation See also chapter Chapter 3 3 2 Chapter 3 3 8 Data set names The data set name is the name that DAx uses to distinguish data sets The data set name equals the proper file name used for the initial data set as it was measured Alternatively the data set name equals the
172. lar when you are hiding empty cells 217 User defined bins can be limited to certain trace types so that only data from data sets matching the specified trace type will end up in the bin The bins equal peaks option causes all existing peaks to be removed from data sets that are added to the binning sheet New peaks with coordinates exactly equal to all user defined bins are then created in the data sets The peaks that were created this way cannot be manually edited As long as a data set is in the binning sheet it is excluded from the Peaks Find peaks menu option User defined bins can be copied from the binning sheet dialog and pasted into the marker peak dialog chapter Chapter 18 6 or the meta analysis list chapter Chapter 18 13 If bins have been automatically assigned they can be converted to user defined bins and thus fixated using the Analysis Convert to user defined bins menu option Chapter 18 12 4 Displaying curve samples Small curve samples of the data in each bin can be plotted instead of displaying the quantifying parameter Use the View Curve samples menu option to turn curve samples on or off the option can also be set in the binning sheet dialog box The line height of the binning sheet is adjustable when curve samples are displayed To change it place the mouse cursor between two binning sheet lines where it will change to an up down shape click and drag to set the new line height A triangul
173. lation Use the derive from data check box to make the volume limits equal to the range of volumes in the GPC calibration list 3 The Molecular Weight limits determine which molecular weight values calculated when plotting Molecular Weight axis or calculating molecular weight peak data will be accepted as valid Becale Calibration 165 Use the derive from data check box to make the limits equal to the range of molecular weights in the GPC calibration list for points that are within elution volume bounds 4 Do not alter points usage ensures that the same points will be used in the new calibration that were used in the old calibration This option is useful when points have been manually excluded from the calibration 5 The Recalculate calibration button activates a possible new calibration type using linear or logarithmic Mw values polynomial degree elution volume limits or molecular weight limits 6 Add to existing calibration is used to add calibration points to an existing calibration If the option is not checked a new calibration will be derived from the selected data sets 7 Detector measures weight indicates if the detector measures weight or concentration abundance If two substances A and B are present with equal concentration and B has a molecular weight of 2 A a detector that measures concentration will give equal signal values but a detector that measures weight will have a doubled signal value
174. let you modify the width and height of the markers To modify a peak move the mouse cursor over one of the markers and drag it to a new location This procedure can be used for peak begin end as well as top markers When you move the mouse cursor over a peak marker if there is a peak list window chapter Chapter 10 11 the appropriate peak will be highlighted in the list1s Another way peaks will be highlighted is when you zoom in on part of the graph the peaks that lie in the zoomed window are highlighted DAx 1 0 PP 18 1 96 Volt 0 3 30 40 time s Example graph with peak markers peak markers have been enlarged for clarity e f a peak top is dragged beyond the begin or end of the peak the peak will be removed You will only be asked for confirmation if peaks are very close together e f the Ctrl key is pressed while a peak top is dragged and released a new peak will be added DAx will search for reasonable peak boundaries either the point where the signal crosses over the baseline or the point where the direction of the signal changes i e a valley between peaks Peaks will be adjusted in such a way that the new peak does not overlap any existing peak with the exception of shoulder peaks see chapter Chapter 10 10 4 13 This behaviour is suspended if you press the Shift key or if Scroll Lock is active 125 If the Ctrl key is pressed while a peak begin or end marker is dragged and
175. lie in the middle of the intervals that contained the N points 86 generate results from it such as peaks Click the Add button in the information dialog box to be able to manually add remarks 87 Chapter 9 Data Set Graphics Window Data menu The Data menu of a Data Set graphics window contains various options that modify the data or derive new data Many of these options can be performed on one or more of the data sets in the graphics window These options use a dialog box that has a list of the data sets in the window at left In this list the user may select check those data sets that should have the operation performed on them Not all operations can be performed on all data sets The dialog box has New window and New Curve options These will copy the data before performing the operation either to a new graphics window or to a new data set Various restrictions to data set selection may apply If a data set cannot be selected the tool tip that is displayed when the mouse cursor is held over the data set will list the reason Chapter 9 1 Data Relationships amp Sorting This command invokes a dialog box that displays how the different data sets are related to each other Each copy of a data set belongs to the original copy For instance when a raw data set is filtered because of GLP chapter Chapter 3 3 2 a copy of the original data set has to be created This copy is a child of the original data When a data se
176. line Press the Shift key while clicking the menu option to remove the peaks and baselines for all data sets in the graphics window 11 The undo option in the tag peak menu undoes moving adding or deleting operations Chapter 8 3 1 Showing Boundary Conditions Use the Show Boundary Conditions menu option to switch between displaying peak boundary conditions and hiding them There are two types of boundaries shown if they were used e thresholds the baseline is shifted up by the positive threshold value and down by the negative threshold value to show where the thresholds values are crossed by the data curve e derivative thresholds lines are drawn at the angles at which a data curve derivative is thought to become high enough to indicate the start of a peak 74 Press the Shift key while clicking the menu option to show or hide peak boundary conditions for all data sets in the graphics window Chapter 8 3 2 Fixating Peak Names and Concentrations Normally when peaks are named and quantified using an Identification Database the names and concentrations can change if a different Identification Database is loaded or if the Identification Database is edited To prevent names or concentrations from being changed fixate the peaks using the Peaks Fixate menu option Saving data to disk also fixates the peaks Modifying data such as by filtering them or by manually adjusting peaks will unfixate peaks The Peaks Unfixate menu op
177. ll these identifications The peak name will consist of multiple identifications separated by semicolons with the best matching identification listed first If there are too many identifications to fit in the peak name the peak name will end in The peak type and peak annotation will be assigned from the identification database item that best matches the peak Multiple peaks per identification can also be allowed If multiple peaks are within tolerance for a single identification database item each of these peaks will be assigned the identification database item s name Marker peaks can also be multiply assigned However if a peak is assigned with a marker peak it cannot also be assigned with an identification database item Marker peaks are only assigned to multiple peaks if they use the near coordinate search method but a single peak can be assigned with multiple marker peaks using any search method Chapter 18 6 Marker amp Reference Peak Allocation Dialog Box If the menu option Analysis Marker peaks is not present you need to use File Customise gt Extensions and check Use Marker Peaks This dialog box is used to set up the automatic recognition or allocation of a number of marker peaks It is invoked using the Analysis Marker peaks menu option One or more marker peaks can be marked as reference peak if a data set contains multiple marker peaks marked as reference peak 194 the last one recognised becom
178. locally stored calibrations The Parameter retrieval menu options retrieve parameters that have been stored with the data set Use the File Customise gt Extensions option to enable storing parameters with data 72 10 Use the Data Table menu option to create a text window listing all data points in the data set The information of the text window may be printed copied to the clipboard or saved as a text file 11 Use the Copy to New Curve or Copy to New Window menu options to copy the data set to a new window or to an extra data set Copying a data set is also achieved by pressing the Ctrl key and dragging the data tag to a Data Set Graphics window or to the DAx client area to create a new window 12 Use the Delete Data menu option to remove the data set from memory The data will be stored in the waste barrel from where they may later be recovered If the waste barrel is not available you will be asked for confirmation Note that the waste barrel has a limited capacity and may overflow losing the oldest data it contained Alternatively drag the data tag to the waste barrel Chapter 8 3 Data Tag peaks menu 1 Use the Mark Peaks menu option to switch between displaying peak markings and hiding them Peaks Bring to top are marked by markers at their begin and end limits and at their one top These markers may be arention dragged to move the top or the colur gt limits See also chapter Chapter
179. lse as baseline 10 5 a Blank measurement Press the Shift key while clicking SENEE the menu option to show or hide ae s Show boundaries peak markings for all data sets in e v Show labels the graphics window mounier nker A eE 2 The Show Boundaries menu splines gt Plot curves option switches between showing saks Total curve d hidi b d diti Turn total curve into data set and hiding boundary conditions Add label E SLEY see chapter Chapter 8 3 1 ar Cale RMS noise radien 3 Use the Show Labels option to TAE switch between displaying peak mires Fixate labels in the graphics window and serail Unfixate hiding them See also chapter Datatable EESO O O Chapter 10 11 6 Use the E z z i Delete data Show meta analysis File Customise gt Plotting Add to binning sheet Peaks menu option to select which of the columns in a peak list window should be used as label __ Clear analysis Press the Shift key while clicking Log in analysis log the menu option to show or hide peak labels for all data sets in the graphics window 4 The Fit Gaussians menu option e Remove from binning sheet Display analysis assay Undo peak Redo peak 13 causes Gaussian curves to be fitted to all peaks in the data set using the parameter settings most recently entered using the Peaks Fit Gaussians menu option chapter Chapter 10 6 Plot curves causes the Gaussians to be marked If no Gaussians have been fitted yet or if th
180. menu options are not displayed If the CE calibration has been changed but has not been saved to disk yet when DAx is about to terminate you will be asked to confirm the loss of the new calibration Saving the calibration to a floppy disk does not prevent this you must save to hard disk Chapter 16 4 CE Apparent mobility CE Effective mobility CE Track mobility menu options In graphics data set windows either apparent or effective mobility can be tracked in the coordinate display In order for effective mobilities to be tracked a reference peak chapter Chapter 10 10 4 must have been defined and an effective mobility for the reference peak s component must have been entered as an annotation for the reference peak If no effective mobility is entered in the reference peak s annotation the default reference mobility from the CE Capillary Parameters dialog box will be used instead If no default reference mobility has been entered for a data set the default reference mobility last assigned to any data set will be used The peak top is used as the migration time coordinate for the reference mobility Use the CE Track mobility option to determine if mobility coordinates are displayed in the coordinate tracking bar in the first place Chapter 16 5 Mobility Axes The menu option CE Mobility axis displays graphics where the time axis has been replaced by an axis that contains the mobility at a given time coordinate using the curr
181. move unselected button causes unselected preserved data to be removed unselected rather than selected data sets are removed because they are not displayed when the OK button is clicked Chapter 18 12 2 Binning sheet popup menu options The Binning Sheet has the usual text window menu options Other than in the usual text export formats binning sheet contents can be exported in PAUP and PHYLIP formats using the File Export menu option When the right mouse button is clicked in the sheet a popup menu appears This popup menu offers the ability to locate or highlight a peak or bin in a graphic window press the Shift key to hide all other data sets or to highlight all selected peaks 215 The peak in the cell that was clicked on can be marked for attention Its data set will also be marked If bin annotations have been defined they can be applied to the peak in the cell that was clicked on Press the Shift key when choosing the bin annotation to also mark the peak and data set for attention NB If the Shift key is pressed when the right mouse button is clicked a shortened popup menu containing only the bin annotations is displayed The Show source data menu option brings the graph window that a peak was derived from to the front If user defined bins are used the Insert peak and Remove peak menu options are used to insert a peak in the location of the bin cell that was clicked on or remove an existing peak Double clicking
182. nce peak as an internal standard If no internal standard peak is marked as a reference peak the peak top time will automatically be used instead of the peak top time offset Chapter 17 1 Setting Up the Calibration Type The type of calibration DAx will calculate is set up by clicking the Set Calibration Type button in the calibration dialog box which is invoked using the Calibration Calibration menu option A calibration is defined by the following parameters e ashort string naming the calibration e g DNA a Calibration parameter see below name of calibrated value e g base pair count short name for calibrated value e g BP number of significant digits to be used when writing calibrated values A value of 0 causes whole numbers to be written 175 The following parameters can be used as calibration parameter Peak top time Peak moment Peak top time offset Capacity factor Peak elution volume Apparent mobility Effective mobility By default DAx is set up for DNA base pair count calibrations Chapter 17 2 Creating a calibration To create a calibration 1 2 3 4 7 measure one or more samples with known calibrated values e g known base pair counts construct a baseline refer to chapter Chapter 10 1 for details detect peaks refer to the chapter Chapter 10 4 for details if peak top time offset capacity factor or effective mobility is used as a calibration parameter mark the appropriate interna
183. nd height of the peak markers This will affect all graphics windows You can stop plotting markers altogether by un checking the Draw Markers box Chapter 21 3 Why are the Molecular Weights in the peak list blank why can t calculate molecular weights No GPC Calibration has been loaded Use the GPC Load calibration menu option to load a pre existing calibration or set up a new calibration refer to chapter Chapter 15 1 for step by step instructions Make sure the flow rate is entered correctly Use the data tag sizing menu to enter it refer to the Measurement Window manual for details Another reason why molecular weights may not be calculated for some peaks is that the time coordinates may not be in the valid range for the calibration Remember that only multi linear and 245 polynomial calibrations can do any extrapolating beyond the time coordinates spanned by the calibration points In other words if you use a cubic spline calibration the unknown peaks must have time coordinates within the range of time coordinates the calibration peaks had Keep in mind that even though multi linear and polynomial calibrations can extrapolate the calibration may still contain limits for elution volumes and molecular weights Make sure you set these limits wide enough in the GPC Calibration dialog box chapter Chapter 15 5 Chapter 21 4 Why is the Effective Mobility column in the peak list blank Refer to what is said
184. no more peaks lie within the tolerance interval the process is repeated until there are no data sets left with peaks that have not been put into a group yet 206 Chapter 18 12 Binning Sheets The Analysis Binning sheet menu option is used to create a text window that compares the peaks found in different data sets grouping corresponding peaks together The option first displays a dialog box that lets you select which data sets to include in the comparison The binning sheet is a global sheet which means that it can contain data from any open graphics window Peaks in different data sets will be grouped into clusters of corresponding peaks using a qualifying parameter and tolerance The qualifying parameter and tolerance will be copied to the current Identification Database By default the current values from the Identification Database are used Whereas a comparison sheet chapter Chapter 18 11 lists all peak parameters with successive peaks being listed on successive lines a binning sheet only lists the qualifying coordinate and a quantifying parameter of corresponding peaks on a single line Below is an example of a binning sheet DAx 7 3 14 09 2003 14 59 51 PP Binning sheet qualifying on Base pairs with absolute tolerance of 1 Index Number Average Base pairs Height Base pairs Height Data b02 ET ROX b02 FAM 1 2 100 0 100 0 2706 2 100 0 4828 1 2 1 102 2 o 102 2 208 29 3 1 103 3 103 3 176 36 4 2 106 9 106 8 1
185. normalise peak areas The Data Overlay chapter Chapter 9 16 command can adjust signal coordinates for data sets in such a way that the normalisation peaks in each data set have equal height The normalised peak area for each peak can be displayed in peak list windows If a normalisation peak has been defined all areas are expressed as a percentage of the area of the normalisation peak if no normalisation peak has been defined the highest peak is used to normalise peak areas Chapter 3 3 15 Reference Peaks Reference peaks are used in four ways e The Data Overlay chapter Chapter 9 16 command can adjust time coordinates for data sets in such a way that the reference peaks in each data set have equal time coordinates e In chromatography the capacity factor k is calculated as k mto with t the retention time of the component and ty the 0 retention time of an inert component Peak tops not the first peak moments are used as retention time coordinates in DAx The peak top time of the reference peak is used as the reference time t If no reference peak has been defined the inert time is 28 used The inert time is a sizing parameter cf chapters Chapter 7 7 and Chapter 8 5 e Reference peaks are used to derive the electro osmotic flow in capillary electrophoresis Refer to chapter Chapter 15 for details e Reference peaks are used to denote an internal standard when relative peak top time capacity factor or effective m
186. nt views The Display list form is aimed at presenting filenames file dates and operator information but cannot present component information The Statistics List is meant to present component information derived from peak lists stored in files Note that only information stored in files is presented the peaks are not re qualified when the file is scanned 232 Chapter 19 5 Data List Find data The Find data menu option searches the hard disk for measurements files meeting specific search criteria The difference with the Edit Find menu option is that the search does not occur in the current data list but in the files that are on the hard disk Chapter 19 6 Edit Sort list menu option This menu option can be used to sort the data list in order of any of the properties The list may also be sorted by clicking on any of the column headers Clicking a second time sorts the list in descending order Chapter 19 7 Edit Find Edit Find next Edit Select menu options The Find menu option displays a dialog box with a comprehensive list of search properties The Data List will be searched for the first measurement that satisfies the specified search properties The search is started from the current line in the data list not from the top and follows the current sort order The Find next menu option finds the next measurement that satisfies the search properties The F3 function key can be used as a shortcut for th
187. ntification of all entries belonging to the species Chapter 18 13 1 Meta analysis patterns The wanted peaks for a species can be made to have to satisfy a pattern using the pattern pattern quantifier and pattern target columns in the meta analysis list displayed using the Analysis Edit meta analysis menu option Available pattern quantifiers are height area normalised area migration time corrected area and migration time corrected normalised area The pattern target specifies the pattern that the quantifiers of the peaks must satisfy For instance three peaks may be specified to have pattern targets of 1 1 1 If all three peaks are present but they do not have equal heights or area etc they are considered to be absent If the pattern target is specified as 1 2 1 the middle peak must be twice 23 The Show meta analysis menu option is also displayed when the peak list button at the left side of a data tag is right clicked 222 as high as the first and last peak or they will again be considered to be absent Each species has at most a single pattern consisting of all wanted peaks that use a pattern The procedure used to match the pattern is as follows the quantifying value of all patterned peaks for a species is totalled The value for each peak is divided by this total and multiplied by the total of the pattern targets specified The resulting fractions must then match the pattern target to within 0 2 If they do
188. nto window Changed nodes only GPC calibration gt i 5 i ie Undo spline Automatically Calibration gt i Redo spline Parameter retrieval gt Data table Copy gt Delete data 1 Use the Add Spline option to display a spline for the data set If a spline is already present clicking this option will hide it but it will not be removed Press the Shift key while clicking the menu option to add splines to all data sets in the graphics window 2 Use the Cubic spline option to switch between straight line splines where straight lines are drawn between the spline nodes and cubic splines where third order polynomials are drawn Press the Shift key while clicking the menu option to switch all splines in the graphics window to the new value straight or cubic 3 Use the Replace curve with spline or Turn spline into curve window options to replace a data set with a spline turn the spline into a separate data set in the current graphics window or turn the spline into a data set in a new graphics window When replacing a curve with a spline you can either replace the entire curve just the part currently visible or just the parts that were changed The latter is useful for partial changes If the data set contains raw data that is if the data set is not Modified Data the data set cannot be replaced with the spline because of GLP cf chapters Chapter 3 3 2 and Chapter 3 3 16 Replace curve with spline gt automatically causes the data
189. o multiple data tags e select multiple tags by pressing the Ctrl or Shift key when clicking additional tags Use the Ctrl key to select individual additional tags use the Shift key to select all tags between the last clicked and currently clicked tag Selected tags will be displayed with a check mark on the right side To invoke the data tag menu without changing the selection of tags click on one of these check marks 71 click the data tag with the right mouse button instead of the left mouse button to select an enslavement group rather than a single tag while choosing a data tag menu option press the Shift key to select all data tags this is possible only for certain menu options The data tags give visual feedback about whether or not the currently highlighted menu option will be applied to a single or multiple data tags all affected data tags are depressed 1 The Attention menu option can be used to mark the data Use the Use as Baseline menu option to make this data Use the Blank Measurement menu option to indicate The Bring Tag to Top menu option makes the data set the default chapter Chapter 3 3 4 and the tag the top 3 F tag This means that its data will be drawn on top of all Info other data in the graphics window v Plot The Info menu option displays the data set info dialog tention Refer to chapter Chapter 8 10 Colour The Plot menu option can be used to switch between Use
190. oard refer to chapter Chapter 5 for an overview of common text window options e Peak List windows chapter Chapter 10 11 These windows list the peaks that were detected for a data set e Colour view chapter Chapter 7 3 18 A window displaying a coloured map representation of a data set graphics window 20 Gradient Corrected windows chapter Chapter 14 6 These windows display signal values from which the defined signal gradient has been subtracted Volume axis windows chapter Chapter 15 3 These windows display the same signal values as graphics windows but they use an elution volume horizontal axis Molecular Weight axis windows chapter Chapter 15 3 These windows use the current GPC calibration to calculate a molecular weight for each data point based on its elution volume Molecular weight is then used as the horizontal coordinate in linear or logarithmic form The vertical coordinate may be signal value or signal value multiplied by molecular weight Vertical coordinates may be plotted differentially or cumulatively or both GPC Calibration windowg chapter Chapter 15 3 The current main GPC calibration which defines a relationship between elution volume and molecular weight or a local GPC calibration is plotted in this window GPC Calibration list t chapter Chapter 15 6 The current main GPC calibration or a local GPC calibration is listed in this window For each calibration point the elution volume and mole
191. obility are used as calibration parameter Refer to chapter Chapter 17 Chapter 3 3 16 Marker Peaks Any peak can be the marker peak for another peak Peaks do not need to be marked as a marker peak but you do need to set the marker peak for a certain peak This is done using the peak popup dialog box Some automatic analysis methods will automatically assign marker peak names Marker peaks are used to calculate relative times for peaks and to recognise peaks based on their position relative to other peaks internal standards Chapter 3 3 17 Splines Traditionally the craftsman s spline is defined as a flexible piece of wood hard rubber or metal used in drawing curves In mathematics splines are curves that intersect a predefined table of points They are defined in such a way that they are continuously derivable that is there are no sudden changes in direction or derivative Although DAx has the option of using splines in this way their primary use is slightly different DAx can represent a data set by means of a limited number of spline nodes i e points through which the spline is plotted Spline nodes are connected either by straight lines or by cubic polynomials cubic splines These nodes can be moved added or deleted When the user is done moving spline nodes the data set that was being represented by the spline may be replaced by the spline This technique is used to manually adjust data sets particula
192. ommand to rescale the display Chapter 7 3 5 View Set colours This menu option is discussed in chapter Chapter 9 3 Chapter 7 3 6 View Data set names This command switches the display of data set names at the left side of the graph between Do not label Label above curve and Label below curve This option will typically be used in conjunction with data set stacking cf chapter Chapter 9 17 Chapter 7 3 7 View Tags setup This command invokes a dialog box that determines where the data tags are displayed in the window as well as the width of data tags and the number of lines of data tags Chapter 7 3 8 View Edit scale This command invokes a dialog box that allows the scale to be set Each window has one horizontal axis but may have several vertical axes The scale for all of these may be entered Some scales have an always use this scale box next to them If you check that box the scale entered will always be used To start using an optimised scale again re invoke View Edit scale and uncheck the box Chapter 7 3 9 View Zoom in The zoom command zooms in on the centre one third of the graph Chapter 7 3 10 View Zoom out Zoom Out undoes one level of zoom 60 E EE Chapter 7 3 11 View Reset scale Reset Scale undoes all zooming and optimises the scale so that all curves will be optimally visible Chapter 7 3 12 View Seconds axes View Minutes axes These commands switch between displaying time
193. on the binning cell has the same effect If curve samples are being drawn the Copy curve sample menu option places a copy of the peak sample on the clipboard The Display single bin menu option limits the binning sheet to displaying a single bin You can then use the Previous bin Next bin View Previous bin View Next bin options to navigate to the previous or next bin Use All bins or View All bins to once again display all bins The menu also offers options to change the order of the columns in the sheet including invoking a dialog box that allows the columns to be sorted by one or more criteria The column order can also be changed by clicking in a column header and dragging the column to a new position This functionality is not available when binning lines are wrapped using the View Wrap lines menu option or when trace types are used or when empty cells are hidden When curve samples are being drawn use the Mark data set as master menu option to start drawing a curve sample from the selected master data set in all cells Use Preceding data set as master to draw a curve sample from the preceding data set in 216 each cell21 The No master menu option stops displaying a master curve sample The Data set needs attention option marks the data set corresponding to the bin cell that was clicked on for attention Remove data set is used to remove a data set from the binning sheet If user defined bins are used bins can b
194. ons 233 Chapter 19 8 View Selected lines 234 Chapter 20 Reports and the Report Editor 235 Chapter 20 1 The Report Menu 235 Chapter 20 2 Editing Reports The FontBar 236 Chapter 20 3 Editing Report Definitions 237 Chapter 20 3 1 Changing Position and Size of objects 237 Chapter 20 3 2 Changing the Type of an Object 237 Chapter 20 3 3 Editing Multiple Objects 238 Chapter 20 3 4 Removing Objects 238 Chapter 20 3 5 Copying Objects 238 Chapter 20 3 6 Customising the Report Window 239 Chapter 20 4 Report Objects 239 Chapter 20 4 1 Spreading report objects across multiple pages 241 Chapter 20 4 2 Text Object Variables 241 Chapter 20 5 Customising the Report Editor s Appearance 242 Chapter 20 6 Previewing Reports 243 Chapter 20 6 1 File Write report 243 Chapter 20 6 2 Report Optimise scale 243 Chapter 20 6 3 Report Single graph Separate graphs Graph per set 243 Chapter 20 6 4 Customising the Report Preview Window 244 Chapter 21 Frequently Asked Questions and Troubleshooting 245 Chapter 21 1 Can I search for peaks in a time interval 245 Chapter 21 2 I don t like the way peaks are marked 245 Chapter 21 3 Why are the Molecular Weights in the peak list blank why can t I calculate molecular weights 245 Chapter 21 4 Why is the Effective Mobility column in the peak list blank 246 Chapter 21 5 Why are there no curves in my Mobility Axis Window 246 Chapter 21 6 How do I exclude the EOF peak from area calculations 246 Ch
195. ons are being used the new main calibration will also be stored as the local calibration in all data sets in the current graphics window unless they already have a local calibration and were not used to derive the new main calibration and are not in the same enslavement group as any data set used to derive the new main calibration Copy Main Calibration from retrieves a stored local calibration and stores it as the main calibration If the calibration parameters are changed they are applied to the local calibration before it is copied to the main calibration If the main calibration is subsequently changed the local calibration is not affected Store Main Calibration In stores the current main calibration in the selected data sets as local calibrations If the calibration parameters are changed they are applied to the main calibration before it is stored in the selected data sets After this if the main calibration is changed the local calibrations are not affected Cf Chapter Chapter 17 9 Change Configuration For changes the calibration parameters for the selected local calibrations without changing the calibration points The parameters for the main calibration are also changed I Derive from data la 179 2 The calibration curve type determines the model for the calibration curve that is fitted through the calibration points The straight line spline and local Southern type calibrations will intersect all points used
196. op Mp Molecular Weight at peak top Mn number averaged Molecular Weight calculated as _ signal M signal 2 Mw weight averaged Molecular Weight calculated as _ Yi signal M ad signal M _ NM gt signal M me NM X signal M ins M D dispersivity D T The following items are CE related App Mob apparent mobility calculated as Magy with Mapp the apparent mobility in 10 m Vs kot length of capillary laet length from injection to detection t migration time and V voltage over the capillary Eff Mob effective mobility in 10 m Vs calculated as My Mp EOF with EOF the electro osmotic flow in the capillary determined using the reference peak Area Mig migration time corrected area calculated as signal At Areal Mig _ j 2 time Area Migration Marker migration time corrected area of the peak divided by the area of a marker peak Other area items corrected in the same way for migration time are Area Mig Fraction Norm Area Mig LPeakFract Mig and RPeakFract Mig The following items are calibration related Calibrated value calibrated value based on calibration parameter value of the peak available for start top and end of the peak Calibrated value at start of peak Calibrated value at end of peak 138 Some versions of DAx have the ability to fit Gaussian curves to all peaks using the Peaks Fit Gaussians menu option The fitted values can be displayed in the peak list
197. ors enter the appropriate concentrations when setting up the Identification Database For instance let s assume the database is being set up with IS as the internal standard and A and B unknown components In the sample that is being used to set up the database let s assume all three peaks have the same size We further know that A has the same response factor as IS but B has a response factor that 203 is twice as high Since the peaks are the same size the concentration that should be entered for A should be twice as high as the concentration being entered for B Chapter 18 11 Comparison Sheets The Analysis Comparison sheet and Analysis Global sheet menu options are used to create a text window that compares the peaks found in different data sets and optionally performs statistical analysis on them The option first displays a dialog box that lets you select which data sets to include in the comparison Peaks in different data sets will be grouped into clusters of corresponding peaks There are three possible ways in which DAx determines which peaks correspond e using peak names e using peak indices used to compare data known to contain the same peaks e using a qualifying parameter and tolerance The qualifying parameter and tolerance will be copied to the current Identification Database By default the current values from the Identification Database are used gt Select Data Sets for Comparison Sheet 2 x HUE
198. oss over Peaks 121 Chapter 10 5 Peaks Baselines amp peaks 123 Chapter 10 6 Peaks Fit Gaussians 123 Chapter 10 7 Peaks Clear analyses 124 Chapter 10 8 Peaks Un Fixate analyses 124 Chapter 10 9 Peaks Recalculate peaks 124 Chapter 10 10 Modifying Peaks 124 Chapter 10 10 1 Peak Popup Dialog Box 127 Chapter 10 10 2 De Activating Peaks 127 Chapter 10 10 3 Normalisation Peaks 128 Chapter 10 10 4 Reference Peaks 128 Chapter 10 10 5 Marker Peaks 129 Chapter 10 10 6 Shoulder Peaks 129 Chapter 10 10 7 Automatic Shoulder Peak Detection 130 Chapter 10 10 8 Shoulder Peak Area Calculations 131 Chapter 10 10 9 Connecting and Combining Peaks 132 Chapter 10 11 Peak List Windows 132 Chapter 10 11 1 Peak Highlighting 133 Chapter 10 11 2 Deleting Peaks from the Peak List Window 133 Chapter 10 11 3 Locating or Highlighting Peaks 133 Chapter 10 11 4 Peak Annotations 134 Chapter 10 11 5 Setting Marker Peaks 135 Chapter 10 11 6 Labelling Peaks in the Graph 135 Chapter 10 11 7 Columns in Peak List Windows 135 Chapter 10 11 8 How Relative Concentrations are Calculated 139 Chapter 10 11 9 Only List Named Peaks 140 Chapter 10 11 10 Only Show Peaks of Certain Types 140 Chapter 10 11 11 The Peak List window Popup Menu 140 Chapter 11 Filtering Techniques 143 Chapter 11 1 Fourier Filtering 143 Chapter 11 2 Deriving Fourier Filtering Characteristics 146 Chapter 11 3 Savitzky Golay and Moving Average Filters 147 Chapter 11 4 The Spike Removal Algorithm 147 C
199. ot been saved since they were last modified the user will be given the 53 opportunity to save the data Data will not be sent to the waste barrel chapter Chapter 11 Chapter 7 1 5 File Scanning Scanning for files is described in Chapter 4 2 Chapter 7 1 6 File Preview report This command creates a preview window for the report that can be printed using the File Print Report menu option Chapter 7 1 7 File Print report This command prints a report of the contents of the graphics window using the current report definition Chapter 7 1 8 File Print This command prints the contents of the graphics window The Print command is very versatile It invokes the following dialog box 54 hh lx Printer Name BUFFYSHP Properties Status Ready Type HP DeskJet 710C Where BUFFYSHP Comment J Print to file m Curve Selection Copies Asis j Number of copies 1 Plot per enslavement group 3 3 Collate Curves baselines separately aE je ae m Scaling Extra items Il Optimise Scale 2 V Curve line samples T Pit bands 3 I Include nfo amp LogFiles 4 oscar TET ma J Include Peak Lists 5 Bands per page 4 Cancel 1 You can either print all visible curves on one page or print each curve on a separate page 2 The scale of the axes can be optimised prior to each print This is especially useful if curves are being printed on separate
200. ow The Calibration Curve window has the usual graphics window functionality chapter Chapter 1 When the mouse cursor is moved over a calibration point in the graphics window the corresponding line in the text window will be highlighted if a Calibration list window is open Double click the mouse on a calibration point to include or remove the point from the calibration The View Linear and View Logarithmic menu options may be used to switch between linear and logarithmic calibrated value axes Chapter 17 8 3 When the calibration changes Whenever the calibration is changed either by removing lines from the calibration list or by double clicking points in the calibration curve the following steps are taken e Anew calibration will be calculated e The calibrated value data in peak list windows chapter Chapter 10 11 will be recalculated e Any graphics windows with Calibrated Axes chapter Chapter 17 5 will be re plotted Chapter 17 9 Local Calibrations If Local Calibrations are enabled using the File Customise gt Extensions menu option each data set can store and use its own calibration This makes it possible to 182 re analyse data each time using the calibration that pertains to that specific data set If a data set does not have a local calibration the current main calibration will be used There are two ways in which a data set can obtain a local calibration e Using the Store Main Calibration in
201. pages Do not use this option if you want to print zoomed graphs 3 Some versions of DAx have the ability to print bands This means that the current horizontal scale is divided into a number of slices the total number of slices is specified in the Print dialog as well as the number of slices per page 4 Data set information can be included with the prints Refer to chapter Chapter 8 10 for details on data set information 5 Peak lists can be printed along with the graphs The peak lists will have the same columns as are currently being used in peak list windows chapter Chapter 10 11 Chapter 7 1 9 File Printer setup This command lets you choose and set up the printer that is to be used 55 Chapter 7 1 10 File Load analysis procedure File Save analysis procedure These commands load and save Analysis Procedures chapter Chapter 3 3 1 These procedures fully define the way in which measurements are interpreted Chapter 7 1 11 File View message log This command inspects the log of error messages and important warnings Cf GLP chapter Chapter 3 3 2 Chapter 7 1 12 File Operator This command is used to change the operator name Chapter 7 1 13 File Customise This command invokes the customisation dialog box cf Chapter 3 1 Chapter 7 1 14 File Exit This command closes DAx You will be asked to confirm closing windows that contain unsaved data If a measurement is still running you will be asked
202. pe gt lt all gt ROX Offset to marker 498 bp First in interval 400 0 1200 0 _ SS Pa Use extemal markers in J related data J any data 4 Do not use intemal markers Help Hi F2 to edit inside item Add Line Copy Peak Data Edit Cut Paste Print Marker Peak dialog box For an explanation on 1 see Chapter 18 6 2 Marker peaks can be allocated automatically based on a number of qualifying coordinates peak top time first statistical peak moment peak begin time peak end time peak top volume peak top molecular weight number averaged molecular weight for the peak weight averaged molecular weight for the peak Mz for the peak apparent mobility for the peak top calibrated value for the peak top tis also possible to use peak top time minus marker peak top time peak top time divided by marker peak top time relative position index between two markers 195 Clearly the markers relative to which new markers are being allocated must have been allocated already In other words later markers can be allocated relative to earlier markers in this context later means lower down in the list of markers not necessarily later in a measurement NB When you select lt none gt as the qualifying coordinate for a marker peak the peak will not be found This can be used to switch certain markers off Depending on the measurement
203. peaks 17 Chapter 3 1 6 File Customise gt Plotting peaks 18 Chapter 3 1 7 File Customise gt Data 18 Chapter 3 1 8 File Customise gt Measurement window 19 Chapter 3 1 9 File Customise gt Measurement data directory 19 Chapter 3 1 10 File Customise gt Fields setup 19 Chapter 3 1 11 File Customise gt Report headers 19 Chapter 3 1 12 File Customise gt Data lists 19 Chapter 3 2 Window Types in DAx 20 Chapter 3 3 Terminology 23 Chapter 3 3 1 Analysis Procedures 23 Chapter 3 3 2 GLP 23 Chapter 3 3 3 Data Sets 24 Chapter 3 3 4 Default Data Sets 25 Chapter 3 3 5 Measurement Data Types 25 Chapter 3 3 6 Horizontal Axis Units 25 Chapter 3 3 7 Modified Data 26 Chapter 3 3 8 Data set names 26 Chapter 3 3 9 Enslavement 26 Chapter 3 3 10 The Waste Barrel 27 Chapter 3 3 11 Baselines 27 Chapter 3 3 12 Blanks 28 Chapter 3 3 13 Peaks 28 Chapter 3 3 14 Normalisation Peaks 28 Chapter 3 3 15 Reference Peaks 28 Chapter 3 3 16 Marker Peaks 29 Chapter 3 3 17 Splines 29 Chapter 3 3 18 Andi Files 30 Chapter 3 3 19 Trace Files ABI MegaBACE SCF 30 Chapter 3 4 Saving and Loading Analysis Procedures Chapter 3 5 DAx Command Line Parameters Chapter 4 Measurement window Scanning for files Chapter 4 1 DAx measurement window Chapter 4 2 Scanning for files Chapter 4 2 1 File Scanning Scan setup menu option Chapter 5 DAx Graphics Windows Chapter 5 1 Zooming in on graphics Chapter 5 2 Graphics Window Menu Commands Chapter 5
204. played in peak or data lists Labels can be moved added and deleted e To add a label double click at the point in the graph where you want to add it then select Add Label from the popup menu The label editing dialog box see below will appear and you can enter a text You can also set font font size attributes and alignment for the label e To move a label place the mouse cursor over it the mouse cursor will change click the mouse and drag the label to a new position e To delete a label click on it In the label editing dialog box click on the delete button item 10 below e To change the text for a label click on it and enter a new text in the label editing dialog box Text label entry 3 21 xi First peak after baseline flattens H B PaT j a Arial viis Text M Background ff x ee a z Delete Label entry dialog box 1 text entry 2 rotate 90 degrees 3 text attributes 4 alignment 5 font and font size 6 foreground and background colours 7 thickness of box around text and arrow lines 8 number of arrow lines 9 display arrow point 10 delete label Labels can have arrows attached typically pointing to details of the data The arrows consist of one two or three lines selected using buttons in the label editing dialog box item 8 above To remove the arrow click the currently depressed button again The first line starts at the edge of the rectangle surrounding the label
205. pline to the current default data set indicated by the data tag with the thick black border and bold text 2 drag spline nodes to new positions or add spline nodes by pressing the Shift Ctrl keys and double clicking at the position a new node should be added refer to chapter Chapter 8 7 for further information on modifying splines 3 use the replace data with spline menu option in the data tag spline menu to replace the baseline data with the spline data You may also use the replace with spline button on the toolbar Steps 2 and 3 may be repeated several times 4 finally hide the spline using the add spline menu option or add spline toolbar button 111 Chapter 10 4 Peaks Find peaks This command invokes a dialog box that starts a search for peaks in selected data sets A baseline needs to have been constructed in order for peak detection to be possible Alternatively a baseline may have been subtracted from a data set Refer to chapter Chapter 10 1 for information on constructing baselines There are two ways in which a peak can be detected e the signal traverses a detection limit Separate detection limits for positive and negative peaks are available in DAx As soon as the signal becomes higher or lower for negative peaks than this threshold a peak will start Peaks do not need to be baseline separated but do need to be threshold separated i e the signal does not need to drop to the baseline level but does nee
206. pplying any additional filtering select a Savitzky Golay filter with filter width 3 DAx contains several methods for filtering data and removing noise from a signal e Moving average filter This filter replaces each data point by the average value of a number of points around the data point A moving average filter can be used synchronously in which case the smoothed value is the average of a number of points 94 preceding the current point and the point itself or asynchronously in which case values both before and after the point to be smoothed are used DAx uses asynchronous smoothing only e Savitzky Golay filter This filter is based on the principle that a polynomial with a certain degree is fitted through the points around each data point The data point is then replaced by the value of the polynomial This method can be used to smooth data and to determine first and second derivatives DAx only allows derivatives to be calculated for data sets that are not already a derivative of some sort e Fourier filtering and Optimum filtering Refer to chapter Chapter 11 1 for details The Data Filter menu option contains a low pass Fourier filter The most important parameter in any filtering process is the Filter Width Filter Width determines the number of points around a point that will be used to smooth its value Higher filter widths lead to smoother signals If the filter width is too high however too much information will
207. procedure so the Load Save calibration menu options are not displayed If the Calibration has been changed but has not been saved to disk yet when DAx is about to terminate you will be asked to confirm the loss of the new calibration Saving the calibration to a floppy disk does not prevent this you must save to hard disk Calibration Stop calibration stops using the calibration and removes it from memory Chapter 17 4 Calibration Track value menu option Use the Calibration Track values menu option to determine if calibrated values are displayed in the coordinate tracking bar when you move the mouse cursor across a graphics window Keep in mind that the horizontal coordinate of the mouse is rounded to the nearest actual data point before the corresponding calibrated value is determined so that calibrated values may not change smoothly as the mouse is moved left to right Chapter 17 5 Calibrated Axes The Data Set Graphics window menu option Calibration Calibrated axis displays graphics where the time axis has been replaced by an axis that contains the calibrated value corresponding to the time coordinates using the current calibration See also Calibrated Axis options chapter Chapter 17 5 1 DAx has the option of converting the horizontal axis of a measurement to calibrated values instead of time coordinates Use 177 the Calibration Axis conversion menu option for this You will be asked to specify the lower and upp
208. proper file name of a disk file data were imported from Chapter 3 3 9 Enslavement For measurements DAx has the option of enslaving channels to other channels Enslaved channels share certain attributes with their owner channels Enslavement will typically be used for multi detector measurements Refer to the Measurement Window manual for details on how to enslave measurement channels DAx extends the principle of enslavement to other aspects of data sets as well Each copy of a data set belongs to the original copy For instance when a data set is filtered and a copy of the original data set is created this copy is a child of the original data 1 The proper file name is the file name without the full path 26 When multiple data sets are imported from a single file such as a trace file chapter Chapter 3 3 19 the data sets are also enslaved When a data set is added to a graphics window by moving it from another graphics window chapter Chapter 7 or by adding it from a disk file chapter Chapter 7 1 2 DAx tries to re establish relationships Data that share the same data set name chapter Chapter 3 3 8 will belong together Refer to chapter Chapter 9 1 to see how data set relationships can be visualised and altered Chapter 3 3 10 The Waste Barrel DAx has a small window that depicts a chemical waste barrel This barrel is used to delete data sets A number of the most recently deleted data sets can be restored
209. ps are used as retention time coordinates in DAx The peak top time of the reference peak is used as the reference time t If no reference peak has been defined the inert time is used The inert time is a sizing parameter cf chapters Chapter 7 7 and Chapter 8 5 e In CE a mobility may be entered as the annotation for the reference peak right click the peak top marker then enter the annotation in the peak popup dialog box Electro osmotic flow will be calculated using the reference peak s apparent and entered mobilities Chapter 10 10 5 Marker Peaks Any peak can be the marker peak for another peak Setting a marker peak for a peak is done using the peak popup dialog box or the peak list window Some automatic analysis methods will automatically assign marker peak names Marker peaks are used to calculate relative times for peaks and to recognise peaks based on their position relative to other peaks internal standards Use the marker peak set up dialog box to set up automatic allocation of marker peaks cf chapter Chapter 18 6 Chapter 10 10 6 Shoulder Peaks DAx has the option of working with small peaks that lie on top of wider peaks The smaller peaks are said to lie on the shoulder of the wider peak The graph below gives an example of two shoulder peaks on top of a shouldered peak 129 DAx 1 0 PP 19 01 96 Volt 0 15 0 10 time s Example of shoulder peaks lying on shouldered peak Notice the m
210. ption This menu option is used to indicate where to scan for files how to analyse them if files should be moved after analysis and how DAx should determine which files have been analysed already Scope Look for files in D harvard8 au File name wild card fsa 2 V Scan subdirectories 3 m Actions Add files to Data List 4 IV Auto analysis Contig 5 m Cleanup IV Move files after analysis le Move files to D harvard8 done E Only scan files since 1 04 2008 19 22 53 OK Cancel 1 Look for files in specifies the location to monitor for the appearance of new files When new files appear they will be opened and possibly added to the data list or analysed 2 The File name wildcard is used to specify which files to scan for Examples are txt and A1000 fsa 2 3 Check Scan subdirectories to scan subdirectories for new files also If move files after analysis is checked subdirectories will be recreated in the output location 4 Use Add files to Data List to create an entry in the data list for each new file that appears 5 Files can be analysed automatically when they appear Check the Auto analysis box to do this use the Config button to specify what type of analysis is to be performed These analysis settings are separate from the measurement and File Open analysis settings that DAx also keeps Analysis results are saved as DAx files either in the current location or in the output location if mo
211. ragged the current coordinates will be displayed in the toolbar at top of the DAx window When the left slider is positioned at the time coordinate of data point M all signal changes that occur within M points are discarded When the right slider is positioned at the time coordinate of point N all signal changes that require more than N points to occur are discarded NB1 The slider controls always pertain to the default data set chapter Chapter 3 3 4 that has the data tag with a slightly thicker border and bold text 14 or more accurately by discarding the extra amplitudes of the frequency data at high frequencies 143 NB2 The slider controls can use a logarithmic scale15 At the left side the slider needs to be moved a lot to cause a small difference in coordinate At the right side coordinates are much closer together This is convenient because low Fourier coordinates need to be set more accurately than high ones NB3 Fourier techniques assume that the data values are distributed roughly around zero To accommodate for this before converting to the frequency domain a linear trend is removed from the data Three possible forms of linear trend are available e straight line connecting first and last data points e straight horizontal line at average signal value e straight horizontal line at initial signal value Use the File Customise gt Data menu option to choose which form is to be used The figures below demon
212. rameters indicate what the lowest highest acceptable value of the baseline signal level is If it is Known that the measurement signal should never be lower higher than a known signal value X X is the logical limit For instance many data acquisition cards are known to have a range of 0 2 Volt The lower limit might be set to 0 01 Volt and the higher limit to 1 99 Volt e maximum acceptable drift This parameter determines what the highest acceptable amount of drift in the signal is The absolute value is used so no distinction is made between decreasing and increasing baseline signal values If the baseline first decreases and then increases back to its original level there is still drift present e maximum acceptable noise This parameter determines the maximum acceptable RMS noise level If baseline parameters are limited to an interval the noise level is calculated using only data within that interval e maximum acceptable signal jump Sometimes the measurement signal level will make a sudden jump to a new level This parameter determines the maximum acceptable value of such a jump This parameter will typically be lower than the limit for drift Chapter 18 15 3 Peak Parameters These parameters inspect the peaks that have been detected in the measurement It is possible to limit the time interval where peaks are inspected by setting starting and ending coordinates e maximum allowable skew If any peak in the measurem
213. raphics windows data set graphics windows 69 graphics windows data set graphics windows 89 graphics windows gradient corrected signal axis windows 158 graphics windows volume axis windows 162 graphics windows molecular weight axis windows 162 graphics windows mobility axis windows 171 graphics windows time corrected signal axis windows 173 graphics windows calibrated axis windows 177 grouping measurements 90 headers 19 highlighting lines in text windows See Selecting Lines highlighting peaks 47 125 133 Horizontal axis unit 25 78 HPLC 15 155 158 Identification Database 189 Identification Database list window 200 inert time 28 64 78 129 136 information dialog box 85 injection volume 199 248 installation 13 Internal Standards 203 kurtosis 137 labels 152 adding 63 leptokurtic 137 Local calibrations 179 182 Local GPC calibrations 168 local Southern method 180 locating peaks 133 log file 17 24 76 86 log of analyses 223 logos 239 main data set definition 25 make child 90 marker peak See peaks marker peaks 129 135 194 definition 29 external markers 197 marker tacks 151 match curve shapes 103 maximum jump 226 maximum noise 226 Measurement data directory See Measurement Window manual measurement data type 15 25 measurement driver selecting 16 measurement drivers 14 measurement set up 64 measurement window 20 33 customising 19 opening 33 MegaBACE fil
214. rated so you would need to use slope limits to recognise them as separate peaks The following table gives an overview Method threshold slope slope peak requiring threshold 4 amp 5 together 4 5 Vv Ivy 6 amp 7 Baene V 6 7 separatey Chapter 10 4 6 Limiting Peak Search Intervals Sometimes you may want to search for peaks only in a limited part of the measurement To do this first construct a baseline for the entire measurement Now zoom in on the part of the measurement in which you want to detect peaks Invoke the Peaks Find Peaks menu option On page 4 of the peak find wizard check the limit search to zoomed window check box This limits the peak search to that part of the data set that is currently visible in the data set graphics window Any peaks already present in the window will be removed but peaks outside of the window will not be affected This makes it possible for instance to search for peaks in two small time intervals and disregard the rest of the data It s also possible to use peak events to limit the peak search to certain time intervals Refer to the next subchapter 119 Chapter 10 4 7 Peak Detection Events Peak detection events are used to adjust the way peaks are detected in certain time intervals Peak events are entered on page 6 of the peak find wizard 5158 4 Dal V Use Peak Events Hit F2 to start 5158 4 Dal1 editing item STAND 1 Da1 STAND 1 Da1
215. re report will be repeated that is additional pages will be added DAx uses Single graph by default if the report definition contains a single data plot item and Separate graphs if more than one data plot item is present The Report Graph per set menu option causes enslaved data sets to be grouped together in single graphs with additional graphs for additional enslavement groups If the report preview has more than one page use the PgUp and PgDn buttons to move between pages Chapter 20 6 4 Customising the Report Preview Window Use the View Attributes menu option to customise the appearance of the preview window Use the View Centimeters menu option to display coordinates in centimeters with the origin at the top left of the page Use the View Inches menu option to display coordinates in inches 244 Chapter 21 Frequently Asked Questions and Troubleshooting Chapter 21 1 Can I search for peaks in a time interval Yes you can Refer to chapter Chapter 10 4 6 for details Chapter 21 2 don t like the way peaks are marked The way peaks are marked can be changed in several ways e go to the data tag peaks menu chapter Chapter 8 3 and use the Mark peaks option to stop marking peaks for the data set If you press the shift key when invoking the menu option all other data sets in the window will also no longer have their peaks marked e use the File Customise gt Plotting Peaks option to change the width a
216. rea 103 Instead of specifying a target area the Derive from first data set encountered box can be checked to use the area of the first data set that is overlaid as target area The absolute value of peak areas is used If the 75 smallest peaks are used but there are only two peaks then all of the area of the smallest peak and half the area of the remaining peak will be used In general if the given percentage of the peaks is not a whole number the left over fraction of the next smallest peak will be included in the calculated area Scaling will also be applied in such a way that the smallest signal value encountered will be 0 e Vertical scaling will be applied in such a way that the total area of corresponding named peaks in the data sets with the specified trace type will be normalised to the specified target area Further data sets will be sized in such a way that the named peaks that occur in both the current and the first data set will end up having an equal total area This means that data sets can be sized correctly even if they do not have the exact same named peaks Data sets that are not the specified trace type will be scaled using the most closely related data set of the specified trace type If no closely related data set of the specified trace type is found the data will be excluded from the overlay Instead of specifying a target area the Derive from first data set encountered box can be checked to use the area of
217. rected axis 173 CE Track mobility 171 Data De Convolution 100 Data Arithmetic 64 97 Data Average 98 253 Data Correlate 99 Data Derivative 94 Data Filter 94 Data Integrate 96 Data Interpolate 93 Data Overlay 102 Data Plot select 91 Data Reduce 92 Data Relationships amp Sorting 89 Data Remove trend 94 Data Set colours 91 Data Slice 94 Data Sort 99 Data List Add data 232 Data List Assert data 232 Data List Close list 232 Data List Display list 232 Data List Display statistics 232 Data List Find data 233 Data List Load list 232 Data List New list 232 Data List Save list 232 Data Tag Menu 71 Data Tag Peak Menu 73 Data Tag Sizing Menu 76 Data Tag Spline Menu 79 Edit Copy 45 58 Edit Copy Graph 38 Edit Create Metafile 38 Edit Find 233 Edit Find next 233 Edit Invert selection 58 Edit Redo 59 Edit Select 57 233 Edit Select all 45 57 Edit Sort list 233 Edit Undo 59 File Close 53 File Customise 15 56 File Exit 56 File Export 43 File Export selected 43 File Load analysis procedure 56 File New 33 48 File Open 48 File Open data group 52 File Operator 23 56 File Preview report 54 243 254 File Print 38 43 54 File Print report 54 File Printer setup 55 File Save 52 File Save analysis procedure 5
218. reference peak in the peak list window the default reference mobility as entered in the CE Capillary Parameters dialog box will be used instead If no default reference mobility has been entered for a data set the default reference mobility last assigned to any data set will be used Chapter 16 2 CE Capillary Parameters Dialog Box This dialog box is used to enter parameters about the capillary being used in a CE measurement It is invoked using the CE Calibrate menu option Length of capillary m ft Z al Length till detector m o s pJ Voltage kv 10 000 pA Default Reference Mobility 10 9 m s 0 00 A Parameters will be stored for selected data sets and as global defaults Cancel Help 1 The parameters entered in this dialog box will be stored with the data sets selected here Other data sets will retain their old CE parameter values 2 The default reference mobility is used if a reference peak has been marked but no effective mobility has been entered as its annotation Chapter 16 3 CE Load calibration CE Save calibration menu options CE capillary parameters may be saved to disk using the CE Save calibration option They may be loaded from disk using the 170 CE Load calibration option f the Separate Parameter Files item in the File Customise gt Extensions dialog box is unchecked CE parameters can only be saved as part of an analysis procedure so the Load Save calibration
219. released a new peak will be added just before or just after the existing peak The peak will start or end at the point where the mouse was released DAx will search for the peak top of the new peak automatically Pressing the Ctrl key and double clicking the mouse button also adds a peak If this is done inside an existing peak a shoulder peak will be added Double clicking the mouse button without pressing the Ctrl key will display a popup menu This menu includes the option to add a peak near the clicked coordinate If the coordinate is inside an existing peak the menu will include the option to split that peak at the clicked coordinate Pressing the Ctrl key then clicking the mouse and dragging a rectangle displays a popup menu from which you can add a peak If you also press the Shift key a peak is immediately added without first displaying a popup menu The peak will have boundaries equalling the rectangle that was dragged The peak top is searched automatically Peaks can be added inside other peaks this way You will be asked if the new peak should be a separate peak or a shoulder peak Peak begin markers cannot be dragged beyond the top marker of the previous peak nor beyond the top marker of the current peak If a peak begin is dragged beyond the end marker of the previous peak the end marker of the previous peak will be adjusted in such a way that the peaks will not overlap with the exception of shoulder peaks cf chapter Chap
220. river options You can choose to automatically open the measurement window each time DAx is started f you use DAx to analyse data files generated using third party software you will not need the measurement window You may also select which measurement driver is to be used The driver should correspond to your measurement hardware It can only be changed when no measurement window is open Chapter 3 1 3 File Customise gt GLP Good Laboratory Practice dictates among other things that all measurements always be saved and that no measurement can overwrite an other measurement However these aspects may be customised to suit your needs The following customisation aspects are available e Ask if aborted measurements need to be saved Measurements that end because of a stop trigger or time out are always saved regardless of this setting e Overwrite Raw Data If this option is enabled operations that normally require Modified Data can also be performed on original measurement data e Overwrite Files If this option is enabled files can be overwritten Naturally DAx asks for confirmation when an existing file is to be 16 overwritten If the option is not enabled no existing file can be overwritten e Keep Logfiles Normally all irreversible operations on data and some others such as finding peaks are logged with the data You have the option of not keeping the log file e Keep error logs Normally all error messages
221. rly baselines chapter Chapter 3 3 11 29 Chapter 3 3 18 Andi Files The Andi file format sometimes also referred to as the AIA file format is supported by a wide range of data analysis software DAx supports both reading and writing of this file format Use the File Open menu option to load Andi files use File Save As to write Andi files In both cases use the list Files of Type item in the dialog box to select Andi files Chapter 3 3 19 Trace Files ABI MegaBACE SCF Some versions of DAx can read gene analysis files in the ABI Genescan Amersham MegaBACE and SCF formats When files of these types are opened a dialog box will be displayed which allows the user to set a number of trace analysis parameters such as which colours to use to display the traces and which colour separation matrix to apply to the data if any Use the File Open menu option to load trace files Use the list files of type item to select ABI Genescan Amersham MegaBACE or SCF files A separate quick start manual is available that explains how to analyse trace files Please contact your supplier for more information Chapter 3 4 Saving and Loading Analysis Procedures The File Load analysis procedure and File Save analysis procedure menu options let you read and create analysis procedure files These files can contain a number of parameters Not all parameters need to be included in an analysis procedure file when you create
222. rogramme is entered as one of the gradient parameters Based on this parameter the corresponding axis in the graphic window will be correctly labelled Chapter 14 1 Defining Gradients step by step To be able to calculate gradients DAx needs to know a number of parameters Use the HPLC GC Set gradient menu option to enter these parameters or use the HPLC GC Load gradient menu option to load parameters from disk Chapter 14 2 HPLC GC Gradient Parameters Dialog Box This dialog box is used to enter parameters about the gradients being used in a HPLC or GC measurement It is invoked using the HPLC GC Set gradient menu option Each gradient step is defined by four parameters 155 signal percentage signal ee ae 0 1 where time and time are the start and end time coordinates for the gradient and signal and signal are the start and end signal values In addition to these values gradient percentage values or programme temperature values can be entered in DAx They can be plotted as a separate curve using the HPLC GC Show percentages or HPLC GC Show temperature programme menu options HPLC Gradient GC Temperature Programme Parameters 2 xi PIPO TXT Number of steps h x IV Derive signal values Y0 Y1 from data ME Piro txt t0 s eo A wv A Ar a vw A aff A M Gradient Type 7 t1 s hsa PA rT oY lon A rT rT Y GC Temperature Programme 3 y1
223. rs menu option To display the analysis assay use the Peaks Display analysis assay menu option in the data tag peak menu cf chapter Chapter 8 The Display Analysis button on the toolbar displays the analysis assay for the default data set in a graph window the default data set has a data tag with a thicker border and bold text Not all versions of DAx contain analysis assays Chapter 18 15 1 Assay Weighting The principles of assay weighting are as follows e each individual parameter starts out at 100 quality e if a parameter is better than the specified suspect value its quality is 100 e if a parameter is worse than the specified bad value its quality is 100 minus the specified parameter weight e if a parameter is between the specified suspect and bad values quality is determined as quality 100 _ parameter suspect bad suspect e total assay quality is determined as the product of individual qualities If individual qualities are 80 70 and 75 overall quality is 80 70 75 42 225 Chapter 18 15 2 Baseline Parameters These parameters inspect the baseline of a measurement It is possible to limit the time interval where the baseline is inspected by setting starting and ending coordinates It is possible to enter baseline limits as a relative value In this case the signal values of the original measurement are taken to range from 0 100 e low high limit signal level These pa
224. s Select allocation methods and limits IV If slant gt 3 0 Al IZ If skim gt joo p4 IV If relative width gt 15 0 Al Select all lt a r eect IV IF relative area gt 3 0 9 A epey Noa ci Fifth page of the peak find wizard Sets up automatic shoulder peak detection cf chapter Chapter 10 10 7 Depending on your version of DAx the peak find wizard may have additional pages Those are discussed in chapters Chapter 10 4 7 Chapter 10 4 9 and Chapter 10 4 10 Chapter 10 4 1 1 Selecting data sets The Select all button in the peak find wizard can be used to select all data sets that contain a given string of characters in their names Enter the string or choose one of the pre defined strings listed Check deselect to deselect all data sets that contain the string If the Alt key is pressed when the Select all button is clicked all previous selection will be removed before the new selection is applied Chapter 10 4 2 Area Limit Sometimes DAx will find a great number of small peaks in a data set even though there may only be a few components present 116 One way to find fewer peaks would be to set higher peak detection limits for instance a higher threshold If automatic thresholds are being used the automatic threshold might be set to equal 10 times the noise level instead of 4 However there is an alternative If you are not interested in any component with a relative concentration of 1 or if the anal
225. s There are Data Value Table windows and Peak List windows chapter Chapter 10 11 DAx GPC has a GPC calibration list window chapter Chapter 15 6 DAx Calibrations has a calibration list window chapter Chapter 17 8 1 All text windows share the way in which they are used as well as a number of menu commands These common attributes will be discussed in this chapter Chapter 6 1 1 Selecting Lines Lines may be selected in a text window by dragging the mouse cursor or by using Shift cursor key to navigate the text window chapter Chapter 6 1 5 Certain menu commands operate on selected text For instance it is possible to copy the selected text to the clipboard Chapter 6 1 2 Editing Text Window Contents Some text windows have the option of editing their contents If this is the case click the mouse cursor on the item you want to edit to display an edit control Refer to chapter Chapter 6 1 5 for information on how to move the edit control to a new position Not all columns are always editable Sometimes only one column can be edited Some columns may not display a free form edit control but a checkbox Click the checkbox with the mouse or press the space bar to toggle the value of the item Some columns display a drop down combo box Hit the F2 key to be able to use the cursor keys to change the selection Hit the F2 key again to be able to move between cells with the cursor keys Some columns display a small
226. s To remove criteria select one or more used criteria then click Deselect to remove them to the list of available criteria Finally click sort to sort the data sets All data sets will be sorted not just the selected sets If one or more running measurements are present in the graph window data set sorting is disabled Chapter 9 2 Data Plot select The Data Plot Select menu option is convenient to do large scale adjusting of data set visibility To show or hide individual data sets 2 x the data tag menu chapter oO C DAxDTest4SiTEST3 DAX C DA Chapter 8 2 is more convenient STAND 1 Da1 MEJ stanp 1 ba1 5158 4 Da1 The Select all button can be used mo 5159 4 Dal to select all data sets that contain a given string of characters in their names Enter the string or choose one of the pre defined strings listed Check deselect to deselect all data sets that contain the string Select all 1 Desel If the Alt key is pressed when the al Gees neo Select all button is clicked all previous selection will be removed before the new selection is applied Chapter 9 3 Data Set colours This menu option is used to change the colours of data sets typically when many colours need to be changed at once To 91 change the colour of an individual data set the data tag menu chapter Chapter 8 2 is more convenient Data set colour Selection 2 xi Cx Aea e 2 Assign colour to select
227. s and multiple peak parameters e Binning map chapter Chapter 18 12 A window displaying a coloured map representation of a binning sheet Chapter 3 3 Terminology For a complete understanding of this manual a number of terms need to be explained Chapter 3 3 1 Analysis Procedures DAx uses an Analysis Procedure file to save filter parameters baseline construction and peak find parameters GC HPLC CE GPC parameters as well as qualification quantification and assay parameters This means that a single Analysis Procedure file can fully define the way in which measurements are interpreted Chapter 3 3 2 GLP GLP stands for Good Laboratory Practise a system of rules and regulations intended to formalise the way in which measurements are performed in a laboratory Some GLP options can be switched on and off using the File Customise gt GLP menu option e Name registration is required for any user An operator name has to be entered when the program is started The operator name may be changed using the File Operator menu option available in all menus e Data operations that cause irreversible changes can only be performed on Modified Data data sets in order to ensure the original raw data will always be preserved Irreversible operations must therefore be performed either on Modified Data or in such a way that a new copy of the data set is formed All irreversible operations have the option of creating a new copy of a da
228. s at the start and end of each slope The markers may be hidden using the Mark gradient menu option The markers may be dragged to change the gradient Dragging a marker past the previous or next marker will remove the gradient Pressing the Ctrl key while dragging a marker will add an extra gradient There is a maximum of ten gradient steps Click the right mouse button on a gradient marker to display a popup menu This menu lets you copy the gradient to other data sets This is useful when you have manually adjusted the gradients in a single data set and want to use the same gradients in other data sets The undo option in the tag gradient menu undoes moving adding or deleting operations Press the Shift key while clicking the Show gradient or Mark gradient menu options to show or hide gradients or markers for all data sets in the graphics window 158 Chapter 14 8 1 Modifying Gradient Percentages Programme Temperatures Gradient Percentage or Programme Temperature values cannot be changed using the mouse Instead use the tag menu to display a data set information dialog box chapter Chapter 8 10 Click the HPLC button This displays an extra dialog box in which gradient percentages or programme temperatures can be changed 159 Chapter 15 DAx GPC Gel Permeation Chromatography Extensions If the menu option GPC is not present you need to use File Customise gt Extensions and check GPC Options Not all versions of
229. s has been replaced by an axis that contains either the eluted volume at a given time coordinate or the molecular weight that corresponds to that volume using the current GPC calibration 162 Chapter 15 4 1 Molecular Weight Axis Options There are several ways in which data can be plotted with a molecular weight axis All of these are selected using View menu options e Either a linear or logarithmic scale can be used along the horizontal molecular weight axis Logarithmic axes can either be marked linearly or logarithmically e Either a differential or a cumulative distribution or both can be plotted To plot a cumulative distribution the baseline should be subtracted from the data chapter Chapter 10 2 before plotting e Either the concentration abundance or the weight can be plotted along the vertical axis e It is possible to automatically subtract baselines from the data Baselines are plotted as straight lines at 0 level e Cumulative distributions can be plotted with absolute values or normalised as a percentage In a normalised plot the total area of all peaks present in the data correspond to 100 Data points outside of peaks are ignored When no peaks are present a straight horizontal line is plotted e Distributions can also be normalised on height so that the highest point in each distribution corresponds to 100 e Mw Mn Mp and Mz values for peaks can optionally be marked in the graph Chapter 15 4 2 Conv
230. s to be a numeric value indicating the calibrated value for instance a number of base pairs In both these cases the values molecular weight or calibrated value can also be entered as the peak name If at most one peak annotation has a numerical value DAx assumes that value to be a molecular weight or calibrated value and the peak names will be used to derive calibration values Annotations are entered in the peak list window by clicking in the annotation column for the appropriate peak Finding the peak is facilitated by the fact that as you move the mouse cursor over a peak marker in the graph window the line in the peak list window that pertains to that peak is highlighted The menu option Peaks Fill annotations can be used to copy the contents of any column in the peak list window into the annotation field NB There is a difference between a peak annotation and a peak label An annotation is a text you enter about a peak A label is any peak value that is entered or calculated by DAx displayed above the peak To select which peak values must be used as labels use the File Customise gt Plotting Peaks option There are three more columns in the peak list window that can be used to enter values the name and the concentration columns and the peak type column 134 Chapter 10 11 5 Setting Marker Peaks The peak list window can be used to quickly set the marker peak name s to be used by one or more peaks Highlight the p
231. se e selecting the qualifying parameter e setting a tolerance for the value of the qualifying parameter Intermediate Qualification Pass causes the Identification Database to be built in two stages First a qualification database is built This is then used to qualify peaks Only then is the quantification database built This means that you do not have to enter component names for each peak DAx will try to establish the component name Obviously you do need to enter concentrations for every peak that is to be included in the quantitative calibration 192 Use the OK button to create a new database or add entries to an existing database Check the Add to existing calibration box to add quantitative calibration points to a calibration The Change Qualification button is used to change the qualifying coordinate and set new tolerances without adding or removing entries in the database The Quantification coordinate and Calibration Curve type selected in the dialog box are initially applied to all quantitative calibrations for new components Additional points may be added to existing quantitative calibrations but the calibration type will not change Refer to chapter Chapter 18 8 for details on how to change the calibration type later Finally concentration limits can be set Concentrations outside the limits will still be calculated but will be flagged in the peak list with an L or an H The concentrations entered are initially appl
232. sis Stop quality assays stops performing analysis assays Use Analysis Load assay parameters or Analysis Edit assay parameters to start performing assays again Chapter 18 15 9 Displaying an Analysis Assay Use the Peaks Display analysis assay menu option in the data tag peak pop up menu to display the analysis assay for a data set The Display Analysis button on the toolbar displays the analysis assay for the default data set in a graph window the default data set has a data tag with a thicker border and bold text 229 Chapter 19 Data Lists If the menu option Data List is not present you need to use File Customise gt Extensions and check Data Lists DAx has the ability to compile a list of measurements either by inspecting files on the hard disk or by adding measurements as soon as they stop running The Data List can be used to quickly overview all measurements Measurements can be loaded directly from the list The list can be sorted on a variety of properties A search facility lets you quickly find a specific measurement The search facility can also be used to select all measurements that fulfil the search criteria Data Lists can list the following properties File name File data amp time Number of files in a data set File type Data Set name Operator name Measurement date amp time Acceptance measurements can be okay suspect or bad or have failed controls Data type Number of data poin
233. small peaks Available quantifiers are height area normalised area migration time corrected area and migration time corrected normalised area e use pattern Determines if the meta analysis entry should satisfy a meta analysis pattern see below e pattern quantifier e pattern target Meta analysis is set up by displaying the meta analysis list using Analysis Edit meta analysis To change multiple entries in this list at once select the entries right click and choose the Configure selected menu option This displays the meta analysis configuration dialog box When the meta analysis list is edited entries with the same identity of species are always kept together for instance when you change the identity of an entry the entry may be moved up or down so that it is grouped with other entries with the same identity User defined bins chapter Chapter 18 12 3 and marker peaks chapter Chapter 18 6 can be copied and pasted into the meta analysis list The meta analysis list is part of the Identification Database it cannot be loaded or saved separately Meta analysis typically looks for multiple peaks per species These peaks do not all have to be present in a single data set They can also be located in different data sets in the same enslavement group cf chapter Chapter 3 3 9 or in any data set in the same graphic window In DNA analysis typically the middle option should be used Use the Analysis Configure meta analysis
234. sorted see below 3 Data sets can be grouped automatically based on the files they were originally loaded from The Relationships dialog can be used to change the relationships between data sets and to change the order of data sets Selecting a data set and clicking the gt Make Child button will make the data set owned by the preceding data set The primary use of this button is to group measurements If one of those measurements is a blank measurement it needs to be grouped with the other measurements in order to be used as a baseline Remember to mark it as a blank measurement using the data tag menu The lt Emancipate button breaks the ownership relationship between a data set and the preceding data set You can select data sets then press the Bring to Top button the Move Up button the Move Down button or the Drop to Bottom button to change the order of the data sets Data sets can be moved up or down by clicking on them and dragging them up or down to a new location Dragging them to the left will emancipate them dragging them to the right will enslave them 90 Chapter 9 1 1 Sorting data sets Data sets can be sorted using one or more criteria in ascending or descending order Under available criteria select the criteria to be used then click the Select button to add them to the list of used criteria To change the order of the used criteria select one or more and click the top up down or bottom button
235. strate the principle of Fourier filtering DAx 1 0 PP 18 01 96 18 55 34 Volt Original Data 15 use the File Customise gt Data menu option to change to a linear scale 144 DAx 1 0 PP 18 01 96 18 56 33 Volt Data with high frequency changes removed DAx 1 0 PP 18 01 96 18 56 08 Volt l l e I l l l l L Corns air es le ee A e Ts l l 4 i l l l l l 4 aa a E ee a ae E Caaan i a a A aa ae a eT l l l l q l l l Ore uerus Merrie as Ra Mls ees ek ate EEN EET EE a cla a A 4 l l 8 a ee a p y 0 500 1000 1500 Data with low frequency changes removed DAx can also use Fourier filtering to derive baselines In this case the applicable frequency interval does not have a limit for low frequencies since signal drift should typically be included in the baseline A baseline is constructed by discarding all frequency data that makes the signal change faster than some percentage of the total horizontal scale 145 Chapter 11 2 Deriving Fourier Filtering Characteristics DAx has the ability to determine how the signal to noise ratio for a data set changes as a function of the applied amount of filtering i e as a function of which frequency data are discarded For best results make sure that peaks have been detected in the data set cf chapter Chapter 10 Then invoke the Fourier Derive characteristics menu option in the data tag menu use the tag m
236. t all button is clicked all previous selection will be removed before the new selection is applied 98 Chapter 9 12 Data Correlate If the menu option Data Correlate is not present you need to use File Customise gt Data and check Data Correlation This command invokes a dialog box that is used to determine correlation between two or more data sets Two types of correlation determination can be used e correlation as a function of horizontal shift between data sets This is used to determine if data sets can effectively be considered as equal except from a shift along the horizontal axis A new data set is created that plots the correlation between the entire data sets as a function of horizontal shift e correlation between corresponding intervals of data sets A sliding window with a set interval width is moved across the data and the correlation is determined repeatedly A new data set is created consisting of all correlation values determined in this way The width of the sliding window is entered as a parameter in the dialog box The new data set is displayed on a separate vertical axis to make scaling more convenient Obviously at least two data sets need to be selected for correlation determination If more than two data sets are selected each possible combination of two data sets is correlated However as an option only data sets inside the same enslavement group can be correlated This might typically be used
237. t concentration 139 concentration 139 cross over peaks 121 customising finding 17 customising plotting 18 definition 28 events 120 first peak moment 136 fixating peak names and concentrations 75 highlighting 47 125 133 kurtosis 137 labelling 135 labels 73 256 limited searches 119 locating 133 manually adding peaks 126 manually modifying peaks 124 marker peak 29 marker peak 194 marker peaks 129 modifying peaks 124 moment 136 naming peaks 127 normalisation peak 28 85 128 137 normalising peak widths 120 number of theoretical plates 137 peak highlighting 133 peak list window 74 reference peak 28 85 128 134 138 171 reference peak 194 reference peak 198 relative concentration 139 removing 133 resolution 137 setting peak types 127 shoulder peak area calculations 131 shoulder peaks 126 129 skew 137 slope limits 117 too many peaks 116 pen thickness 39 plates 137 platykurtic 137 point indices 25 Point to point noise See noise See noise preserving data in binning sheet 215 previewing reports 54 243 print 38 43 84 printer setup 55 printing 54 printing reports 54 programme temperatures 156 157 programme temperatures 159 qualifying peaks 192 quality control 193 198 quantifying peaks 192 quantitative calibration 198 quantitative window 202 quantitative calibration list window 201 reading data files 48 recalculate 75 124 reference peak 13
238. t is added to a graphics window by moving it from another graphics window by dragging the data tag chapter Chapter 1 or by adding it from a disk file chapter Chapter 7 1 2 DAx tries to re establish relationships Data that share the same data set name chapter Chapter 3 3 8 will belong together The Relationships dialog box shows the relationships that exist between different data sets in a window Owned enslaved measurements are displayed with a horizontal displacement 89 Data Relationships ote Data set sorting all data sets Aana TP53BP_BTMCL_01N_NO12_POP_LOO1 LO01 eee oan ani ra E TPS3BP_BTMCL_O1N_NO12_POP_LOO1 L001 _Bing to Top Plate name 3 TP536P_BTMCL_01N_NO12_POP_LO02 L002 Move Up ell row A H of JIB TP53BP_BTMCL_O1N_NO12_POP_LOO2 L002 12 COMM TP53BP_BTMCL_01N_NO12_POP_LO03 L003 o LIE TP538P_BTMCL_OLN_NO12_POP_LO03 L003 CM TP538P_BTMCL_01N_NO12_POP_LO04 LO04 Pi rase Ewa OB TP53BP_BTMCL_01N_N012_POP_L004 L004 lt Emancipate Select all CME TP53BP_BTMCL_01N_NO12_POP_LO05 L005 JB TP53BP_BTMCL_01N_NO12_POP_LOOS LO0S CON TP53BP_BTMCL_01N_NO12_POP_LO06 L006 JB TP53BP_BTMCL_01N_NO12_POP_LO06 L006 Move Down Drop to Bottom Use criteria in order Title name Press the Ctrl key to select Group automatical multiole items y Bl e 1 A yellow B indicates a data set is a baseline chapter Chapter 3 3 11 2 Data sets can be
239. ta set An example is filtering chapter Chapter 10 23 Operations that modify the data but are considered reversible and are therefore allowed to be performed on original data sets are horizontal and vertical moves and sizing chapter Chapter 7 7 and flow rate adjustment e DAx will not overwrite an existing data file with a new data file and will warn the user when an attempt is made to do this e All data operations are logged in a log file Log file entries are dated and carry the name of the operator e All error messages and important warnings as well as program start and stop and changes of the operator name are logged in a message log The file menu contains the option View message log which lets the user inspect the error log New logs are started automatically when a log file becomes larger than 50 000 bytes Use the File Customise gt GLP menu option cf chapter Chapter 3 1 3 to turn off log files or to force the creation of a new log file e Data files and parameter files have a checksum attached If the file is changed by any program other than DAx the checksum will become invalid and a warning message will be displayed e DAx has the ability to store copies of the parameters that were used to analyse data with the data Use the File Customise gt Extensions menu option to switch on Store parameters with data You should also switch on Local Calibrations and Local GPC Calibrations Chapter 3 3 3 Data Se
240. ta tags are used for notifications in graphic windows e They flash blue when a popup menu appears that pertains to the data set such as when the right mouse cursor is clicked on a peak marker e They also flash blue when a data set is changed such as when a peak marker is dragged to a new location e Data tags are depressed when the mouse cursor is placed over the data set s curve e They are also depressed when the mouse cursor is placed over a peak tack or spline marker e Data tags are depressed in stacked views created using the Data Stack data menu option when the mouse cursor is in the vertical range of one or more of the stacked data sets The last three of these are tracking behaviour Tracking behaviour is suppressed when the Shift key is held down A thick black line around a tag and the use of bold text indicate this is the tag for the default data set chapter Chapter 3 3 4 which is used in certain operations A yellow padlock symbol on the right side of the data tag indicates that the data set has fixated peaks A blue padlock symbol indicates read only data A yellow B at the left side of the tag indicates the data set is being used as a baseline A blue B at the left side of the tag indicates the data set has been marked as a blank measurement Data tags for data sets that are not currently plotted are displayed using a lighter background colour By default data tags are located at the top of graphics windo
241. tal axis can be converted to calibrated values Click on Convert axis to calibrated to configure the conversion process Finally automatic analysis can convert the horizontal axis to mobility values The Identification Database should be set up to recognise a reference peak to use in determining mobility values Click on Convert axis to mobility to configure the conversion process Chapter 7 1 2 2 Dropping Files DAx supports dropping files from the file manager onto its client area to open them Dropped files are always loaded into a new graphics window Chapter 7 1 2 3 Opening Groups The File Open data group command is similar to File Open but will scan a directory tree for files with the same names as selected files These files can then all be opened at once Chapter 7 1 3 File Save File Save as This command saves one or all of the data sets in the graphics window to file A list of the data sets in the window will be displayed from which the user may then select the sets that are to be saved Data sets that are enslaved to each other cannot be saved separately so when one is selected all are selected 52 PE 21x Select Data Save in apex o t fe c EJ STAND 1 Da1 STAND 1 Da1 LCI WINVOLT JTEST2 DAX Ge TEST4 DAX MB s158 4 ba1 L WVBLANK TATEST3 DAX IETEST5 DAX MEI 5158 4 Dal1 iIpipotest dal eltest3b dal eltest5b dat He TEST1 dal fe Test3c dat Geltest5c dat Ha TEST1 DAX Geltest3d
242. ted for the data set Spline Curve the data set was generated from a user modifiable spline Filtered Moving Average data were smoothed using a moving average filter or a Savitzky Golay filter 1st or 2 Derivative Savitzky Golay first or second derivative of data set with respect to the horizontal axis was determined using a Savitzky Golay technique De convolution curve was de convoluted chapter Chapter 9 14 using a theoretical curve or another data set Peaks present peaks were searched in the curve Fourier Transform a Fourier transform technique was used to filter the data Spike Removal spikes were removed from the data chapter Chapter 11 4 85 Moving Average Baseline curve data points were calculated from a data set DerLim Baseline Fourier Baseline Average below average Modus baseline Constant Arithmetic Curve Arithmetic Data Reduced Sorted Data Curve Time Moving Curve Time Sizing Flow Rate Adjustment Curve Vertical Moving Curve Vertical Sizing Gradient Set Gradient Subtracted Blank Measurement Sliced Data by using the moving average baseline derivation technique chapter Chapter 10 1 the DerLim algorithm is generally the best algorithm to use to derive baselines especially baselines that are not strictly horizontal curve data points were calculated from a data set by using the Fourier baseline derivation technique horizontal baseline horizontal baseline data were modified by an ar
243. tention time of the component minus the peak top time of a marker peak cf chapter Chapter 3 3 16 The marker peak is selected in the peak popup dialog box and can be any named peak retention time of the component divided by the retention time of the marker peak index between markers relative position of the peak top between two width WHM sDev LWHM LsDev RWHM RsDev Area Area Fraction 136 named marker peaks end time begin time s peak width at half height s standard deviation of peak time with respect to signal gt signal time m ye y signal m the first peak moment peak width of left side of peak at half height standard deviation for left side of peak peak width of right side of peak at half height standard deviation for right side of peak area of peak area of peak divided by total absolute area of all active peaks value s calculated as o with Norm Area normalised area area of peak divided by area of normalisation peak or area of highest peak if no normalisation peak has been marked Area Marker Area area of the peak divided by the area of a marker peak The marker peak is selected in the peak popup dialog box and can be any named peak Height Marker Height height of the peak divided by the height of a marker LPeakFract RPeakFract asymmetry plates net plates stat plates peak area of left side of peak divided by area of peak area of right side of peak divided
244. ter 10 10 6 Peak end markers cannot be dragged beyond the top marker of the next peak nor beyond the top marker of the current peak If a peak end is dragged beyond the begin marker of the next peak the begin marker of the next peak will be adjusted in such a way that the peaks will not overlap All of these peak edits may be undone Type Alt BackSpace to do this Typing Shift Alt Backspace redoes the edit Undo redo is also available from the data tag peak menu cf chapter Chapter 8 3 126 Chapter 10 10 1 Peak Popup Dialog Box When the right mouse button is clicked on a peak top a dialog box is displayed The dialog box can be used to enter a name type concentration and annotation for the peak It can also be used to mark the peak as the reference peak or the normalisation peak or to activate or deactivate the peak see also chapters Chapter 10 10 2 through Chapter 10 10 4 Finally the name s of the peak s to be used as marker peak for the peak can be entered A faster way to set marker peak name s for a number of peaks is by selecting the peaks in the peak list window clicking the right mouse button and using one of the Set marker peak menu options Items that have been entered or changed by the user are displayed in red in the dialog box Chapter 10 10 2 De Activating Peaks Peaks can be either active or inactive Inactive peaks are not taken into consideration when area fractions are calculated To de ac
245. ter 12 1 The Waste Barrel Pop up Menu The waste barrel also has a menu The menu is invoked by clicking the right mouse button on the waste barrel Waste Barrel Cursor Restore Items Destroy Contents Help Chapter 12 1 1 Waste barrel cursor Select the Waste barrel cursor item from the pop up menu The mouse cursor now turns into a portable waste barrel Click a data tag to delete the data set To return the mouse cursor back to normal click any part of the screen that is not a data tag or graphics window Chapter 12 1 2 Restore items Select the Restore items menu option to display a list of the contents of the waste barrel Select the items you want to be 149 restored into a Data Set Graphics window chapter Chapter 1 Data sets that are restored will be removed from the waste barrel If more data sets are selected than will fit in the graphics window they will be left in the waste barrel Chapter 12 1 3 Destroy contents Select the Destroy contents menu option to remove all data from the waste barrel Chapter 12 2 Moving the Waste Barrel The waste barrel may be moved across the screen by clicking the left mouse button on it and dragging it 150 example tack reo Chapter 13 Remark Tacks and Labels DAx has the ability to add tacks and labels to a data set Both are used to add descriptive texts at a selected time coordinate Tacks are attached to the data curve whereas labels have freely sel
246. ters Chapter 18 7 Chapter 18 6 Horizontal coordinates for all but the first data set will be adjusted in such a way that the shapes of the curves match best A trace type can be specified if it is data sets with the specified trace type will be matched and the other data sets will be stretched and moved an equal amount A maximum amount of stretching and moving can be specified The interval in which curve shapes are matched can be specified otherwise the entire curve is matched Possible vertical adjustments No adjustment Vertical scaling will be applied in such a way that all data sets will have values running from 0 to 1 vertical unit Vertical scaling will be applied in such a way that the top of the normalisation peak in each data set has its baseline at O vertical units and its top coordinate at a specified target height Instead of specifying a target height the Derive from first data set encountered box can be checked to use the height of the first data set that is overlaid as target height If no normalisation peak has been defined the reference peak will be used If no reference peak exists either the highest peak will be used Vertical scaling will be applied in such a way that a given percentage smallest peaks in the data set will have a specified target total area For instance if there are four peaks and the 75 smallest peaks are used then the total area of the three smallest peaks will become the target a
247. tical axis The latter option requires that the data have a time horizontal axis not e g an index axis If a baseline is available for the data it will be subtracted 172 Chapter 16 6 Time Corrected Signal Axes The menu option CE Time corrected axis displays graphics where the signal values have been divided by migration times Only data sets that have a time horizontal axis as opposed to e g an index axis can be included in the graph If a baseline is available for the data it will be subtracted 16 This differs from a mobility axis with migration time corrected signal values in that the original horizontal axis will be plotted not a mobility axis 173 Chapter 17 DAx Calibrations If the menu option Calibration is not present you need to use File Customise gt Extensions and check Calibrations Not all versions of DAx contain this option The Calibrations version of DAx has a number of extra options that make it possible to use the program to determine calibrated values from chromatograms or electropherograms The procedure is to find a relationship between a measurable parameter and the calibrated value For instance the number of DNA base pairs may be related to the relative migration time Possible calibration parameters are Peak top time Peak moment Peak top time offset Capacity factor Peak elution volume Apparent mobility Effective mobility NB Peak top time offset is determined using a refere
248. time date and time the data were measured accepted acceptance state of the data 241 points frequency ordinate unit description description2 platename wellname totalconc dilution analysis procedure gpc parameters number of data points measurement frequency Hz ordinate name typically Volt ordinate unit typically V first descriptive line second descriptive line name of titer plate trace files only name of well trace files only total chemistry value of the data file name of the analysis procedure used to analyse the data file name of the GPC parameters that were used to analyse the data may be identical to analysis procedure if GPC parameters are contained in the analysis procedure calibration parameters file name of the calibration parameters used to analyse the data identification database file name of the Identification field name n extra field n Database used to qualify and quantify the data n 1 10 Extra field name Use File Customise gt Fields Setup to enter names for these fields n 1 10 Extra field value Chapter 20 5 Customising the Report Editor s Appearance Use the View Attributes menu option to customise the appearance of the Report Editor Specifically this menu option is used to show or hide grid lines 242 Chapter 20 6 Previewing Reports Reports can be previewed on screen using the Report Preview report menu option A report preview
249. ting derivatives 94 calibrated axes 177 calibrated axes 21 calibrated axes 182 Calibration 51 calibration curve 22 calibration quality 228 calibration type 175 calibration window 37 calibrations automatic calibration derivation 181 calibration curve window 182 calibration list window 181 calibration windows 181 creating calibrations 176 global Southern method 180 local calibrations 182 local Southern method 180 selecting the calibration type 175 Calibrations 15 175 capacity factor 28 129 136 Capillary Electrophoresis See CE CE 15 37 129 134 169 173 changing gradient percentages 159 changing programme temperatures 159 channel config dialog box See Measurement Window manual Clearing analysis results 75 clipboard 20 38 41 44 45 58 84 closing windows 53 colour views 62 65 columns in peak list windows 135 combining peaks 132 command line parameters 31 comparison sheet 70 204 concentration ranges 193 198 Concentration to weight conversion 163 configuring text windows 43 connecting peaks 132 Converting to a calibrated axis 52 178 Converting to a mobility axis 52 171 Converting to mobility axis 172 Converting to molecular weight axis 51 163 convolution 100 copy 38 45 58 84 copying data 69 correlating data 99 Cross over peaks 121 curve samples 218 customising DAx 15 customising report windows 239 data lists customising 19
250. tion can also be used Press the Shift key while clicking the menu option to fixate or unfixate peaks for all data sets in the graphics window Chapter 8 3 3 Recalculating Peak Data Use this menu option to force peak parameters to be recalculated DAx normally recalculates all parameters as needed this menu option can be used to make doubly sure all values are up to date Press the Shift key while clicking the menu option to recalculate peaks for all data sets in the graphics window Chapter 8 3 4 Clearing Analysis Results The Clear analysis menu option removes peaks that have been found for the data set If a baseline has been created it will also be removed Press the Shift key while executing this menu option to clear the analysis results for an enslaved group of data sets 75 Chapter 8 3 5 Calculating RMS Noise Levels The Calc Noise option calculates the Root Mean Square noise level for the visible part of the curve for the data set The gt signal baseline numberofpoints Because the calculation is limited to the visible part of the data set if you have zoomed in on part of the graph only the visible part of the data will be included in the calculation A baseline needs to have been constructed for the data set Peaks are excluded from the calculation The result of the calculation is output to screen and to the log file calculation is RMSnoise NB The peak detection algorithm in DAx also uses noise levels If
251. tivate a peak in the data set graphics window chapter Chapter 7 2 move the mouse cursor over the peak top then click the right mouse button The peak popup dialog box appears Use the Active check box to switch between active and inactive states Peaks may also be activated and deactivated in the peak list window chapter Chapter 10 11 Highlight a range of peaks to de activate Click the right mouse button and select either Activate selected or Deactivate selected from the pop up menu Inactive peaks are not numbered in peak list windows Inactive peaks can still function as reference or normalisation peaks chapters Chapter 10 10 3 Chapter 10 10 4 127 Chapter 10 10 3 Normalisation Peaks One peak in each measurement can be marked as the normalisation peak To mark a peak as a normalisation peak in the data set graphics window chapter Chapter 7 2 move the mouse cursor over the peak top then click the right mouse button The peak popup dialog box appears Use the Normalisation Peak check box to mark the peak as the normalisation peak this will unmark any existing normalisation peak Repeat the procedure to unmark the peak Peaks may also be marked as a normalisation peak in the peak list window chapter Chapter 10 11 Click the right mouse button on the peak you want to make normalisation peak and select Set normalisation peak from the pop up menu Normalisation peaks are listed with norm following the peak number
252. to confirm aborting the measurement If any parameters have been changed but not saved you will be asked to confirm the loss of the Analysis Procedure and possibly the loss of individual sets of parameters such as GPC CE parameters you are asked this if separate parameter files are enabled cf chapter Chapter 3 1 1 56 Chapter 7 2 Data Set Graphics Window Edit menu The edit menu contains options that copy the contents of the window to the clipboard as well as options that undo or redo editing operations Chapter 7 2 1 Edit Select all This command selects all data sets in the graph window See the following section for details on the use of selected data sets Chapter 7 2 2 Edit Select This command invokes the data set selection dialog box 2 xi Ois C win winvolt harvard3 testBeter dal MAB HarvarD_1_oN_ ROx The Select all button can be used to HARVARD__1 0N_ ROK select all data sets that contain a given string of characters in their names Enter the string or choose one of the pre defined strings listed Check deselect to deselect all data sets that contain the string If the Alt key is pressed when the Select all button is clicked all previous selection will be removed before the new selection is applied HARVARD__1_ON_ FAM HARVARD__1_ON_ FAM HARVARD__1_ON_ IC HARVARD__1_ON_ VIC HARVARD__1_ON_ NED HARVARD __1_ON_ NED HARVARD__1_0T ROX HARVARD__1_O0T ROX H
253. ts A data set is a collection of signal values as a function of time or another parameter cf Chapter Chapter 3 3 6 DAx uses only data sets where points are measured at a constant frequency points are equidistant The first point may have a coordinate different from zero A data set contains additional data such as operator name measurement time data set name file name Furthermore data sets may contain derived data such as frequency domain data peak lists and splines DAx represents each data set with a data tag shaped like a button chapter Chapter 7 5 DAXSIM1 PRN i 24 Chapter 3 3 4 Default Data Sets The default data set or main data set is a data set that is used preferentially as the target for some operations such as when the mouse is double clicked in a graph window The default data set can be recognised because its data tag has a thicker edge and bold text Chapter 3 3 5 Measurement Data Types The measurement data type is used to increase ease of use by hiding options that are not relevant for certain types of measurements Measurement types are e Generic All options are available e GC Gas Chromatograms can have temperature profiles e HPLC Liquid Chromatograms can have solvent gradients e CE Electropherograms have parameters that are used to calculate mobilities e GPC SEC Options include flow rate and molecular weight calibrations The measurement data type is set in the Channe
254. ts Measuring frequency Ordinate Name Unit Name Measurement descriptions Extra fields An alternative view the Statistics List is meant to show component information Only component information stored in the files is presented The files are not re qualified when they are added to the data list 231 Chapter 19 1 Data List New list Data List Load list Data List Save list Data List Close list menu options Data Lists may be saved to disk using the Data List Save list option They may be loaded from disk using the Data List Load list option A new list may be started using the Data List New list option New lists can also automatically be generated Use the File Customise gt Data lists menu option to indicate when a new list should be started each day every week on a certain day when there are a certain number of items in the list or when a new sequence of measurements is started Data List Close list removes the list from memory Chapter 19 2 Data List Add data menu option Use this menu option to add files to the list The files will briefly be opened to scan their contents Chapter 19 3 Data List Assert data menu option This menu option checks if all the measurements in the list are still present The other properties in the data list are also checked and possibly updated Chapter 19 4 Data List Display list Data List Display statistics The Data List can be presented in two differe
255. ts you choose which information you want to export e the information being displayed e information about the current data tag and the ones it is related to ONOoSF 84 e information about all data in the graphic window e information about all data currently visible plotted in the graphic window The Print option in data set graphics windows has the option of including the Data Set Information with any graphics prints Refer to chapter Chapter 7 1 8 for details Chapter 8 11 The Data Set Action List Dialog Box This dialog box lists the actions that have been performed on the data set Some actions may have been performed more than once Refer to logging in the information dialog box for a complete lists of individual actions that have been performed Possible actions are listed below Enslaved Measurement the data set was measured on a channel that was enslaved chapter Chapter 3 3 9 to another channel Modified Data the data have been modified in some irreversible way Data moving or sizing is not irreversible but filtering is Some operations can only be performed on Modified Data This means that the original data will never be overwritten by an_ irreversible operation Baseline the data set is marked to be used as a baseline Baseline subtracted a baseline has been subtracted from the data set Has Normalisation Peak a normalisation peak has been selected for the data set Has Reference Peak a reference peak has been selec
256. u __ Chapter 9 1 Data Relationships amp Sorting Chapter 9 1 1 Sorting data sets Chapter 9 2 Data Plot select Chapter 9 3 Data Set colours Chapter 9 4 Data Reduce Chapter 9 5 Data Interpolate Chapter 9 6 Data Slice 4 86 89 89 91 91 91 92 93 94 Chapter 9 7 Data Remove trend 94 Chapter 9 8 Filtering Data Filter derivative 94 Chapter 9 9 Data Integrate 96 Chapter 9 10 Data Arithmetic 97 Chapter 9 11 Data Average 98 Chapter 9 12 Data Correlate 99 Chapter 9 13 Data Sort 99 Chapter 9 14 Data De Convolute data 100 Chapter 9 15 Data Generate data 101 Chapter 9 16 Data Overlay 102 Chapter 9 17 Data Stack data 105 Chapter 9 18 Data Best overlay 105 Chapter 10 Baselines and Peaks Data Set Graphics Window Peak Menu 107 Chapter 10 1 Peaks Construct baselines 107 Chapter 10 1 1 Using Gradients 110 Chapter 10 2 Peaks Subtract baselines 110 Chapter 10 3 Modifying Baselines 111 Chapter 10 4 Peaks Find peaks 112 Chapter 10 4 1 The Peak Find Wizard 113 Chapter 10 4 1 1 Selecting data sets 116 Chapter 10 4 2 Area Limit 116 Chapter 10 4 3 Automatic Thresholds 117 Chapter 10 4 4 Using Slope Limits 117 Chapter 10 4 5 Why use slope requires threshold 118 Chapter 10 4 6 Limiting Peak Search Intervals 119 Chapter 10 4 7 Peak Detection Events 120 Chapter 10 4 8 Automatic Shoulder Peak Recognition 120 Chapter 10 4 9 Normalising Peak Widths 120 Chapter 10 4 10 Removing Bleedthrough Cr
257. u cannot de convolute a curve with itself If New Curve or New Window which automatically means New Curve as well has been selected this restriction does not apply If the data set to be used to de convolute has a non zero time offset zero values will be inserted at the start It is recommended to use data sets with the same frequency as the object data 10 The exception to this are Savitzky Golay and Moving Average filters chapter Chapter 9 5 which are also convolutions but are performed in the time domain 100 De Convolution k 2 xj STAND 1 Da1 C Convolute Deconvolute 4 DE stanp 1 ba1 Ei or ane z O epp Gaussian Curve l2 Top ts foo DA C Triangle Stand Dev s 1 00 Asymmetric A C Block 3 fo C Exponential Gauss symmetry s fi 00 A A C Curve T New Window IW New Curve Cancel Help Filter Fraction 0 05 M Auto Check the Convolute item to convolute data Otherwise data will be deconvoluted The Curve Top item determines the time coordinate of the curve top for a theoretical curve DAx creates data points starting at the left side of the theoretical curve or zero seconds whichever is smaller The Curve Width item determines the width of the theoretical curve If the theoretical curve is asymmetric the curve width item determines the width of the left side of the curve and the Rwidth item determines the width of the right side of the curve
258. ulated but will be flagged in the peak list with an L or an H This can be used in quality control situations The dialog box is invoked from a quantitative calibration list or quantitative calibration graphics window using the Analysis Config quantification menu option Refer to chapters Chapter 18 9 2 and Chapter 18 9 3 for details The dialog box can also be invoked from the Identification Database list window by right clicking the mouse and can then be used to change quantification parameters for several components at once Refer to chapter Chapter 18 9 1 198 Quantitative Calibration 2 xi Quantification coordinate Normalised Peak Area Peak Area Migration Normalised Peak Area Migration m Calibration Curve Type C Multilinear C Cubic Spline Degree lal Polynomial ft z m Concentration Limits Low f5E 19 7 High 04 PA Derive from data Relative limits in Cancel Help 1 The calibration type determines the model for the calibration curve that is fitted through the calibration points Both the straight line and spline types calibrations will intersect all points used The polynomial calibration does not necessarily intersect the points Chapter 18 8 1 Dilutions DAx has the ability to handle dilutions You may indicate that a sample has been diluted a number of times DAx uses dilutions in the following way e when a measurement is added to a quantitativ
259. ursor tracks the data of the stacked band it is in and the data tags of the data sets in the band are depressed Chapter 7 6 Mouse Double Click Commands If the left mouse cursor is double clicked while the mouse cursor is in a data set graphics window several commands may be executed Most of these commands operate on a data set The data set affected is the one whose curve is nearest to the position where the mouse was double clicked with a bias towards the default data set The data tag for the default data set has a slightly thicker edge around it and has bold text e double click without pressing either the Shift or Ctrl keys to display popup menu The menu lets you e zoom out of the graph e create a bounding rectangle Drag the markers along the edges of the bounding rectangle to resize the data set or drag the entire rectangle to move the data set e add a peak split a peak set a relative origin or add a spline node or add a tack or label e double click while pressing the Ctrl key to add a peak at the time coordinate of the mouse cursor e double click while holding down the Shift key to set a relative origin When the mouse cursor is moved over the graph 6 This is true only if View Track data set is checked 63 coordinates will be referred to the relative origin Shift Double Click again to remove the walker To add peaks with specific begin and end coordinates press the Ctrl key and drag a rectangle betw
260. use wheel to zoom in on the position of the mouse cursor 37 Chapter 5 2 Graphics Window Menu Commands The menu of each graphics window contains a number of standard options Chapter 5 2 1 File Print This command prints the contents of the graphics window Chapter 5 2 2 Edit Copy graph This command copies the contents of the graph window to the clipboard as a so called metafile picture This picture may then be imported into a drawing program or word processor using the Edit Paste command in that program Chapter 5 2 3 Edit Create metafile This command creates an Aldus placeable metafile of the contents of the graphics window This metafile can be read into drawing programs Chapter 5 2 4 View Attributes This command invokes a dialog box that configures many aspects of the graph Among these are e the area of the window to be used for the actual axes picture e font to be used for axis markings Font size may be entered separately for screen graphics printed graphics and metafile graphics e colours e axis label texts e line width colour and dotting type for all curves The View Attributes dialog box lets you make the choice between coloured lines and typed i e dotted lines Typed lines can be particularly useful for use with black amp white printers 38 Axes Setup Axes Labeling Grid Curves Graph Background Thickness pts 4 Axes BackGround Screen Printer Clipboard Axes
261. used to limit the extent to which the calibration can be extrapolated beyond the scope of the calibration points e indicating if the detector measures weight or concentration abundance m Calibration Curve Type DAXSIM1 PRN oe DAXSIM1 PRN MV Use Logarithmic Mw s C Multilinear Cubic Spline Polynomial Degree ft A m Elution Volumes ml Low fo pA Hien fe GA MV Derive from data 2 I Do not alter point usage 4 Molecular Weight 5 Low fo Al High iE 20 WA Add to existing calibration 6 IV Derive from data ef se Derive from data Tl Detector measures weight 7 1 The GPC calibration type determines the model for the calibration curve that is fitted through the calibration points Both the straight line and spline types calibrations will intersect all points used The polynomial calibration does not necessarily intersect the points Use the GPC Calibration curve menu option to display the calibration as a graph Refer to GPC Windows chapter Chapter 15 6 for details on how to include or exclude individual points from the calibration Points can also be excluded by setting elution volume limits 2 The Elution Volume limits determine which points will be used in the calibration The volume limits are also used in both drawing Molecular Weight axis windows and calculating molecular weight peak data Any data points outside the volume limits will not be plotted or used in the calcu
262. ve files after analysis is checked 6 Check Move files after analysis to move files to a new location after they have been analysed Use of this option is recommended Current time 2 The wildcard should be specified as narrowly as possible In particular problems may occur if the analysis result files that DAx generates are included in the scanning operation files will be re analysed ad infinitum This will occur only if files are analysed files are not moved after analysis and if the wildcard allows da1 files to be read 34 Move files to specifies a location that files will be moved to after they have been handled DAx will create a file called DAx scan output in this location to prevent the location from being scanned for input files Check Only scan files since to limit scanning to newly appearing files This option is required if files are not moved to a new location after analysis The Current time button sets the time limit to the current time The time limit is inclusive files with the specified time will be included in the scan 35 Chapter 5 DAx Graphics Windows DAx contains several types of graphics windows There are Data Set Graphics windows chapter Chapter 6 DAx GPC has Volume axis windows graphics windows chapter Chapter 15 3 Molecular weight axis windows graphics windows chapter Chapter 15 3 and a GPC calibration window chapter Chapter 15 6 DAx CE has Mobilit
263. ve samples 218 View Darker 65 View Data set base calls 62 View Data set names 60 View Detector units curves 61 View Extra space 65 View First derivative curves 61 View Fit columns 44 View Frequency domain curves 61 View Gradient 157 View Group per lane 66 View Halftones 65 View Inches 239 244 View Inter curve derivatives 61 View Mark bins 62 66 View Mark peaks 66 View Minutes axes 61 View Only named peaks 140 View Plot all 59 View Plot data sets 59 View Plot groups 59 View Plot next 59 View Plot previous 59 View Plot select 59 View Reset scale 61 View Second derivative curves 61 View Seconds axes 61 View Select Peak Types 140 View Selected Lines 234 View Separate lanes 66 View Set colours 60 View Shrink amp fit columns 44 View Subtract baselines 65 View Tags setup 60 View Temperature programme 157 View Text size 44 View Track data set 61 percentages View Use trace colours 65 219 Window Close all 48 Window Minimise all 48 mesokurtic 137 message log 17 24 56 meta analysis 220 Meta analysis patterns 222 metafiles 38 59 migration 173 minutes 61 mobility 21 37 169 171 apparent mobility 171 effective mobility 171 mobility axes 21 171 Mobility axis 172 Modified Data 23 85 110 definition 26 modifying baselines 108 111 modifying
264. will be closed as soon as they are not needed anymore 8 DAx can create comparison sheets in which peaks from various analyses are grouped and compared By checking Add to Global Sheet the analysis results are automatically inserted into the global comparison sheet For details about comparison sheets refer to chapter Chapter 18 11 By checking Add to Binning Sheet analysis results are automatically added to the binning sheet cf Chapter Chapter 18 12 Check Exclude Calibrations to prevent size calibration traces from being added to the comparison and or binning sheets 9 Automatically overlay the data The settings last entered in the overlay dialog box chapter Chapter 9 16 will be used or you can click the blue underlined Overlay button to configure overlaying parameters Generally overlaying requires peaks to have been detected Certain forms of overlay use a target height or target area which can be marked as Derive from first data set encountered Normally this means the value is derived from the first data set encountered during the overlay operation In the case of automatic analysis the value will be derived from the first data set that is automatically overlaid in the window 10 Check Print Report to print a measurement report load a report definition using Report Load report 11 Check the Print Curves box to automatically print a measurement If a baseline has been constructed it will also be printed if peaks have been d
265. window The columns are called Fitted coordinate Fitted standard deviation Fitted height Fitted area and Fitted asymmetry factor Chapter 10 11 8 How Relative Concentrations are Calculated How relative concentrations are calculated depends on two things whether or not an Identification Database is available and whether or not a total concentration has been entered for a data set Use the Data Tag Sizing Sizing dialog menu option to enter a total concentration or enter it in the sequence dialog box e When no Identification Database and no total concentration is available relative concentrations are equal to peak area fractions and concentrations are not calculated e When no Identification Database is available but a total concentration is available relative concentrations are equal to peak area fractions and concentrations are calculated as relative concentration total concentration e When an ldentification Database is available but no total concentration is available concentrations are derived from the calibration in the Identification Database and relative concentrations are calculated as fractions of the sum of the concentrations for all components e When an ldentification Database is available and a total concentration is available concentrations are derived from the calibration in the Identification Database and relative concentrations are calculated as concentration divided by total concentration 139
266. ws in up to three lines For windows containing many tags the number of 70 lines of tags can be changed by placing the mouse cursor just below the data tags where it will change shape to an up down arrow Click and drag the mouse to change the amount of space allocated to data tags Chapter 8 1 Selecting Data Tags There are several ways to select data tags e click a data tag with the left mouse button to select it A check mark is displayed on the right side of the tag e click a data tag with the right mouse button to select an enslavement group refer to chapter Chapter 9 1 for details on data sets belonging together in enslavement groups e select multiple tags by pressing the Ctrl or Shift key when clicking additional tags Use the Ctrl key to select individual additional tags use the Shift key to select all tags between the last clicked and currently clicked tag As always selected tags will be displayed with a check mark on the right side To invoke the data tag menu or to start dragging a tag without changing the selection of tags click on the check mark of a selected data tag Chapter 8 2 Data Tag menu The data tag menu is displayed when the user clicks on a data tag with the left or right mouse button Some menu options can be applied only to the data tag that was clicked on whereas other options can be applied to multiple data tags at once There are several ways to indicate a menu option should be applied t
267. y axis windows chapter Chapter 16 5 and Time Corrected Signal Axis windows graphics windows chapter Chapter 16 6 DAx Calibrations has calibrated axis windows graphics windows chapter Chapter 17 4 and a calibration window chapter Chapter 17 8 2 All graphics windows share the way in which they can be zoomed as well as a number of menu commands These common attributes will be discussed in this chapter Chapter 5 1 Zooming in on graphics The mouse may be used to zoom in on parts of the graph To do this click the left mouse button at the top left coordinate of the part of the graph you want to zoom in on Now drag move the mouse while keeping the button depressed the mouse cursor to the lower right coordinate of the part you want to zoom in on then release the mouse button To zoom out or cancel a zoom operation click the right mouse button If the right mouse button is clicked while the graph was not zoomed in on you will be zoomed out to get a wider view of the graph Click the right mouse button again to go back to the normal view When the mouse cursor is moved across the graphics window the coordinate in the graph corresponding to the position of the mouse cursor will be displayed in the toolbar below the menu bar If your mouse has a scroll wheel rolling the scroll wheel will scroll the graph window left right or up down press the Shift key to change the scrolling direction Press the Ctrl key then roll the mo
268. y the user are displayed in red type peak type Peak types can be used to limit the peaks that are displayed to certain types use the 135 concentration View Select peak types menu option A peak can have multiple types concentration of the component Used in calibrations Concentrations that have been entered by the user as opposed to derived from the Identification Database are displayed in red A concentration unit can be entered using the File Customise gt Plotting Peaks menu option relative concentration relative concentration of the component See below signal to noise moment top time offset relative peak time k time marker time marker to find out how relative concentrations are calculated ratio of signal value at peak top and RMS noise level first moment of peak s calculated as 7 gt time value j gt value peak top time minus peak top time of the reference peak chapter Chapter 10 10 4 retention time of the component divided by the retention time of the reference peak m t t capacity factor is calculated as k with t the 0 peak top retention time of the component and to the peak top retention time of an inert component The peak top time of the reference peak chapter Chapter 10 10 4 is used as the reference time to If no reference peak has been defined the inert time is used The inert time is a sizing parameter cf chapters Chapter 7 7 and Chapter 8 5 re
269. ysis method has a resolution of 1 set the Area Limit on page 1 of the peak find wizard to 1 Any peak DAx finds with an area less than 1 of the total area of all peaks found will be removed Chapter 10 4 3 Automatic Thresholds You may use the Automatic Thresholds check box on page 2 of the peak find wizard to set a threshold value based on the data s noise level A factor should be entered and the threshold will equal the noise level multiplied by this factor It s advisable to also use slope requires threshold chapter Chapter 10 4 5 Chapter 10 4 4 Using Slope Limits If slope limits are being used to detect peaks the following conditions must be met for two consecutive points to start a peak we ll assume a positive peak is being detected negative peaks are the same just upside down e the signal value must exceed the noise level If the slope requires threshold option is in effect the signal value needs to ultimately reach at least the threshold value away from the baseline for a peak to be recognised based on slope e the derivative of the signal minus the derivative of the baseline i e the net derivative exceeds the slope limit e the derivative of the baseline is smaller than the slope limit or the derivative of the signal is at least twice the derivative of the baseline If the above conditions are met but the algorithm is already detecting a peak a new peak will be started if e a positive slope limit is exceede

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