SPR User Manual
Contents
1. Fig 2 Temperature Controller 1 Turn on the power strip 2 Turn onthe temperature controller 2 2 3 Turn the display select switch 2 1 to SET T and rotate the Adjust Knob 2 3 to choose the desired setpoint temperature between 14C and 45C 4 Turn the select switch 2 1 back to ACT T and press the Output button 2 4 1 Laser Key 2 TEC Button 3 Laser Button Fig 3 Laser Controller 5 Turn the key of the laser control to on 3 1 6 Turn onthe TEC 3 2 7 Turn on the Laser 3 3 8 Wait for both to stabilize usually 30mins at 6C and 20mA SPR User Manual Author Sidd Shenoy 2 Fig 4 Hamamatsu CCD Controller 9 Turn onthe Hamamatsu CCD Fig 4 SPR User Manual Author Sidd Shenoy Software Fig 5 SPR ARIA 1 Start the SPR Aria Software Fig 6 Multiline Control 2 Choose one or many in the multiline option depending on the situation Fig 6 Fig 7 Acquisition Control SPR UserManual Author Sidd Shenoy ss 4 3 Choose the AcqT ExpT and Gain default 0 5 0 02 0 Fig 7 Fig 8 Action Panel 4 Press Sample in the Action Panel Fig 8 to see a real time curve of the sample Fig 9 SPR Curve Fitting Panel 5 Choose Y limit in the Curve Fitting Panel Fig 9 to be the Y value below which you want to fit the SPR curve 6 Choose Width Position in the Curve Fitting Panel Fig 9 to be below the curve 7 Click File in the Ac2. the pattern on the SPR linear array is not sharp and SPR response is overlapping you can remove specified number of lines from the end of every line For example If Remove is set for 2 then 2 CCD lines from boundary of 2 closest lines total 4 lines will be removed from processing If we recall example with 600 um wide line the measuring width will be reduced to 4 x 24 um 96um or by 48 um from each side of 600 um line reducing it to 504 um Lines to show will specify data that will be displayed in the graph 1 and 2 windows This allows to see the process that goes at all lines or selected SPR User Manual Author Sidd Shenoy 10
3. SPR User Manual Sample Cell 1 Screw Gauge 3 Clamp Screw 4 Positioning 2 Temperature Screw Control bo 5 Cell Bolt 1 Pe N Fig 1 Sample Cell properly attached to the stage 1 Turn the screw gauge 1 1 to raise the stage off the glass slide 2 Loosen the screw on the right side of the stage 1 3 thereby separating the stage from the vertical clamp stand 3 To change cells loosen the two bolts 1 5 holding the cell to the stage Tighten the bolts with the desired cell in place 4 Clean the prism with isopropyl alcohol or ethanol 5 Completely cover the surface of the prism with 1 52 refractive index liquid 6 Gently place the new gold coated glass slide onto the prism with the gold side facing up ensuring it is held tightly in place by the built in grooves on the slide holder 7 Attach the stage onto the vertical stand and lower the stage so that the O ring of the cell presses on the slide and is held in place by the weight of the stage and the built in spring system The screw gauge 1 1 completely lifts off the base 8 Attach the temperature control cable of the cell to the controller 1 2 SPR User Manual Author Sidd Shenoy Experimental Apparatus amp Equipment MODEL LFI 3751 STATUS TEMP LIMIT T UMIT UMIT 3 Adjust Knob SHIF l 4 Output Button 2 Power a PP Watt Switch Temperature Controller 1 Display Select Switch avelength Electronics RP
4. es of reagents solutions that should be pumped through the SPR cell In time column Time type the time interval for how long this solution will interact with chip surface In comments you can type information about this particular solution protein concentration buffer etc After typing protocol for experiment Sequence of solutions with time and comments press Apply button and OK The window Fig 12 will be closed and in the window Fig 11 will be seen protocol in real time and it execution SPR User Manual Author Sidd Shenoy Flow Control Fig 12 Flow Control Modification Support Valve avle Control Fig 13 Support Valve Support valve is used to fill out empty tubing before experiment start and for washing or substituting reagents connected to particular input of flow control valve To open the support valve window double click on the Valve button of flow control window Fig 11 Use the same procedure to put information in the table as for flow control valve Support valve has 8 ports Numbers 8 7 and 6 are closed for output Ports from 1 to 5 are connected to the flow valve Always return to the closed port after using support valve SPR User Manual Author Sidd Shenoy Appendix Fitting Options Fitting options window is used to submit the fitting calculation and miscellaneous parameters that software uses to determine minima positions of SPR curve Fig 14 Fitting Contr
5. ol Fit calibration is the conversion factor between pixels and angle angle calibration pixel Smoothing passes dictates the number of smoothing passes which the smoothing function will use Offset of Win H specifies the value of offset for curves plotted in the graph windows in Multiline regime Polynomial Order specifies the order of the polynomial used to fit the data Y Limit is the upper boundary for data used in polynomial fitting Data with Y values greater than Y Limit are not used in the fitting function Point Number The fitting function will use every nth data point where n is the Point Number value Width Position specifies where to determine the half width calibration is the factor used to accommodate the S polarization All data values are multiplied by this factor Action Panel Fig 15 Action Panel Action panel is used for experiment preparation and execution Clicking File button the dialog box Fig 16 opens allowing set a file name for sensogram data the data plotted in the graph 1 will be stored in this file with supported information SPR User Manual Author Sidd Shenoy 8 Captured Data File TAR Fig 16 Captured Data File Clicking Data File button the dialog box Fig 17 opens allowing set file name for row data presented in the graph These data will be put in the file only when button Save Data is pressed any time during experiment This
6. orner of the Ul appears word Calibrate indicating that calibration mode is running Linear Array Options SPR User Manual Author Sidd Shenoy 9 Fig 18 Line Control Linear array options Fig 18 allow switching between single line button One and linear array or multiline button Many measurements By the default program works in single line mode To switch to multiline mode click on the button Many Fig 18 This will activate dimmed numbers in the dials in linear array options panel Fig 14 and Offset of Win H dial in fitting options panel Fig 14 The dials First of lines and Remove are used to set the number of CCD lines to be measured and processed Total number of CCD lines are 255 numbered from 0 to 254 Size of every pixel of CCD is 24x24um e Dial First specifies what will be the first line used for measurements For example if number will be set for 8 all data for lines between 0 and 8 will be omitted e Dial of lines specifies how many lines you like to measure For example if you specify 10 lines and First equal 5 you will have 255 5 250 CCD lines available for measurements Every line on the CCR chip will correspond to 25 lines on CCD array and total width of the line measured will be 24 um x 25 lines 600 um Thus you will have 10 lines with width 600 um e Dial Remove specifies how many lines should be skipped from processing If
7. procedure allows to store row data during experiment if necessary especially on critical points of experiment say during background detection when analyte is introduced in the cell during adsorption and washing in the same time avoiding to collect big files of unprocessed data Measured Data File Fig 17 Measured Data File Clicking Capture button the experiment will start executing in the sequence protocol that you put in the flow control window In the left top corner of the Ul appears word Capture indicating that experiment is running Clicking Stop button will stop the experiment running Clicking Wash button will start procedure that flushes and wash flow system all tubing valves and pump using the protocol that you put in the flow control window It similar to capture mode but runs without data measurement In this case in the left top corner of the Ul appears word Wash indicating that washing procedure is running Clicking Sample button allows detection of the SPR signal in the graph 2 without processing data Useful for preparation of the experiment In the left top corner of the Ul appears word Sample indicating sampling mode is running Clicking Calibrate button allows change polarization from p to s and acquires the calibration curve Calibration curve will be stored in special file and in the graph 2 will be displayed reflectivity curve R I I In the left top c
8. tion Panel Fig 8 and choose a file name for the SPR Minimum vs time plot to be saved 8 Click Data File in the Action Panel Fig 8 and choose a file name for the SPR Curve to be saved a suffix with timestamp is added everytime the curve is captured 9 Click Capture in the Action Panel Fig 8 to start taking SPR minimum vs time data At any point press the Save Data option to save the current SPR curve 10 When done press Stop in the Action Panel Fig 8 11 All data is stored in Desktop SPRARIA DAT folder SPR User Manual Author Sidd Shenoy 3 Software Flow Cell Pump control Fig 10 Flow Speed The pump is controlled through the center option panel The speed of the pump is the value of the dial in ml min The pump works in withdrawal mode when the sign of number is The pump will stop if the speed is set to 0 if less than 0 it pumps in opposite direction and when the Stop button is pressed stops Pump will NOT automatically start when Capture starts FLOW CONTROL Fig 11 Flow Control Display Flow control display window Fig 11 shows the order of the execution of the experiment After double click on flow control display window black space the window of flow control modification Fig 12 opens You can use flow control window to program your experiment In position Posit column of the table you can type the position number from 1 to 5 of flow valve Valve connects the volum
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