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Installation and Operation of the Cedex System

1. 14 Creating User Profiles and Delegating Access Rights u 44444 nn nennen nennen 15 Description of Access Rights 444444Hnnnennennnnnnnennnnnnnnnnnnnnnennnnnnnnennennnnnnn nn 16 Defining Target Directories for the Analysis Data 2 44snnnnnnnnnnennnnn nennen 16 Calibrating the Measurement System 244unsssnnnnnnennnnnnennnnnnennnnnnen nenn nennnnnnn nn 17 4 GENERAL ADMINISTRATOR FUNCTIONS 2 4444444420000000000000000n 21 4 1 Administration of User Profiles cccsescecsseecessseeeseeeseeeseeeseeeseeeeseeesesesneesesesnensesesenenss 21 Creating User Profiles m een 21 Deleting User Profiles u nee E AEEA 21 Modifying the Access Rights of a User Profile ussssrsnnesennnnnnnennnnnnnennnnnnn ernennen 21 Modifying a Username 2 cn een seen 22 Deleting a User Password 2 42 2 an eel All adenine 22 Automatic LOg Olt nn a ech neunten 22 4 2 Adaptation of the Counting Routine nrssussersonnnnnnnonnnnnnnnnnnnnnnnnnnnnnnnsnnnnnnnnnnnnnnnennnnnn 23 Description of the Individual Parameters 4 44us4444444444Hnnnnnnnnnnnnennennnnennnnnnn nenn 24 Modifying a Cell Type 1a een 26 Creating a New Cell Type nennen en een 26 Deleting a Cell Type een anna REEE 27 4 3 Using the CD BUMO nessas RER 28 5 PASSWORD MANAGEMENT scsssiccnsiscccestscadscensdescccnssstcosnirescsecsen
2. Figure 3 Cedex Workbench in the Administrator Mode Follow the instructions below to secure access and define the administrator password Enter the username Administrator Click on the traffic light symbol in the User Identification area Click on the Setup menu on the Cedex Workbench PF wn gt Click on Change my Password and the Change Password area will appear 14 Administrator Manual Cedex Version 1 6 1 Installation and Operation of the Cedex System amp 5 Define the administrator password Since no password exists leave the first field vacant Enter the new password in the second field and confirm by reentering in the third field again The password is case sensitive 6 Click on OK to confirm the password A message will appear to confirm the entry After the login the administrator or any user can change their password anytime by following the same procedure Creating User Profiles and Delegating Access Rights There are no user profiles installed on the Cedex analysis software as presettings Once the administrator password has been defined provides security for access to the software and related data The administrator can login now and create additional user profiles with specific access rights Follow the instructions below to create a user profile Click on the Users menu on the Cedex Workbench Click on Add and the New User area will appear Enter the name for the new user profile in the first fie
3. Figure 5 Cedex connections 20 User Manual Cedex Version 1 6 1 Installation and Operation ofthe Cedex System D Figure 6 PC connections Connecting the Cedex System to the Main Power Supply Connect the measurement system PC and monitor to the main power supply with the cords enclosed Observe the technical data User Manual Cedex Version 1 6 1 21 Cedex Preparing the Measurement System Cleaning the Reagent Containers To ensure optimal performance of the measurement system the reagent containers should be rinsed with distilled or de ionized water prior to first use as well as before every reagent fill up recommended once a week This prevents a possible contamination with dust or fluff particles Filling the Containers and Attaching the Sample Cup to the Sample Port Avoid running out of reagents during measurements because it leads to inaccurate measurement results and damages the syringe and 8 way valve Always check the volume level of the individual reagent containers prior to measurement Rinse all reagent containers well before refilling and always have a sample cup attached to the sample port Follow the instructions below 1 Place the reagent containers in their correct positions on the container tray refer to Figure 1 on page 13 2 Connect the capillaries to the screwing lid through the holes Ensure that the capillary tubes are not bent and almost reach the base of the containers 3 Use the l
4. If the Cedex system is not connected to an earthed power supply or the power supply is incorrectly polarized If the instructions for printer installation are not observed Software License Agreement Grant of License This is a license agreement between you the purchaser or licensee of the Cedex system and innovatis AG Innovatis AG grants the licensee the following license on condition that the licensee accepts all provisions of this agreement The licensee agrees to be bound by the terms of this agreement once commissioning or starting the Cedex system User Manual Cedex Version 1 6 1 7 Cedex The Cedex software is based on the following e application software to control the hardware components analysis module for processing registered images and data analysis e data base module to archive and process obtained data graphical user interface for the complete software all technical descriptions and instructions on the use of the software innovatis AG grants the licensee a non transferable non exclusive sublicense except as outlined below to use a copy of the software and documentation belonging to the Cedex system in compliance with the terms of this agreement Any property and commercial protection rights regarding the software product shall remain with innovatis AG The software is only licensed not sold No property rights regarding the software product shall be transferred to the licensee Th
5. 0 ccccccsecsecceeeeeeeeeeeeeeeeeeeeeeseseccaeaeeeeeeeteteneaees 76 Saving the Measurement Data cccceceeeseeecee cece ee eeeeeeeeee seer eeeeesseeeeeeseeeeesenseeaneeseeeseseeseeanes 78 MAINTENANCE OF THE MEASUREMENT SYSTEM eesseeeeeeeeeee 79 Cleaning and Maintenance Check List uuursenssnsnnnnnnnnnnnnnnnnannnnnnnnnnnnnnnannnnnnnnnnnnnnnn anne 79 Daily Cleaning and Maintenance uusrrssnnssnnnsnnnennnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnsnnnnnnnsnnnnn namen 80 Checking the Reagent Containers 0 s 4440040 snaenannenn 80 Emptying the Waste Container 44444444440Rnennnnnnnnnnnnnnnnnnnnnnnnnnnnnennnnnnnnennnnnnnennnnnnnn nn 80 Cleaning Routines Carried out by the Liquid Management 4ss424440eenssnnnnnnnn nennen 80 The Daily Events Area a Aaa ad ea Ae aed 81 The Cleaning Area cecncieinc distin wediie acti naaie intel l dads andl AEE 82 The Syringe Handling Area 20a we kl dled 83 Carrying out a Cleaning Routine 2444usrsnnnennnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnn 83 Interrupting Terminating a Cleaning Routine uunsessensnnnnnnennnnnnennnnnnennnnn nenn 84 User Manual Cedex Version 1 6 1 5 Verifying the Success of the Individual Cleaning Stages 4s nennen 84 9 3 Syringe Cleaning and Maintenance nessnnnseeennnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnsn nn 85 General Cleaning and Mai
6. 3 The ResultViewer is displayed by clicking on the right mouse button or clicking on Show IP Result in the Special menu User Manual Cedex Version 1 6 1 67 Cedex 4 Select the required options for the markings and settings The image can be enlarged with the magnifying glass or with the left mouse button see The Aggregate Histogram on page 44 Holding the right mouse button pressed down and moving it results in the enlarged image being moved within the frame 5 Click on Close to shut the window or use the X button NOTE IF SINGLE CELLS ON THE IP RESULT VIEWER ARE NOT MARKED AT ALL OR ARE MARKED AS DEAD WHEN THEY ARE VIABLE OR VICE VERSA THEN THIS IS AN EXCEPTION AND DOES NOT INFLUENCE THE STATISTICAL RELIABILITY OF THE ANALYZED RESULTS Excluding Cell Images from the Evaluation Process Including Excluded Images Follow the instructions below 1 Select the required cell image with the arrows in the Cell Image area or click on the overview button H to display all the single cell images as thumbnails 2 Select the mode of the single cell image from the drop down list below the image Valid image only refers to cell images included in the calculation process Excluded image only refers to cell images manually excluded from the calculation process and Invalid image only refers to cell images automatically excluded by the software from the calculation process 68 User Manual Cedex Version 1 6 1 The Analysis Results
7. 6 1 Description of the Analysis Results The Result Data area displays two values for each parameter forming two columns The Image column relates to the image currently being analyzed and displayed in the Cell Image area and the Overall column relates to the average value of all the valid images analyzed so far m Result Data Used images 20 Image Overall Viable Cell Density x105 13 88 15 63 Total Cell Density x107 18 59 21 19 Viability 6 74 6 FA Total Cell Count 71 1619 Average Compactness 1 091 1 068 Average Diameter um 172 10 40 Aggregate Rate 16 9 10 1 Standard Deviation 10 2 42 Aggregate Histogram Diameter Histogram Measurement Chart Compact Histogram Print results Cultivation Time Chart Liquid Management Figure 10 Analysis results The Viable Cell Density Parameter This parameter indicates the density of viable cells x10 5 Cells mL The Total Cell Density Parameter This parameter indicates the total cell density x10 5 Cells mL The Viability Parameter This parameter indicates the proportion of viable cells in to the total amount of cells viable and dead cells The Total Cell Count Parameter This parameter indicates the total number of counted cells User Manual Cedex Version 1 6 1 41 Cedex The Average Compactness Parameter This parameter indicates the average shape variation of viable cells from a sphere with the ideal sphere shape equaling gt
8. 1 2 3 4 5 Carry out an LM ShutDown Shut down the analysis software Shut down the operating system Switch off the individual devices at random Empty the waste container Carrying out an LM ShutDown All the capillary tubes in the liquid management system are rinsed with distilled or de ionized water during an LM ShutDownThe Trypan Blue is withdrawn from the 8 way valve syringe and the complete liquid management system replaced with air to prevent the capillary tubes and measurement chamber flowcell from getting damaged Follow the instructions below to carry out an LM ShutDown 3 Click on Liquid Management in the Result Data area on the Cedex Workbench The Liquid Management window will appear Click on the LM ShutDown button in the upper left hand corner of the window The time required for the LM ShutDown is then displayed Click on XJ to shut the window Refer to Maintenance of the Measurement System in The Daily Events Area on page 81 User Manual Cedex Version 1 6 1 39 40 User Manual Cedex Version 1 6 1 The Analysis Results O 6 The Analysis Results The analysis results obtained from a measurement are displayed in the Result Data area on the Cedex Workbench during and after the analysis process The obtained data can also be displayed as a graphic diagram by activating the Diameter Histogram Compact Histogram or Measurement Chart buttons Each parameter is explained in the next section
9. Empty the waste container often to prevent it from overflowing Always observe the safety regulations when handling Trypan Blue detergent ethanol or acids and bases e Waste contaminated with tissue cells e g sample cups pipette tips etc must be disposed of according to the recommended rules for handling bio hazard materials e Any leaks in the liquid management system must be repaired immediately Contact your distributor e Always ensure that there is a sample cups attached to the sample port Only use Cedex Detergent or a detergent with identical specifications for the cleaning process in the liquid management system Only use sample cups from suppliers specified in the Cedex Operating Instructions for the conveyance of cell samples to the sample port Other sample cups with other catalogue numbers can cause irreparable damage to the Cedex Always wear protective clothing when handling samples particularly gloves To prevent damage to the measurement chamber flowcell no particle larger than 80 um should enter the measurement system Do not use Microcarriers for this device e Ensure that all utensils required for sample preparation e g sample cups pipette tips etc are stored in a dust free environment to guarantee precise measurement results Ensure that the Trypan Blue solution is free of contamination otherwise filter prior to use Check that the sample cup is free of contamination before use
10. R Reactor ID 31 Reagent containers ne 80 Reagent fill up s nennen 22 Reprocess mode renser 72 Reprocessing uunnesnenneennnnnnennnnen 71 Result data nn nn 32 Result diagram s nennen 49 ResultViewer sn 66 Rights 7 S Safety instructions 9 Safety sheets unssnnenneennnnnneennnnn 93 Sample ID sense 34 Sample port 36 Sample properiies ne 32 S o 00 0 ee 28 Saving a View uunnnseenesnnnnnnennnnnnnnennnnn 52 Select Source 24neneneeennenenennn 58 Selected Measurements 59 Selection of Data Series 57 DOU Dina iaar aa as 74 Setup menu uuueeeseeenssnnnnennnnnnnnennnnnenn 31 Show IP Result sen 30 SNUTCOWM CORSEBERNESHERICHEESERERAEBEFELUDESUEBRFERFEER 27 Ssilic ne tube u rsaseenineine 22 SMEs a 47 SONG ea wed dee ea eee 86 Specials menu 4 nennen 30 Sphere reed 42 Spindel ee 89 Spindle cniinne nal 87 Standard Cells nen 75 Standard Clean nenn 82 Standard deviation en 42 Suitable location 20 S10 0 0 93 Switching Off essesnnneennnnnnennnn 39 Switching ON uuuessnseesssnnnnnneennnnnn nennen 27 Syringe 22242nnsnenenenneennennennn
11. Some data fields are offered in two formats e specification in 10 5 Cells mL e specification in 10 6 Cells mL Once one of these formats has been included in the Export Data area all data fields with the other format are automatically shielded Furthermore date and time formats in the Date Time Format area characters for the inversion of strings text fields for export in the CSV format in the String Terminator area and the decimal point symbol in the Decimal Point area can be selected 64 User Manual Cedex Version 1 6 1 The Analysis Results O The Start Export Button The export process is initiated with the Start Export button A window with progress bars appears in which the time progress of the export is displayed The export procedure can be interrupted using the Cancel button After successful export its termination is displayed OK closes the export window The Hide Button The Hide button fades out the Export window and the Cultivation Time Chart window is displayed again When the Export window is called again it appears in an unchanged form with the same contents Transposing an exported file into Excel Open the exported CSV file with a double click on the file name in Explorer If Excel is started earlier and used to open the file depending on the Excel version formats will possibly not be correctly transposed 6 4 Viewing and Selecting Cell Images Any single image that could lead to incorrect results durin
12. The Cedex program version 1 6 1 has been equipped with a detailed password management thereby providing better compatibility to CFR 21 Part 11 guidelines 5 1 Access Control at the Operating System Level We recommend that the system administrator ought be engaged as a Windows administrator Operating System Administrator This person should undertake the configuration of the settings that regulate access to the system at the operating system level The system administrator should login with a valid password as an administrator under Windows 2000 factory default Administrator and for security change this password The Security Update 1 application with which previous Cedex versions were equipped see Cedex User Manual has been replaced in Cedex version 1 6 1 by a new configuration In Windows NT the Administrator and Cedex were both available as users The Windows administrator could with the help of Security Update 1 select whether the Cedex users should have full access on the desktop or not In Windows 2000 this type of configuration has been replaced by installing an additional user CedexSA SA secure access The Windows administrator must now undertake setting whether the Cedex or the CedexSA user should be given access The Cedex user has access rights for the desktop He she can use all available applications e g the CD burner software The CedexSA has no access to the desktop and can therefore only use the Cedex
13. 1 6 1 35 Preparing the Sample and Attaching it to the Sample Port Prepare the sample prior to measuring according to the following instructions Transfer 1000 uL 300 uL for Cedex Small Volume of a well mixed cell suspension to a sample cup The measurement system tolerates volume variations of 10 Any greater difference will lead to inaccurate results and can cause air bubbles to enter the measurement chamber flowcell A cell density range between 5 x 10 1 x 10 Cells mL can be determined by the measurement system and any higher density would make a manual dilution necessary Cells with a diameter range between 8 um 40 um and cell clusters of up to 10 cells can be assessed accurately by the measurement system Remove the present sample cup from the sample port Attach the prepared sample cup containing the cell suspension to the sample port The sample cup is correctly attached to the sample port when it is in an upright position directly benea th it and the capillary tube almost reaches the base of the sample cup Figure 8 Attaching the sample cup to the sample port NOTE THE SAMPLE VOLUME CAN BE REDUCED TO 300 uL WITH THE OPTIONAL ACCESSORY SOFTWARE CEDEX SV CEDEX SMALL VOLUME 36 User Manual Cedex Version 1 6 1 To Start Measuring Working with the Analysis System 5 First ensure that the entire measurement system including the sample is ready for measurements by following the instruct
14. 1 lt Higher values indicate the extent of variation and therefore the overall state of the cell culture e g damage caused by shear force etc The Average Diameter Parameter This parameter indicates the average diameter of viable cells um The Aggregate Rate Parameter The Aggregate Rate specifies the percentage of cells Total Cell Count present in aggregates clusters The Aggregate Rate is given for each single Image displayed as well as for the complete measurement Overall while taking the excluded images into consideration The Standard Deviation Parameter This parameter only Overall column indicates the standard deviation of the single measured results of a sample results per image compared to the total cell density x10 Cells mL It therefore indicates the homogeneity of a sample and if measurement errors exist 6 2 Viewing Individual Analysis Results after Completion of the Measurement Process Once measurements have been completed the Result Data area displays the analyzed results of the last cell image Image and the average results of all the cell images Overall The Cedex software can also display single cell sample results from a previous set of measurements Select the appropriate sample data by clicking on the floppy disk symbol and following the instructions below Select the appropriate cell image using the arrows in the Cell Image area The chosen image is then displayed e g 17 20 is the 17 image of
15. 4 6 5 6 6 6 7 7 1 7 2 7 3 7 4 7 5 8 1 8 2 8 3 9 1 9 2 Display of the Measurement Series in the Diagram ecceeceeeeeeeeeeeeeeeeeeneeeeeeeaees 56 Measurement Series Offset cccecceceecnece cece ee eeeeeeaeeeeeeeeetegseaeaeeeeeeeeesesenaeeeeeeeeeeesieees 56 The Details Button sitnine ena eirinielddieenahlla 57 Grouping Measurement Series nennen landen landen learn 57 The Selection of Data Series Window 2 244444440onsnennnnnnnnnnnennennnnennnnnnnnennnnnnn nennen 57 The Select Source Area u nein 58 The Buttons in the Selection of Data Series Window 2444440s0nsnnnnennnennennenennnnen 58 The Selected Measurements Area 44 u0ossssnennnnnnnenennnnnnnnnnnnnnnennnnnnnennennnen nn 59 Exporting the Cultivation Time Chart Data 44444444444Hnnnennnnnnennnnnnnennnnnnnennnn nenn 59 TAS EXpOrt WindoW nn nee andre ne a nennen ieh 59 The Output File Name Area 444444000nnnnnnnnnnnnnnnnannnnnnnnnnnnnnnannnnnnnnnnnnnnnnannnnnnnnn 60 The Output Format Area 4uusssssssnnnnennnnennnnnnnnnnnnnnnnannnnnnnennnnnnnannnnnnn nenne EEEE EEEn 60 The Data Series Mixing Area uu s440unssnnnnnnnennnnnnnennnnnnnennnnnnennnnnnnepennnnnnennnnnnn nn 60 The Additional Information Area nssssssnnnsnnnenennnnnnnnnnnnnnnennnnennnnnennnen nn 61 The CSV Options Area 244
16. 5 Number of data sets of the fifth measurement View Name Name of the View used and its creator Export Template Name Name of the export template used and its creator The CSV Options Area Here options that specify the data transfer in the CSV format can be entered If the Use IBM OEM Charset checkbox is activated the IBM OEM code is used for the export file otherwise the Standard Windows fond is used If the Use LF instead of CR LF checkbox is activated line feeding in the export file is carried out only as Linefeed LF OAH otherwise Carriage Return and Linefeed CR LF ODH OAH are used The Export Template Area Once the options for the data export have been set they can be saved in an ExportTemplate This ExportTemplate can then be called any time for further data exports User Manual Cedex Version 1 6 1 61 The Buttons in the Export Template Area Information on name creator etc is listed for a loaded template in the Export Template area The Save Button Templates created can be stored using the Save button The Save an Export Template window opens in which the user can select and modify an existing Export Template name or enter a new name If the selected Export Template name already exists a warning appears with the question whether the existing file should really be overwritten Save stores the Export Template The window can be quit using Cancel without saving the Export Template Save an Export Template Name
17. 60 Area Output Format eenen 60 Area Select Source 58 Area Selected Measurements 59 Area VieW na rerae 52 Audit Trail button 222000000nn nn 32 Audit Trail file 69 Average compactness ennn 42 Average diameter eennn 42 B ASC Peer RIESEN A 86 BUNG a nee 22 Cc Calculator symbol 31 Capillary tube eect eeeeeeeeentteeeeeenaes 36 Capillary tubes eee eeeeeeeesteeeeeeenaes 87 Cedex Workbench ccccccssseceeeeeeeeeeaees 26 Cell ANG Gs aieia sccacdsacieoussivsiaacastases 47 Cell CIUSHEMS 0 ce ceeecccccececeeeeeeeeeeeeeeseeaees 36 Cell densities 2 222040000 nennen 35 Gellidensity u nee 36 Gell image ee 32 Gelliimages 40 2 ee 65 Chamber Clean 1 2u uu22nneeeeeeneeen 83 Chamber Clean 2 cccccccssessseeeeeeeeeeeees 83 Choose workarea uuunenenenenenenenenenenenenenen 30 Cleaning and maintenance 80 Cleaning area 440een nennen 82 Cleaning routine uee een 83 COMBING nun 47 Comment euesssssesseennsesnnnnnnennen nenn 35 55 Compactness histogram 47 Contact details ne4 ne 91 Contents of the delivery 19 Copyright 7 Counting routine eeessnneeeensnneennn nn 75 Cross sectional area 77 CSV Options een 61 Cultivation progress nen 49 Cultivation Time Char
18. Data Series window can be quit using the Cancel button If the data was modified a security question of whether these modifications should really be discarded appears A progress bar displays the adoption progress By using the Cancel button in the progress bar window the adoption can be interrupted whereby the Selection of Data Series window also fades out NOTE Only measurements already loaded are displayed in the Cultivation Time Chart window diagram The Selection of Data Series window must be called once again for loading the rest of the measurements The Details button is used for this Measurements not adopted are marked with red v in the Use column of the Selected Measurements area Data that was not at disposal not found for the adoption is additionally marked with an exclamation mark v When the mouse pointer is positioned on a measurement in the Selected Measurements list an info window that specifies the Workarea the measurement comes from appears when the right mouse button is pressed As soon as the OK button is actuated the missing measurements are reloaded and the process is regularly completed 58 User Manual Cedex Version 1 6 1 The Analysis Results O The Selected Measurements Area In the Selected Measurements area the single measurements that are adopted for each measurement series in the Cultivation Time Chart are listed In addition to the measurement series Reactor ID Sample ID Data Set and
19. ONLY USE THE EMERGENCY STOP BUTTON IN A CASE OF EMERGENCY BECAUSE THE CEDEX PROGRAM MAYBE HAVING TO BE COMPLETELY RESTARTED Verifying the Success of the Individual Cleaning Stages The actual image of the measurement chamber flowcell can be displayed on the Live image ofthe chamber area in the Liquid Management window to confirm the success of the cleaning stages If the Chamber Clean 2 is not done frequently a message prompting the user to start this cleaning routine appears on the desktop after approx 60 measurements L Cedex Workbench LO ix Eile Special Setup Users Help innovatis AG 07 03 03 16 19 02 7 User Identification f Process Parameters Cell Image Username Administrator Cell Type E Password FE Dilution 4 Shut down Logout Images r Processtype 7 Data Source Identification gt Status System needs nnowmr s Cleaning History ann 4 gt X Reactor ID Kopie_von_CHO 1 7 E Sample ID r Result Data DataSet Original data Used images Viable Cell Density x10 E Sample Properties Total Cell Density x105 4 Comment Viability Be Total Cell Count Average Compactness Average Diameter um Aggregate Rate Elan Standard Deviation x10 I Aggregate Histogram Diameter Histogram Creator Administrator Measurement Chart Compact Histogram Print results Cultivation Time Chart Liquid Management Figure
20. The Cedex analysis software offers various options to optimize measurement results by adjusting the Operator Standard Cells The Operator can be adjusted for abnormal cell culture images or for specific internal counting strategies Detailed information is found in the Cedex Administrator Manual The ResultViewer offers the following information and options viable number of viable cells dead number of dead cells e Viable Cells button to mark or unmark viable cells e Dead Cells button to mark or unmark dead cells e Mark Cells button viable cells are marked with a uniform yellow circle and dead cells are marked with a red cross Outline Cells button viable cells are marked with a yellow circle and dead cells with a red circle The circle diameter corresponds to the cell diameter Fill Cells button presently not available image enlargement brightness and contrast settings e navigating a window scroll bar 66 User Manual Cedex Version 1 6 1 E ResultYiewer EN 0004 00666 6 6 6 6 6 6 0 8 6 0 0 0 0 0 0 0 370 0 0 0 0000000 000000006 OO EO Dead Cells dead 47 Mark Cells Dutline Cells Eiligelle Operator Std Cells V3 0 Figure 30 Viewing a cell image using the IP ResultViewer Follow the instructions below to display the IP ResultViewer 1 Select the required cell image using the arrows in the Cell Image area 2 Move the cursor on the displayed image in the Cell Image area
21. Types Users System Detailed information is found in the Cedex Administrator Manual User Manual Cedex Version 1 6 1 23 24 User Manual Cedex Version 1 6 1 Working with the Analysis System 5 5 Working with the Analysis System The operating instructions and guidelines of the Cedex system are divided into the following sections Analysis system general instructions General instructions on working with the Cedex Workbench Starting the Cedex system e Description of the Cedex Workbench Description of access rights e Carrying out a measurement Interrupting a measurement Displaying and evaluating the analysis results Viewing and selecting cell images valid spot checks e Printing the analysis results Viewing and evaluating the data Audit Trail file e Editing existing analysis results data sets Switching off the Cedex system e General functions of the Cedex system e Administrator Functions 5 1 Analysis System General Instructions Switching on off the Cedex System Switch on the hardware components of the Cedex system always in the order below The individual devices have their own switch Proceed as follows 11 Switch on the measurement system 12 Switch on the monitor 13 Switch on the PC Shut down the software and operating system first before switching off the other devices at random NOTE IF YOU TRY TO LOGIN DIRECTLY AFTER THE LOGIN WINDOW APPEARS TWO WARNING MESSAGES MIG
22. W Total Cell Density 2g a F Viability Avg Diameter m e View Name KV_mix_020826 Creator Administrator Designed 26 08 2002 17 03 Modified 26 08 2002 17 08 1207 1 00 Load Save Delete ogo General ogo Print Export 0 401 Comment Hide 22 7 ae m Extras Doe 3 Fit lowest common value Time d os 1046 mf d Data Series _ KV 187 101 POO1 P017 16 10 2001 11 55 Olh Details 7 On New E 2 kvchoo1 POO P023 16 10 2001 11 43 Olh Details On New E 3 Kvchok 101 POOI P017 16 10 2001 11 06 Olh Details On New 4 KYhb5801 POO P031 16 10 2001 12 14 Olh Details On New 5 kvk56201 POOI P017 16 10 2001 12 05 Olh Details On _New Figure 18 The Cultivation Time Chart User Manual Cedex Version 1 6 1 49 Cedex Areas in the Cultivation Time Chart Window The Cultivation Time Chart Diagram In the Cultivation Time Chart window the grouped data see Grouping Measurement Serie Page 57 is displayed in a diagram When the mouse pointer is over a measurement point in the diagram detailed information of the corresponding measurement series is displayed in the View area see Figure 19 page 50 The view area is thereby faded If in addition the mouse pointer is pressed the graphic representation can be enlarged or scaled down again by moving the mouse In order to enlarge particular areas of the diagram do the following Place the m
23. a set of 20 and the corresponding data is shown in the Image column on the Result Data area 6 3 Graphical Display of the Analysis Results Besides displaying the evaluated analysis values in the Overall column the Cedex system can also display graphic diagrams of the results These diagrams can be displayed once measurements have been completed i e when all single sample images have been analyzed and evaluated by clicking on the Measurement Chart Compact Histogram or Diameter Histogram buttons in the Result Data area of the Cedex Workbench The results displayed in these diagrams are based on valid images taken of the sample Cell images that could lead to inaccurate measurement results due to lack of sharpness air bubbles major aggregates etc were already excluded from the calculations Images can be excluded automatically by the analysis software or manually by the user see Viewing and Selecting Cell Images on page 65 42 User Manual Cedex Version 1 6 1 The Analysis Results O Partial View of Diagrams Certain areas of the chart can be enlarged by following the instructions below 1 Place the mouse pointer on the top left hand corner of the area to be enlarged 2 Hold the left mouse button pressed down and move it to the lower right hand corner of the area to be enlarged 3 Release the mouse button and the selected area will fill the complete window The enlarged area or selected parts of it can now be moved by follo
24. area will appear Enter your present password in the first field Enter your new password in the second and third fields Click on OK to confirm the entry and a message will confirm the entry Click on OK to shut the window oa fF ON User Manual Cedex Version 1 6 1 73 Cedex 7 3 Viewing your own Access Rights Follow the instructions below 1 Click on the Setup menu on the Cedex Workbench 2 Click on Show my rights and the Show rights window will appear 3 Click on OK to shut the window 7 4 Data Entry Fields for the Data Source Identification Area Every set of sample data is clearly defined using a set of pre defined fields known as an ID Mask The Cedex system has a default ID Mask with three standard fields Reactor ID Sample ID and Data Set type If an additional ID Mask is required using different fields a customized ID Mask may be created by innovatis AG upon request Where alternative ID Masks are available the user can choose the appropriate mask using the ID Masks sub menu in the Setup menu 7 5 Specifying your own Default Measurement Parameters The logged in user with specific access rights to certain default measurement parameters cell type process type dilution etc can determine these during measurements These parameter settings are only valid for this user and if all the measurements taken are based on these default parameters then only the Data Source Identification area requires to be filled out
25. edited or translated without prior written consent of innovatis AG except as permitted under the terms of the copyright Disclaimer The information in this document is subject to change without prior notice and should not be construed as a commitment by innovatis AG The innovatis AG accepts no responsibility or liability for any errors or inaccuracies the document or for any damage occurring during delivery performance or use of this product Windows 2000 is a registered trademark of the Microsoft Corporation All other trademarks are trademarks or registered trademarks of the respective companies Cedex is a registered trademark of innovatis AG Warranty Standard warranty applies The warranty is void under the following conditions If any software other than the programs supplied with the Cedex system are installed on the computer excluding the printer driver and programs authorized by innovatis AG If damage or malfunction of the Cedex system is caused by abuse mishandling improper use or unauthorized repairs If unauthorized modifications are made to the Cedex system If the Cedex system is opened without authorization If damages occurred during transport are not reported immediately If damages are caused through equipment or software not supplied by innovatis AG If the Cedex system is not connected to an earthed power supply or the power supply is incorrectly polarized If the instructions for prin
26. file and reviewed later see Viewing and Evaluating the Data Audit Trail File on page 69 NOTE THE REPROCESSING PROCEDURE CREATES ADDITIONAL DATASETS ORIGINAL DATA IS NOT OVERWRITTEN 72 User Manual Cedex Version 1 6 1 General Functions of the Measurement System 7 General Functions of the Measurement System The Cedex Workbench offers users with access rights a row of functions besides the basic functions of analyzing modifying and reprocessing of samples and measurement results These functions allow easy and user friendly operations with the Cedex Workbench although some functions are only available to the administrator e g the calibration of the measurement system Detailed information is found in the Cedex Administrator Manual 7 1 Specifying Target Directories for the Analysis Data A target directory work area for the analysis results can only be selected if the user has specific access rights to this function Follow the instructions below to choose a target directory Click on the File menu on the Cedex Workbench Click on Choose Workarea and the Choose a Workarea list will appear Select the appropriate target directory work area ON gt Click on OK to confirm 7 2 Changing your own Password Users may change their own passwords depending on their access rights Follow the instructions below Click on the Setup menu on the Cedex Workbench Click on Change my Password and the Change password
27. management can be set in the Cedex ini file In order to be able to undertake the setting full desktop access must be permitted Open the support folder on the desktop with a double click Double click on Link to the Cedex ini The Cedex ini file opens The Cedex Administrator Using AdminAsUser in the System area it can be decided whether the Cedex administrator undertakes only administrative functions or in addition general functions too e g carrying out or viewing measurements etc AdminAsUser 0 Administrative functions only AdminAsUser 1 default Administrative functions and general functions The BlackList In order to select only passwords that fulfill a specific security standard there exists the so called BlackList This list contains passwords that are not allowed to be used Should a user select a password out of this list the message This password is too simple Choose another one is displayed Selection of whether passwords should be matched with the BlackList can be done in the Cedex ini file of the System area UseBlackList 0 default At the creation of a password it is not checked whether this password is in the BlackList UseBlackList 1 At the creation of a password it is checked whether this password is in the BlackList BlackListlgnoreCase 1 default Capital or small letters are not relevant BlackListlgnoreCase 0 Passwords that exactly match those in the BlackList
28. of cell suspensions 1 Prepare a cell suspension in a balanced salt solution preferably in HBSS Hanks Balanced Salt Solution 2 Transfer 0 5 mL of 0 4 Trypan Blue solution to a test tube Add 0 3 mL of HBSS and 0 2 mL of the of the cell suspension dilution factor 1 5 and mix thoroughly Allow to stand for 5 15 minutes IMPORTANT NOTICE PROLONGED EXPOSURE TO TRYPAN BLUE CAN CAUSE VIABLE CELLS TO STAIN AND LEAD TO FALSE RESULTS 3 Press a cover slip firmly on both chambers of the Neubauer hemocytometer and pipette a small volume of the prepared cell suspension mixture on the edge of the cover slip touching the edge gently with the pipette tip The chamber will fill up due to capillary action Follow the same procedure with the second chamber of the Neubauer hemocytometer Ensure that both chambers are not overfilled or underfilled 14 User Manual Cedex Version 1 6 1 Description of the Cedex System 4 Count the cells in the first chamber using a microscope with a magnifying factor of 100 objective lens x10 and ocular lens x10 Only cells in the four major corner squares and the major center square each containing 16 minor squares in the chamber are counted refer to Figure 2on page 16 Count the unstained viable and stained dead cells in the center minor squares and the minor squares on the top and left borders of the major square Do not count the cells in the minor squares of the right and lower borders refer to
29. plunger is then correctly positioned and the syringe module initialized 7 Ensure that there are no leaks in the liquid management system This can be done with one of the cleaning routines in the Liquid Management area Interrupt the cleaning procedure immediately with the EMERGENCY STOP button if there are any leaks in the syringe module and have them repaired 8 Screw the protective cover back on IMPORTANT NOTICE ONLY USE THE EMERGENCY STOP BUTTON IN A CASE OF EMERGENCY BECAUSE THE CEDEX PROGRAM MAYBE HAVING TO BE COMPLETELY RESTARTED 9 4 Maintenance of the Capillary Tubes The capillary tubes are automatically cleaned with every cleaning routine in the liquid management system However the external surfaces of the capillary tubes that reach into the sample cup and reagent containers should be cleaned often with a lint free cloth 9 5 Maintenance of the Syringe Guide and Spindle The syringe guide and spindle are mounted on either side of the syringe module inside the casing They should be lubricated half yearly or when necessary more often e g conspicuous noise from the syringe guide with the supplied lubricant to guarantee optimal performance of the syringe Follow the instructions below 1 Remove the syringe see Removing the Syringe on page 85 2 Switch off the Cedex system 3 Lubricate the top part ofthe syringe guide and spindle User Manual Cedex Version 1 6 1 87 oS GO mon mn a Ff Switch on the Ce
30. prior to measurements All other entries will be done automatically Follow the instructions below 1 Click on the Setup menu on the Cedex Workbench 2 Click on Defaults and the Defaults area will appear 3 Specify the appropriate default measurement parameters 4 Click on Save to confirm 74 User Manual Cedex Version 1 6 1 Maintenance of the Measuremment System D 8 Administrator Functions The following system functions are in principle only available to the system administrator e Administration of user profiles e Adaptation to the Operator Standard Cells counting routine Calibration of the measurement system 8 1 Administration of User Profiles The user usually has limited access rights All administration rights relating to user profiles i e creating deleting modifying and delegating rights should remain exclusively with the administrator Detailed instructions of Creating User Profiles Deleting User Profiles Modifying the Access Rights of a User Profile Modifying a Username Deleting a User Password can be found in the Cedex Administrator Manual 8 2 Adaptation of the Counting Routine Normally the Cell Type Standard Cells controls the image recognition making it unnecessary to modify the counting routine The Cell Type Standard Cell counting routine which is integrated into the Cedex system is set to suit all types of standard measurements However under certain circumstances it may be n
31. process Automatic Log out The Administrator can set a time limit after which a user is automatically logged out if the keyboard or the mouse button are not being used The log out also takes place during a current measurement The measurement however will thereby not be compromised In Cedex version 1 5 0 the default setting for Auto logout is 600 seconds In version 1 5 1 and 1 6 1 default is set at 0 seconds i e Auto logout is inactive Detailled information is found in the Cedex Administrator Manual 26 User Manual Cedex Version 1 6 1 Working with the Analysis System O To Shut Down the Cedex Workbench Follow the instructions below 1 Click on Shut down in the User Identification area Alternatively shut down by clicking on Xl or clicking on Shut down in the file menu 2 Click on Yes to shut down the program A restart of the analysis software is necessary for the next measurement session 5 3 Starting the Cedex System Always start the Cedex system by following the four steps in the order given below Switch on the measurement system Check the measurement system and have the samples ready Start the analysis software and login e P gt Perform a Prime if new measurements are to be carried out see Carrying out a Measurement on page 33 Switching on the Cedex System The hardware components of the Cedex system must all be switched on before the analysis software is started otherwise the software
32. sample cup containing the cell suspension The cell suspension and dye are then optimally mixed by controlled pump actions between the syringe and sample cup and then incubated with the exact same time for each measurement This prevents any possible decrease in viability caused by the toxicity of the Trypan Blue After the incubation period the stained cell suspension in the syringe is then transported in small portions to the calibrated measurement chamber flowcell through the capillary tubes and 8 way valve A set number of microscopic images of the cell sample is taken in the measurement chamber and transferred to the analysis software via an integrated interface in the measurement system Once the measurement procedure is completed a cleaning routine is automatically performed in the Liquid Management Only after the cleaning routine is it possible to begin with new measurements The Trypan Blue Exclusion Method The Trypan Blue exclusion method is based on the principle that viable cells repel dye and do not stain and dead or dying cells attract dye and do stain NOTE TRYPAN BLUE HAS A GREATER AFFINITY TO SERUM PROTEINS THAN TO CELLULAR PROTEINS IF THE MICROSCOPIC IMAGE HAS A DARK BACKGROUND DUE TO HIGH QUANTITIES OF STAINED SERUM PROTEINS THEN CENTRIFUGE THE CELL SAMPLE AND RE SUSPENDED IT IN A PROTEIN FREE MEDIUM OR PHYSIOLOGICAL SALINE SOLUTION PRIOR TO COUNTING Follow the instructions below on manual staining and evaluation
33. syringe use of special functions Follow the instructions below to customize access rights for a specific user Click on the Users menu on the Cedex Workbench Click on Rights Select the user profile to be modified Click on Edit rights and the Setup rights area will appear Define the access rights by clicking on the appropriate check boxes oa F WN gt Click on Save to store the changes The user is normally denied access rights to certain areas e g administration of user profiles and sensitive system functions calibration etc and therefore cannot access all the menus on the Workbench after login However access rights can be modified any time later by the administrator Defining Target Directories for the Analysis Data The Cedex system automatically creates a target directory work area when a measurement is carried out with the Reactor ID in the directory D Cedex images where all the analysis data are saved Specific target directories can be created with subdirectories of D Cedex images using Windows Explorer and be delegated to individual users 16 Administrator Manual Cedex Version 1 6 1 Installation and Operation of the Cedex System amp Follow the instructions below ak WN gt SO MND Click on the Users menu on the Cedex Workbench Click on Rights Select the user profile that is assigned to the target directories Click on Edit rights and the Setup rights area will appear Click on the A
34. the desired Workarea base directory in which measurement data are stored the Reactor ID and the preferred data type original or reprocessed measurement data are selected under Data Set It can also be determined later for each measurement in the Selected Measurements area whether original or reprocessed data should be used After selecting the Workarea and the Reactor ID the measurement list in the Sample ID area is displayed For a better view clicking on the bar with arrows can enlarge the Sample ID List area Entries selected from the Sample ID List can be adopted in the Selected Measurements list using the Add selected measurements to list button or Drag amp Drop This selection occurs individually for each measurement series 1 to 5 The Buttons in the Selection of Data Series Window Buttons 1 2 3 4 5 enable switching over between different measuring series The Add selected measurements to list button transfers the marked measurements of the Sample ID list to the Selected Measurements list The Review Measurement Data button loads a single marked measurement from the Selected Measurements list onto the Cedex Workbench There this measurement can be viewed in detail as usual The Remove Measurement s button removes marked entries from the Selected Measurements list The Ok button closes the Selection of Data Series window and the data is adopted in the Data series list of the Cultivation Time Chart The Selection of
35. video cable to the Video port of the PC Connect one end of the first serial cable to the Focus port of the measurement system Connect the other end of the first serial cable to the Focus port of the PC oa PF wn Connect one end of the second serial cable to the Syringe port of the measurement system N Connect the other end of the second serial cable to the Syringe port ofthe PC Connect the Hardware key USB Dongle to the USB port ofthe PC see Figure 2 NOTE CHECK THAT YOUR CONNECTIONS ARE CORRECT OTHERWISE AN ERROR MESSAGE WILL APPEAR CAN NOT ACCESS LIQUID MANAGEMENT OR SCOPE FUNCTION DISABLED 100 250V 2 3A 50 60Hz 5 a Made in Germany Focus Syringe Video Serial No Sicherung a age AE Fuse agen CE innovatis GmbH O T6 3 AH 250V CENI Typ Cedex Vor ffnen des Ger ten Notzstockor ziehen nur disconnect Figure 1 Cedex connections 12 Administrator Manual Cedex Version 1 6 1 Installation and Operation of the Cedex System amp Figure 2 PC connections Connecting the Cedex System to the Main Power Supply Connect the measurement system PC and monitor to the main power supply with the cords enclosed Observe the technical data 3 2 Preparation and Configuration of the Analysis Software If the hardware devices are switched on in their correct order the operating system of the PC will start automatically see chapter 5 1 Access Control at the Operating System Level pa
36. will be rejected The BlackList can be freely modified or expanded The following should be adhered to 1 Only one password for each row 2 No backspace before or after the password 3 Pay attention to capital and small letters if BlackListlgnoreCase has value 0 The BlackList is found in the C Cedex directory under Blacklist txt 30 Administrator Manual Cedex Version 1 6 1 Passwortmanagement 5 Specifying Password Characteristics Characteristics of passwords can be specified in the Cedex ini file of the System area The following table shows selectable characteristics of a password PasswordExpiration 0 default PasswordExpiration X Passwords never lose their validity Passwords lose their validity after X days positive whole number PasswordMaxRetry 0 default PasswordMaxRetry X Passwords can be falsely entered as often as possible Passwords can falsely be entered X times consecutively before this user is denied access Only the administrator can reverse denial of access by issuing a new password PasswordMinAlpha 0 default PasswordMinAlpha X Passwords do not require a minimum number of letters Passwords must have at least X letters PasswordMinLen 0 default PasswordMinLen X Passwords do not require a minimum length Passwords require a minimum length of at least X characters PasswordMinNum 0 defa
37. will not be properly initialized Switch on the individual devices in the order given below 1 Switch on the measurement system 2 Switch on the monitor 3 Switch on the computer These devices can also all be switched on at once with one button i e using a multiple contact plug First shut down the analysis software and operating system before switching off the other devices at random User Manual Cedex Version 1 6 1 27 Cedex Check the Measurement System and have Samples prepared Accurate measurements results rely on sufficiently filled reagent containers and the optimal performance of the measurement system Therefore check the measurement system with its reagent containers for possible wear and tear contamination and correct volume prior to each set of measurements as follows 1 Check the syringe module 2 5 mL syringe and 8 way valve for wear and tear and contamination Clean or renew if required see Maintenance of the Measurement System on page 79 2 Check the Trypan Blue solution for possible contamination and dispose of it if necessary Rinse the container well prior to refilling see Preparing the Measurement System on page 22 3 Regularly check if the reagent containers need to be refilled 4 Ensure that the capillary tubes are in their correct positions in the containers 5 Empty the waste container if necessary 6 A sample cup must always be attached to the sample port see Preparing the M
38. 0 6 Cells mL should be displayed on the y axis Correspondingly in the diagram the y axis calibration and division are automatically adjusted 10 5 10 6 Exponent switchover for the density The default value is 10 5 Buttons h and d determine whether the time axis should be displayed in hours h or days d Correspondingly in the diagram the x axis calibration and division are automatically adjusted h d Time axis switchover for hours h or days d The default value is d Buttons lin or log can be selected to determine whether the y axis should display linear lin or logarithmic log scaling for cell density lin log y axis switchover for linear or logarithmic scaling for the density data The default value is lin The Data Selection Area Which curves should be displayed in the diagram is determined in the Data Selection area The following parameters can be selected for this purpose Viable Cell Density number of living cells Cells mL e Dead Cell Density number of dead cells Cells mL e Total Cell Density total number of cells Cells mL Viability Avg Diameter average diameter of living cells um Avg Compactness average compactness of living cells without unit The selection of the parameters is done within the Checkbox m If only one parameter is selected its value Cells mL um numerical value of compactness without unit is displayed on t
39. 000 operating system consists of a better user management through which greater security and thereby better compatibility of CFR 21 part 11 is provided Security for access to Cedex was stepped up Access is allowed only for authorized persons The workstation will be automatically blocked after three consecutive entries of a false password Furthermore image recognition has been upgraded The new Std Cells 5 00 operator has replaced the old Std Cells 3 0 operator The identification performance was improved by the correction of the programmed code Dead cell recognition has been corrected in the Std Cells 5 00 operator This optimized dead cell recognition results in a change of viability towards slightly lower numbers Even cell suspension with low cell densities will only have a resulting difference in viability of maximum 3 The analyses functions of the Std Cells 5 00 operator are adequate to the Std Cells 3 0 operator In general the difference will not be recognized User Manual Cedex Version 1 6 1 11 12 User Manual Cedex Version 1 6 1 Description of the Cedex System 3 Description of the Cedex System The Cedex system consists of two functional units the measurement system which manages the sample handling and preparation and the installed analysis software supplied with the Cedex system which controls the measurement system including the sample processing and analysis of the images syrin
40. 1 lt I gt system Space on drive C 23420 MB 03 06 2003 14 27 51 lt I gt system Space on drive D 48182 MB 03 06 2003 14 27 52 lt I gt system System language DE 03 06 2003 14 31 00 lt S gt system User Administrator logged in 03 06 2003 14 31 26 lt D gt Administrator Loaded measurement Workarea D Cedex Images 03 06 2003 14 31 26 lt D gt Administrator Reactor ID TEST020916 03 06 2003 14 31 26 lt D gt Administrator Sample ID P008 03 06 2003 14 31 26 lt D gt Administrator Data Set Original data 03 06 2003 14 31 41 lt N gt Administrator Saved modified measurement 03 06 2003 14 32 03 lt S gt Administrator User logged out by inactivity timeout 03 06 2003 14 55 47 lt O gt system System shut down List of possible ID characters for the auditTrail file AS20 actions Calibration Data mode Errors Failed security relevant actions Hints Information Internal Liquid management Doing measurements Modifying a measurement System operations User actions with less importance Reprocessing Security relevant actions User management Emergency actions Comment X lt CHONDOOZEZETF TAMVOYS 32 Administrator Manual Cedex Version 1 6 1 Installing a Printer 6 Installing a Printer When a printer is installed on the Cedex system additional software printer driver is required for the PC The printer therefore should only be installed by authorized personnel i e the system administrator 6 1 I
41. 2003 innovatis AG All rights reserved Operating instructions and manuals are protected by copyright No part of this document may be reproduced edited or translated without prior written consent of innovatis AG except as permitted under the terms of the copyright Disclaimer The information in this document is subject to change without prior notice and should not be construed as a commitment by innovatis AG The innovatis AG accepts no responsibility or liability for any errors or inaccuracies the document or for any damage occurring during delivery performance or use of this product Windows 2000 is a registered trademark of the Microsoft Corporation All other trademarks are trademarks or registered trademarks of the respective companies Cedex is a registered trademark of innovatis AG Warranty Standard warranty applies The warranty is void under the following conditions If any software other than the programs supplied with the Cedex system are installed on the computer excluding the printer driver and programs authorized by innovatis AG If damage or malfunction of the Cedex system is caused by abuse mishandling improper use or unauthorized repairs If unauthorized modifications are made to the Cedex system If the Cedex system is opened without authorization If damages occurred during transport are not reported immediately If damages are caused through equipment or software not supplied by innovatis AG
42. 33 Prompt for cleaning Date Time 84 User Manual Cedex Version 1 6 1 Maintenance of the Measuremment System D 9 3 Syringe Cleaning and Maintenance The syringe is subject to wear and tear and must therefore be regularly serviced and certain parts renewed to maximize the life span of the syringe and 8 way valve and to guarantee optimal performance The measurement chamber flowcell tubes syringe and 8 way valve is cleaned during the Intensive Clean routine carried out by the Liquid Management see The Cleaning Area on page 82 The syringe should undergo an intensive cleaning procedure once a week if the Cedex system is frequently used see Cleaning the Syringe on page 86 General Cleaning and Maintenance Instructions To guarantee optimal performance and maximize the life span of the syringe following instructions and guidelines are recommended Should the Cedex system not be used for a longer period then remove the syringe and clean the plunger and syringe barrel thoroughly before replacing The Teflon head seal should be renewed regularly depending on the level of use and syringe maintenance Renew the entire syringe as soon as three gaps between the compact rings on the syringe plunger are clearly discolored taking the basic color of the Teflon head seal into account Removing the Syringe Follow the instructions below 1 Click on Liquid Management on the Cedex Workbench 2 Cli
43. 40ssssnsnnsnennnnnnnnennnnnnnnnnnnnnnnannnnnnnnnnnnnnnannnnrnnnnnennnnennnnnrnn nn 61 The Export Template Area 244444440annnnnnnnnnnnnnnnnennnnnnnnnnnnnnnannnnnnn nenne nnnnen nn 61 The Buttons in the Export Template Area 4snsnnsnennnennnnnnennnnennnnnennnennnnn 62 Available Export Templates for Excel 24444444044nnnnnnnnnnennnennnnnnnnennennennnnen 63 The Export Structure Area unsssssennnnnnsnennnnsnnnnnnnnnnennennnnnnnnnnnnnennnnrnnennennnnennennrnnnnn 64 The Start Export Button uussssssennnnnnnnennnnnnennnnnnnannnnnnnennnnnnnnnennnnrrnnnnennnennnnnn 65 The Hide Button 42442444444400nsnnnnennnnnnnnannnnnnnnnnnnnnnannnnnnnrnnennennnnennnnrrnnnnennnennnnen 65 Viewing and Selecting Cell Images uuusssrrssnnnsnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnsnnnnnnnsnnnnn namen 65 Viewing Individual Cell Images 4444nnnnnennnnnnnennnnnnnennnnnnnnennnnnnnnnnnnnnnnnnnnnnnnn nn 65 Enlarging a Cell Image 4444444snnnnnnennnnnnennnnnnnnnnnnnnennnnnnennnnnnnnnnnnnnnnnennnnnnennnnnnnn ran 66 Viewing a Cell Image Using the IP ResultViewer uunssneenssnnnnnnennnnennnnnnnnnnnnnnnnnn nn 66 Excluding Cell Images from the Evaluation Process Including Excluded Images 68 Printing the Analysis Results 222222000044404n0n0anannnnnnnnnnnnnanannnnnnnnnnnnnnnnannnnnnnnnnn
44. 60 200 220 240 Figure 4 Calibration window Calibration starts automatically and follows in several stages At first the calibrating solution Cedex Calibration Beads is aspired into the measurement chamber flowcell Preparing test liquid In the meantime the microscope stage moves to its lowest position and then to its highest position and the brightness is set Checking brightness and saved in the brightness histogram The complete path of the microscope stage is then scanned Scanning focus path and the obtained image information is analyzed The focus position is set where the image is most vivid and the brightness optimal Checking brightness The area around the located focus position is then examined closely Finding optimal focus and scanned several times to obtain the average values The brightness is finally set again to the determined focus point The calibration process ends with a cleaning routine Cleaning system Every stage indicated by a flashing button and its estimated duration indicated by the blue progress bar is displayed on the Calibration Progress area The brightness histogram indicates the distribution of brightness within the measurement chamber flowcell The blue cube in the focus histogram indicates the optimal focus position The respective value in the yellow field above the cube indicates the position determined for the focus The yellow field below the yellow line indicates the actual
45. 9 Exporting the Results gt 54 Export Templates for Excel 63 User Manual Cedex Version 1 6 1 95 Cedex F Fast Clean 82 File ment nee 30 Floppy disk symbol 31 G Gener l uneeeeesesesesesnnenannenenenenennnnnnn 54 General functions nen 73 Graphical display nn 42 Grouping Measurement Series 57 H Hardware components gt 20 He sans 86 Help MOM neinni 31 Aide DUMON oenina 55 Hide Button export ccccececeeeeeeeees 65 I ID Masks 0u un aan 74 Incubation ssanie iais 14 Individual cell image 65 Installation 22222202 nennen 19 Intensive Clean eenennnnn 82 Interrupting serura ee 38 IP ResultViewer ssesenennnennnnnnnnnnn 66 ISOPFOBYle enden 23 L Language uersnneeensnneenennnnennnnnnnnenn nn 31 License agreement nenn 7 LM eee a cane 51 Liquid Management 32 80 List of Measurement Series 55 Live image of the chamber 84 LM ShutDown 0cccccccceseseeeeeeeeeeeeeees 39 Loader program mennnneennnneneennn 29 Loading a View 53 Lg aa 51 Login ae re 26 LOOV ae 26 M Main power supply 21 Maintenance 28 34 38 79 Maintenance check list 79 Meas
46. Cell Type A new Cell Type can be created on the basis of an existing Cell Type The existing Cell Type is saved under a new name which is then modified Follow the instructions below 1 2 3 4 26 Click on the Setup menu on the Cedex Workbench Click on Cell Types Select New and the Create a new cell type window will appear Select the algorithm that the new Cell Type is to be based on from the Choose algorithm based on field Only the Standard Cells algorithm is available at present Select the new Cell Type based on the existing Cell Type from the Choose the cell type based on field Enter the new Cell Type in the Name of the new cell type field Click on Create to establish the new Cell Type Adjust the individual parameters by clicking on the E and buttons to increase or decrease their values Click on Revert just below the individual parameters to delete any changes Click on Save to confirm the adjustments Click on Cancel to shut the window without saving any changes Administrator Manual Cedex Version 1 6 1 General Administrator Functions O Deleting a Cell Type Follow the instructions below Click on the Setup menu on the Cedex Workbench Click on Cell Types Select Delete and the Choose cell type to delete window will appear Select the Cell Type to be deleted Click on Delete and you will be requested to confirm the entry oa Bon gt Click on Yes to confirm the entry NOTE ALL PARAMET
47. E ee ee Be a Cells of total Cell Court is Figure 14 The Aggregate Histogram total view sum of living and dead cells here percentage The Measurement Chart Click on the Measurement Chart button to display a Column diagram showing the adjacent combination of viable cells green column and dead cells red column as a single image 4 Statistics Screen I viable avg Measurement Statistics D dead avg E viable H dead Figure 15 Measurement Chart Any cell image that is declared invalid and excluded from the calculations appears as an empty space on the chart allowing easy identification of unusual samples These unusual samples can then be displayed and the respective cell image s evaluated see Viewing and Selecting Cell Images on page 65 46 User Manual Cedex Version 1 6 1 The Analysis Results O Two horizontal lines in the diagram show the average number of all analyzed viable cells The blue line represents the number of the living and the yellow line the dead cells Combine two cell images by clicking on the Combine button below the diagram The combination of two images corresponds to the results of a Neubauer hemocytometer Return to the standard display by clicking on the Single button The Compactness Histogram Chart Click on the Compactness Histogram button to display a diagram that compares viable cell shapes to that of a sphere This diagram shows results that do not relate
48. ECHNICIAN IS PERMITTED TO CARRY OUT MAINTENANCE WHEN THE CEDEX DEVICE IS REQUIRED TO BE OPENED User Manual Cedex Version 1 6 1 89 90 User Manual Cedex Version 1 6 1 Appendix amp Appendix Contact Details Information and technical support innovatis AG Meisenstr 96 33607 Bielefeld Germany Phone 49 0 521 2997 295 Fax 49 0 521 2997 285 email sales innovatis com support innovatis com validation innovatis com qa innovatis com internet www innovatis com User Manual Cedex Version 1 6 1 91 Cedex Technical Specifications Cedex Measurement System Method of measurement Digital image and pattern recognition Viable dead cell differentiation Trypan Blue exclusion method 5 x 10 1x 107 cells per mL Material and Diameter of the capillaries Teflon 765 um 10 C 35 C 60 F 95 F Operating humidity 20 80 relative humidity non condensing Dimensions Height 660 mm 26 0 inches Width 300 mm 12 0 inches Depth 600 mm 23 6 inches Weight 27 0 kg 60 0 Ibs Energy requirements 100 250 VAC 50 60 Hz Energy consumption Automatic Sampler AS20 Number of samples Average measurement period for 20 samples Dimensions Height 170 mm 6 7 inches Width 175 mm 6 9 inches Depth 160 mm 6 3 inches Weight 2 6 kg 5 7 Ibs nergy requirements 12 VDC nergy consumption Power supply Input 100 240 VAC 47 62 Hz Output 12 VDC 2 0 A Cedex Computer Newest PC ge
49. EPEPEEELFELBEETEFPFEREEFFFFPEREEPEPPEELE 5 WADA occ cece acces ce ae a se eine ence a HE a a a a ne 5 Software License Agreement uunssnsesnnneennennnnennnnnnnnnnnnnnnnnnnnnnnennnnnnnnnnnsnnnnnnrnsnnnennnsnnnnnrnsn nennen 5 General Provisions 20000000 a ee na 7 FCC Notification a cecracucens aocnc nace ncmaccunnan aeiecnanenenscemaenesecnnexceatenenntesnsbensennaeetanesentions 7 Safety Instr ctions 2522 22 202 21a E ER AEE 7 2 INTRODUC MON 22 une ee 9 3 INSTALLATION AND OPERATION OF THE CEDEX SYSTEM 11 3 1 Installing the Cedex System nnnnnnnnnnnnnennnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnsnnnnnnnsnnnnnnnnnnnnnn nennen 11 Checking the Contents of the Delivery 20us424440nnnnnnnnnnennnnnnnnnnnnnnnnnnnnnennnnn nenn 11 Choosing a Suitable Location for the Cedex System 40snnersnennnnennnnnennnnenn nn 12 Connecting Hardware Components and Hardware Key Dongle u nenne 12 Connecting the Cedex System to the Main Power Supply uuunsssssnsnesnnnnnnennnnnnnennnnn 13 3 2 Preparation and Configuration of the Analysis Software uuunsssnnennnnnnnnnnnnnnnnnnnnnnnnnnnn 13 Starting the Analysis Software uuessnnnnennnnnnnnnnnnnnnnnnennnnnnnennnnnnnnnennnnnnnnnnnnnn nn 13 Configuring the Analysis Software uesnnnsennnnnnnnnnnnnnnnnnennnnnnnennnnnnnnnnnnnnnne nn nn ern 13 Securing Access Rights and Defining the Administrator Password
50. ER OF THE MEASUREMENTS ARE STORED IN THE AUDIT TRAIL FILE SO DELETED CELL TYPES HAVE NO INFLUENCE TO FURTHER MEASUREMENTS ATTENTION DO NOT DELETE THE CELL TYPE STANDARD CELLS Should the Cell Type Standard Cells be accidentally deleted then it can be recovered by using the option based on lt none gt Administrator Manual Cedex Version 1 6 1 27 Cedex 4 3 Using the CD Burner If you intend to download your data onto CD with the help of the CD burner then follow these procedures Login to Windows as Administrator to access the Windows desktop e Start the burner software by double clicking on the Easy CD Creator icon on the desktop or use the Start button and then program Roxio Easy CD Creator The Select a Project window will appear with various types of burning options e Select the data you wish to copy from the directory in the upper split window The Cedex data for particular measurements are factory set and stored in the directory D Cedex images reactor ID sample ID Mark the files accordingly and drag them into the window below as in Windows explorer Start the burner process When this process is completed a message is shown to confirm its success When closing Easy CD Creator you will be asked to save the data list Answer with NO as long as you have not yet set up a specific data list which you might need later 28 Administrator Manual Cedex Version 1 6 1 Passwortmanagement 5 5 Password Management
51. Figure 3 on page 16 5 Record the two values for the first chamber 6 Repeat the same procedure for the second chamber of the Neubauer hemocytometer 7 Record the two values for the second chamber Repeat the entire procedure if more than 10 of the cells are in clusters Ensure an even distribution of cells by gently pumping the cell sample in the pipette a few times or carefully mixing it Repeat the entire procedure should less than 200 or more than 500 cells equivalent to a cell density of 20 50 cells in a square be counted in the 10 major squares of both chambers with the appropriate dilution 8 Repeat the whole procedure with a second sample to verify the results of the first sample 9 Calculate the results as follows If the cover slip is correctly placed on the hemocytometer then each major square of a chamber will have a volume of 0 1 mm i e 10 cm The cell concentration per mL total cell count is calculated as follows Total cell count cells per mL average count viable dead cells per square x dilution factor x 10 cell count for 10 squares Example Average count per square 45 cells 45 x 5 x 10 2 25 x 10 cells per mL Total amount of cells in the sample Cells mL x original volume from which the sample has been taken Example 2 25 x 10 cells per mL x 10 mL original volume 2 25 x 10 cells Cell viability viable cells total cell count dead viable x 100 Exa
52. HT FOLLOW BOTH SHOULD BE CONFIRMED WITH OK CONTINUE BY REPEATING THE LOGIN PROCEDURE YOU CAN AVOID THE WARNING MESSAGES BY AVOIDING SHORT TIME DELAY BETWEEN SWITCHING ON THE PC AND LOGGING IN THE WARNING MESSAGES APPEAR BECAUSE THE DIFFERENT HARDWARE DRIVERS NEED TO BE LOADED PRIMARY TO THE LOG IN User Manual Cedex Version 1 6 1 25 Cedex Starting the Analysis Software Follow the instructions below 1 Start the operating system by pressing the keys lt Ctrl gt lt Alt gt lt Delete gt to login 2 Enter Cedex as username and password and confirm by pressing lt Enter gt The Cedex analysis software will then start automatically The password is case sensitive 5 2 General instructions on working with the Cedex Workbench The Login If the Cedex analysis software has already been started then proceed as follows 1 Enter the username in the Username field 2 Enter the password in the Password field if applicable 3 Click on the traffic light symbol or press lt Return gt The Logout Follow the instructions below 1 Click on Logout in the User Identification area 2 Click on Yes to confirm Alternatively exit with Logout in the File menu This option allows the present user to logout without shutting down the software Logout is also possible during a measurement without influencing it Only the original user or administrator can login again during this process A message indicates that measurements are in
53. O 6 5 Printing the Analysis Results The analysis results obtained by the Cedex system can be printed by clicking on Print results in the Result Data area The printout consists of at least an overview page with all the manufacturer details including all sample data and analysis parameters entered by the user as well as a tabular and diagram of the overall results A more detailed printout is also possible using the Cedex Workbench with the following printing options no details please Limited printout details on the overall analysis results of the measured sample including a tabular displayed in the Result Data area as well as a graphic diagram of the Measurement Chart Compact Histogram and Diameter Histogram 2 images per row up to 5 images per row Limited printout details as above in addition to a printout of all the single cell images including their six analysis parameters The amount of cell images per row determines the number of pages to be printed e print colored select to optimize output for color printing otherwise the output is optimized for a gray scale mono printing Follow the instructions below to print the analysis results 1 Click on Print results in the Result Data area The limited printout details are automatically displayed when the Print window appears 2 Select the appropriate printing option s Click on Print Options to select and set the printer 4 Click on Print to start printing Click
54. Only clean sample cups can guarantee an optimal function of the Cedex If not in use keep the Trypan Blue solution in a refrigerator e Secure your measurement data frequently recommended approx every after 250 measurements NOTES EVERY CEDEX SYSTEM IS THOROUGHLY TESTED IN THE END CONTROL ALL RESULTS OF THIS CONTROL TESTING IS SAVED IN THE SYSTEM FOR POSSIBLE USE LATER THE SOFTWARE AND HARDWARE VERSION OF YOUR CEDEX ANALYSIS SYSTEM CAN DIFFER FROM THE INFORMATION DESCRIBED HERE BUT SHOULD NOT MAKE A DIFFERENCE TO THE OPERATIONAL FUNCTIONS IN GENERAL IMPORTANT NOTICE THE MANUFACTURER IS NOT RESPONSIBLE OR LIABLE FOR ANY PERSONAL OR OTHER DAMAGE RESULTING FROM IMPROPER USE OR FAILURE TO COMPLY WITH THE REGULATIONS IN THE OPERATING INSTRUCTIONS 8 Administrator Manual Cedex Version 1 6 1 Introduction amp 2 Introduction The necessary settings of the prepared and fully mounted Cedex system should be done by the system administrator We recommend to constitute the Windows administrator Operating system operator as the system administrator He she should establish the rights and access to the system at the operating system level The system administrator should log in to Windows 2000 as Administrator with the valid password factory setting Administrator and change this password Apart from the Cedex user which is only allowed to perform measurements the Cedex administrator should be established with main respon
55. Start Date Time when the measurement was recorded are listed The measurement list is automatically sorted according to the time of recording of each measurement The serial number of a measurement within a measurement series is given in the No column This number is given under Point in the Detailed Information area for better relation of the measurement points see Figure 19 page 50 The marked status Y of the Checkbox in the Use column causes the corresponding single measurement to be displayed in the Cultivation Time Chart Should it not be desirable to display a particular single measurement it can be deactivated by means of a mouse click In the Data Set column there is a possibility of also selecting reprocessed or modified data in addition to the original data of a measurement This occurs in the selection of the Drop Down list If the columns for Reactor ID Sample ID or Data Set are too narrow to completely display the entries the width can be adjusted Do as follows 1 Position the mouse pointer on the corresponding column dash in the column heading 2 Hold the left mouse button pressed down and move the mouse pointer with the column dash to the required width Exporting the Cultivation Time Chart Data The Export Window The Export button in the Cultivation Time Chart window opens the Export window In this window one has the possibility of exporting measurement series data currently displayed in the Cultivat
56. Z Roches Roc he CEDEX and INNOVATIS are trademarks of Roche Other brands or products names are trademarks of their respective holders Cedex Analyzer Operator s Guide Software Version 1 6 1 October 2003 For life science research only Not for use in diagnostic procedures User Manual Cedex Version 1 6 1 e QUALITY THAT COUNTS Version Cedex 1 6 1 10 29 03 MM DD YY 2 User Manual Cedex Version 1 6 1 Contents GONTENTS EHER 3 1 RICHT San 7 GOP Yi NG VOPPREUREPEESEEN cose asec REISE RE aE a a N DESSERT aa a a Aaaa ENEA ER PEENNBR 7 Disclaimer ee De En En rk EEE 7 Wa ta thy PPRBPEPPEREEFPPPEREREFFPEFEEEFFPPRREEFFEEFRFEFFPERRFELFPPREFEFFPEREEFEFFPERRFEEFPPFEFEFFPPERFEEFEEFEFEFFFPRREEFFPPFERFEFPFEFEFFPPEFEEFEEFEFR 7 Software LICENSE Agreement sisien ea a a nE A A AEA AERAR Aaa ASEA AAAA AAE AAEE E REEERE RER 7 General Mod eA l 1Le n E ZERPEPPERRREFPEFFEBFEFPEEREETEFEFBREEEPPERFRFEFEPFABLEPEEFEFEFEFERLEFEPEBEPELEFFFEEBFEFEEREFELFFLERFEFERFEFPERLPRREFFERER 9 FCC N tification 4 4uRBe ee ea Ba a ar EEE 9 Safety Instructions sss ee 9 2 INTRODUC TION nee 11 3 DESCRIPTION OF THE CEDEX SYSTEM 13 3 1 The Measurement System iiss cicciccccciecteeecics cgedctecwedecsceccesuheeteedescesuzeectcecceauccerhencecesdenctueredececess 13 3 2 The Measurement Procedure 444444404000nnnnnnnnnnnanannnnnnnnnnnnnanannnnnnnnnnnnnnnnannnnnnnnn nenn 14 The Trypan Blue Exclusion Method
57. a Print Cultivation Time Chart I print colored I PrintOptions Preview Cancel Figure 23 The Print Cultivation Time Chart window If the print colored Checkbox is clicked the colors of the diagram are adopted for colored printing If the Checkbox is not clicked the data is displayed in black and white and is suitable for a black and white printer Printer settings are selected using the PrintOptions button The Preview button displays a printing preview The Print button initiates the printing of the diagram The Cancel button can be used to quit the Print Culivation Time Chart window without using the print function Exporting the Results The Export button opens the Export window from which it is possible to export selected data in various formats see Exporting the Cultivation Time Chart Data page 59 54 User Manual Cedex Version 1 6 1 The Analysis Results O Entering a Comment The Comment button opens the Comment of the View window that offers the possibility of entering a comment on the current grouping of measurement series View The comment is stored and the window is closed using OK Shielding the Cultivation Time Chart Window The Hide button fades out the Cultivation Time Chart window The Cedex Workbench is again visible The Cultivation Time Chart button can again fade in the window without changing it The Extras Area The Fit lowest common value button enables automatic shifting of
58. age 93 22 User Manual Cedex Version 1 6 1 10 4 2 Installation and Operation ofthe Cedex System D Fill 70 Ethanol or 70 Isopropyl in the Ethanol container Fill distilled or de ionized water in the water container The waste container should be empty when placed in its position and should be checked often to prevent it from overflowing There should always be a sample cup attached to the sample port even on delivery refer to Figure 1 on page 13 The sample cup is correctly attached when it is in an upright position beneath the sample port and the capillary tube almost reaches the base of the sample cup Preparation and Configuration of the Analysis Software by the Cedex Administrator If the hardware devices are installed and switched on in their correct order the operating system of the PC will start automatically The Cedex administrator has to execute the following operations before the analysis software can be used regularly Securing access rights and defining the administrator password Creating user profiles and delegating access rights Defining target directories for the analysis data Calibrating the measurement system Detailed information is found in the Cedex Administrator Manual After successfull configuration of a user profile the access rights of this user can be defined Thus the access to the following areas is determined Sample data Reprocess data Available work areas Default work area Cell
59. alibrate focus and brightness available from innovatis AG Germany Cedex Calibration Beads 20 mL Cat No 7 60 00 00 16 Density Reference Standard Beads to check the counting routine innovatis AG Germany Cedex Density Reference Standard Beads 10 mL Cat No 7 60 00 00 17 Spare parts e g for the Syringe available from innovatis AG Accessory device for the Cedex AS20 Auto Sampler available from innovatis AG Material Safety Data Sheets for the Substances used Material safety data sheets for all substances used are available from the appropriate supplier User Manual Cedex Version 1 6 1 93 Cedex Declaration of Conformance O nnowris Cedex DECLARATION OF CONFORMANCE Supplier name innovatis AG Supplier address innovatis AG EMC test EMC test Meisenstr 96 Berg amp Lukowiak TUV Rheinland 33607 Bielefeld GmbH Product Safety GmbH Germany 32609 H llhorst Am Grauen Stein Lohner Str 157 51105 Koln Germany GERMANY Declares under our sole responsibility that the following product Product series no Cedex Product name Cedex System Conforms to the following standards Normative Standards FCC CFR 47 Pt 15 subpart A clause 15 107 and clause 15 109 CENELEC EN 55011 1998 Class B EN 61000 6 3 2001 EN 61000 6 2 2001 Emissions A radiated and conducted Classification Product conformance to cited product specifications is based on sample type testing evaluation or assessment at Berg
60. amp Lukowiak GmbH EMC Services in H llhorst Germany and T V Rheinland Product Safety GmbH in K ln Germany This product or family of products is eligible to bear the appropriate compliance logos and statements Head of production innovatis AG Bielefeld Germany June 28 2003 94 User Manual Cedex Version 1 6 1 Index 8 8 way VAIVE nnenesenneennnesnnnnnnennnnnnnennnnnn 89 A ACCESS rights ccceeeeeeeeeeeeeteees 33 74 AGAn sndddcatvbebuvedsndacvervanaaesaniud 86 Additional Information 61 Administrator functions 75 Aggregate Histogram en 44 Aggregate Size 44 Air bubbles 1 0 0 0 eeeseeceeeeeeaeeeeeeeees 36 42 AlCOhdl nern 82 Analysis parameters en 35 Analysis process ueenneenennnennnnnn 35 Analysis results enea 41 70 Analysis software ueeneeennnnn 23 Archive mode cceeeeeeeeeceeeeeaeeeseeeeeeees 70 Area Additional Information 61 Area CSV Options 61 Area Data Selection 51 Area Data Series uuseesssseeeeeeeeennnnennen 55 Area Data Series Mixing 60 Area Export Structure 61 64 Area Export Template 61 Area Extras a ana 55 Area General cccccccccsaccccssesseccecstecsecsecaace 54 Area Output File Name
61. ck on Remove Syringe in the Syringe handling area The syringe barrel is then moved in a position to allow easy removal Unscrew the protective cover 4 Hold the syringe barrel by the Luer lock and turn it anti clockwise to remove it from the 8 way valve 5 Once the connection is loosened keep holding the syringe barrel in an upright position beneath the 8 way valve to prevent it from jamming with the plunger 6 Remove the bottom screw that connects the plunger to the syringe module with the other hand 7 Remove the syringe from the syringe module User Manual Cedex Version 1 6 1 85 Cedex Cleaning the Syringe The syringe should be cleaned according to the level of use and degree of soiling In the Case of Mild Soiling Follow the instructions below 1 Remove the plunger from the syringe barrel 2 Rinse both parts with distilled or de ionized water and then with 70 alcohol 3 Clean the Teflon head seal of the plunger by wiping it carefully with a lint free cloth 4 Rinse both parts thoroughly with distilled or de ionized water 5 Replace the plunger in the syringe barrel Both parts must be wet when assembled In the Case of Heavy Soiling The syringe should be cleaned thoroughly once a week using an intensive cleaning procedure This is done with either a weak detergent or an acid base combination IMPORTANT NOTICE THE SAFTEY REGULATIONS SHOULD BE OBSERVED WHEN HANDLING ACIDS AND BASES Follow the instructi
62. conds In version 1 5 1 and 1 6 1 default is set at 0 seconds i e Auto logout is inactive 22 Administrator Manual Cedex Version 1 6 1 General Administrator Functions O Proceed as follows to set the time for automatic log out 1 Login as Administrator in Windows NT with the password Administrator and double click on the Support folder on the desktop 2 Double click on connections to Cedex ini and the file Cedex ini will open The following text will appear in the section system in the file Cedex ini AutoLogoutTime 0 which indicates that the auto logout is inactive 3 Replace 0 in the respective row by the number of seconds after which Auto logout should be activated Entries of 0 to 999999999 seconds are allowed IMPORTANT NOTICE ANY MISTAKES IN THE ENTRY LIKE ACCIDENTALLY PRESSING THE SPACE KEY WILL DISRUPT THE PROGRAM 4 Select Save in the File menu to store the modifications Close the file Cedex ini 6 Shut down the Windows program Begin with measurements by starting the Cedex system in the usual way Login as described in the Cedex Operating Instructions with the username Cedex and password Cedex 4 2 Adaptation of the Counting Routine Normally the Cell Type Standard Cells controls the image recognition making it unnecessary to modify the counting routine The Cell Type Standard Cell counting routine which is integrated into the Cedex system is set to suit all types of standard measurements However und
63. creator Modified 03 07 2002 17 58 03 07 2002 17 58 Special Version A JA new Template Save Figure 28 The Save an Export Template window 62 User Manual Cedex Version 1 6 1 The Analysis Results O The Load Button The Load button opens the Load an Export Template window in which the user can select an existing Export Template for loading Name creator Modified Default i 03 07 2002 17 58 Special Version A 03 07 2002 17 58 Standardformat Administrator 03 07 2002 17 58 Default Load l Show my files only Figure 29 The Load an Export Template window Through the Show my files only checkbox the display in the Load an Export Template window is limited to the own Export Templates only The name is automatically adopted entry row by clicking the desired template in the list Likewise it is also possible to enter the name in the entry row directly The use of the button Load results in loading the choosen Export Template Using Cancel the window can be quit without loading an Export Template The Delete Button Stored Export Templates can be deleted using the Delete button The Delete an Export Template window in which the user selects the Export Template to be deleted opens Actuating the Delete button and subsequently OK deletes the Export Template Only self created templates can be deleted Using Cancel the window can be quit without deleting a template Available Export Tem
64. ct an existing one from the list The list contains only Views set up by the user himself There is no possibility of overwriting Views created by others By clicking a list entry the name is automatically adopted in the entry row The entered name can still be modified afterwards If the selected View name already exists a safety challenge on whether the existing file should be overwritten appears If the current View already has a name this name is automatically adopted in the entry field The window can be quit with the Cancel button The View data is stored under the given name using the Save button 52 User Manual Cedex Version 1 6 1 The Analysis Results O Loading a View The Load button opens the Load a View window in which the View file to be loaded can be selected Name Creator Modified Default i 03 07 2002 17 58 Administrator 03 07 2002 17 58 Test Cedex 03 07 2002 17 58 Default Load Cancel F Show my files only Figure 21 The Load a View window Via the Show my files only Checkbox the display in the Load a View window is limited to one s own Views only In the list of the Load a View window the individual name of the View the creator s user name and the date of the last modification are shown The window can be quit using the Cancel button The name of the View is automatically adopted in the entry row at the click of an entry in the list The corresponding data is loaded using the Load butt
65. ctions O 4 General Administrator Functions The following system functions are recommended to be only available to the Cedex administrator Administration of user profiles Adaptation to the Operator Standard Cells counting routine Calibration of the measurement system see Calibrating the Measurement System on page 17 4 1 Administration of User Profiles The user usually has limited access rights All administration rights relating to user profiles i e creating deleting modifying and delegating rights should remain exclusively with the Cedex administrator Creating User Profiles Detailed instructions can be found under Creating User Profiles and Delegating Access Rights on page 15 Deleting User Profiles Follow the instructions below Click on the Users menu on the Workbench Click on Delete and the Select User to delete list will appear Select the user profile to be deleted 1 2 3 4 Click on Delete and you will be requested to confirm the entry 5 Click on Yes to confirm and a message will confirm the entry 6 Click on OK to shut the window PLEASE PAY ATTENTION TO CHAPTER RE ISSUING USER NAMES PAGE 31 Modifying the Access Rights of a User Profile Detailed instructions can be found under Creating User Profiles and Delegating Access Rights on page 15 Administrator Manual Cedex Version 1 6 1 21 Cedex Modifying a Username If an existing user is to be replaced by a
66. d consists of top quality electronic and optical components To guarantee optimal and reliable performance of the Cedex system select the appropriate location according to the safety regulations described in the first section Connecting Hardware Components and Hardware Key Dongle See Figure 5 and Figure 6 Follow the instructions below to connect the hardware components Connect monitor keyboard and mouse to their respective ports on the PC Connect one end of the video cable to the Video port of the measurement system Connect the other end of the video cable to the Video port of the PC Connect one end of the first serial cable to the Focus port of the measurement system Connect the other end of the first serial cable to the Focus port of the PC oa PF wn Connect one end of the second serial cable to the Syringe port of the measurement system N Connect the other end of the second serial cable to the Syringe port ofthe PC Connect the Hardware key USB Dongle to the USB port ofthe PC see Figure 6 NOTE CHECK THAT YOUR CONNECTIONS ARE CORRECT OTHERWISE AN ERROR MESSAGE WILL APPEAR CAN NOT ACCESS LIQUID MANAGEMENT OR SCOPE FUNCTION DISABLED lex 100 250V 2 3A 50 60Hz c u la 25 r Fuse CE innovatis GmbH Z 63H 2500 CED Typ God Made in Germany Focus Syringe Video Sai Noa TS Vor ffnen des Ger ten Notestockor ziehen Before opening disconnect mains Avant Jouve fappared retirez la chem
67. der the microscope and actual cell count is done automatically by the Cedex system The recognition and differentiation of cells as either viable or dead is performed under standardized conditions and therefore provide very precise and objective results These results are evaluated automatically and displayed From version 1 5 0 1 on there is a possibility to analyze display and archive different measurement results in comparison The result data Viable Cell Density Total Cell Density Viability Total Cell Count Average Compactness Average Diameter und Standard Deviation of single measurements Image and overall count Overall are supplemented by the Aggregate Rate This result shows the percentage of total cells in aggregates cluster The graphical representations of the measurement results Measurement Chart Diameter Histogram Compactness Histogram were supplemented by the Aggregate Histogram and the Cultivation Time Chart The Aggregate Histogram gives a quick insight of cluster behavior of cell cultures The Cultivation Time Chart displays the grows curves and cultivation parameters in a time dependent manner The results of the Cultivation Time Chart can be separately printed out For further processing of the results in other programs data export as DIF Data Interchange Format Excel compatible and CSV Comma Separated Values generally applicable is possible The Cedex 1 6 1 version apart from adaptation to the Windows 2
68. dex system Click on Reset Initialize in the Liquid Management window Switch off the Cedex system Lubricate the bottom part of the syringe guide and spindle Switch on the Cedex system Click on Remove Syringe in the Liquid Management window Install the syringe see Assembling and Installing the Syringe on page 87 syringe guide spindle Figure 34 Syringe Guide and Spindle 88 User Manual Cedex Version 1 6 1 Maintenance of the Measuremment System D 9 6 Maintenance of the Measurement Chamber Flowcell Microscope Optic 8 Way Valve and the remaining Hardware Components The Cedex must be serviced every six month by an authorized service technician This service ensures that worn and defective parts are replaced The following routines must be carried out every six months Maintenance of the Syringe Guide and the Spindle Replacement of the fuse by a service technician e Inspection of the crimp connections by a service technician The following routines must be carried out every year e Replacement of the syringe by a service technician Replacement of the 8 way Valve by a service technician The following routines must be carried out every two years e Replacement of the bulb by a service technician Replacement of the pump belt by a service technician Replacement if need arises of the measurement chamber by a service technician IMPORTANT NOTICE ONLY AN AUTHORIZED SERVICE T
69. e software or documentation or failure to comply with the limitations outlined above shall automatically terminate the license 8 User Manual Cedex Version 1 6 1 General Provisions The licensee may terminate this license at any time by destroying all copies of the software or returning all copies to innovatis AG Innovatis AG may terminate this license if the licensee fails to comply with all terms and conditions of this license agreement In such an event the licensee agrees to destroy or return all copies of the software The licensee further agrees to be responsible for all taxes resulting from this license Any unauthorized leasing renting or transferring except as outlined above is illegal If any provisions of this license agreement are invalid for whatever reason this shall have no effect on the remaining terms hereof The license agreement is governed by the laws of the Federal Republic of Germany This is the only agreement between innovatis AG and the licensee and replaces any other negotiations or prior agreements relating to the software and documentation Cedex Reg U S Pat amp TM Off FCC Notification WARNING CHANGES OR MODIFICATIONS TO THIS UNIT NOT EXPRESSLY APPROVED BY THE PARTY RESPONSIBLE FOR COMPLIANCE MAY VOID THE USER S AUTHORITY TO OPERATE THE EQUIPMENT Note This equipment has been tested and found to comply with the limits for a Class A digital device pursuant to Part 15 of the FCC Rules These
70. e Measuremment System D Dead Cells Size Range 40 250 Default 110 Unit um Defines the minimum cross sectional area of a particle in um Increasing this value means that less particles will be recognized as dead cells Dead Cells Appearance Range 10 10 Default 0 Unit absolute measure Dead cells have a variety of appearances under the microscope This can range from sharply outlined small dark dots to less distinct objects with varying brightness and contrast This parameter distinguishes large dark objects which are typical dead cells from less distinct objects Increasing this value means that there are more deviations from typical dead cells resulting in a higher count of dead cells Dead Cells Aggregate Minimum Range 250 1500 Default 360 Unit um Defines the minimum cross sectional area of a dead cell aggregate cluster When this size is reached it is then automatically partitioned into single cells Dead Cells Aggregate Maximum Range 1500 5000 Default 2250 Unit um Defines the maximum cross sectional area of a dead cell aggregate cluster The dead cell aggregate area within this range is automatically partitioned into single cells Any aggregate larger than the specified maximum area is not taken into account Dead Cells Aggregate Appearance Range 10 10 Default 0 Unit absolute measure Defines the automatic evaluation of a dead cell aggregate cluster T
71. e list The measurement series are not separately listed but rather all data is grouped together in a big list The data is thereby directly attached one after the other according to the sequence of the Data Series list List for each data series side by side The single measurement series are listed beside each other in a data block each There is an empty column space between the measurement series List for each data series among each other The single measurement series are successively listed in a data block each There is an empty row space between the measurement series The Additional Information Area Whether additional information should be adopted in the export file can be selected in the Additional Information area Add headline column names Adding headlines to the data to be see The Export Structure Area page 64 e Add creation information Adding further information to the measurement series Current User Name of the user who created this export Date Time Date Time of the export Device ID Appliance number of the Cedex System on which the export was created Cedex Version Programme version of the Cedex application Export File Name Path File name of the created export file Series 1 Number of data sets of the first measurement series Series 2 Number of data sets of the second measurement Series 3 Number of data sets of the third measurement Series 4 Number of data sets of the fourth measurement Series
72. e software product must be treated as any other material protected by copyright However this shall not apply to the following The Licensee may e only use the software on the PC supplied with the Cedex system only produce copies of the software for internal archiving and backup purposes i e on condition that all property and copyright notices shall be included on the copies only transfer the software on a permanent basis excluding leasing and renting in conjunction with a transfer of the entire Cedex system and on condition that the licensee retains no copies and the recipient accepts to be bound by the terms of this agreement The Licensee may not e use the software on more than one computer workstation or file server in a network or multiple user system except if an additional license for every additional computer or workstation is purchased from innovatis AG modify copy adapt reverse engineer decompile disassemble or create derivative works of the software or related documentation excluding a backup copy lease or rent the rights on the software and related documentation in any form whatsoever to a third party remove property and copyright notices and marks in or on the software documentation containers and devices All rights claims use and intellectual property rights relating to the software documentation and any copies made by the licensee remain with innovatis AG Unauthorized copying of th
73. easurement System on page 22 7 Have the cell suspension samples ready for the analysis To Start the Cedex Workbench Follow the instructions below 1 Start the operating system and press lt Ctrl gt lt Alt gt lt Delete gt to login 2 Enter username CedexSA and password CedexSA and confirm with lt Enter gt The CedexSA user has no access to the desktop and can therefore only use the Cedex program for further information ask your Administrator The Cedex analysis software starts automatically and the login window appears The password is case sensitive Enter the username in the Username field Enter the password in the Password field if applicable Click on the traffic light symbol to login or press lt Return gt The Cedex Workbench appears after a successful login and the traffic light symbol changes to green The administrator or user have access to the functions once they have logged into the Cedex Workbench The user normally only has access rights to the routine operations depending on the configuration However access rights may vary according to options chosen for each user profile The administrator has unlimited access rights to all functions and can modify user access rights at any time see Modifying the Access Rights of a User Profile on page 75 28 User Manual Cedex Version 1 6 1 Working with the Analysis System 5 The Loader Program When the Cedex program is started the Loader program automa
74. ecessary to adjust this counting routine e g for cell cultures with an extraordinary appearance or for specific internal counting strategies i e if certain aggregates or cell nuclei are to be counted or not In this case the administrator can adjust the image processing and evaluation strategy of the Cell Type Standard Cells to meet specific requirements Adjusting the Cell Type Standard Cells is also known as creating a new Cell Type because a new Cell Type is created or an existing Cell Type is modified This can be done by changing certain parameters that are based on the Cell Type Standard Cells counting routine The Cell Type Standard Cells is based on seven parameters and one algorithm Each parameter directly influences the image processing and evaluation strategy and therefore the results The present software version has only one algorithm Operator User Manual Cedex Version 1 6 1 75 Operator Y5 0 0 1 Operator Std Cells V 5 00 Algorithm Standard Cells 5 00 Living Cells Dead Cells Dead Cells Dead Cells Dead Cells Dead Cells Min Pollution Appearance Size Appearance Aggreg Min Aggreg Max Aggreg App Area 20 20 fo Revert 250 40 110 Revert 10 10 pi fo Revert 1500 5000 250 T 1500 I 360 2250 Revert Revert 10 Revert 30000 Cancel Figure 31 Setting Cell Type Standard Cells parame
75. ells nn 23 Suitable location en 12 System functions sseeenneenenn 16 T Target directories 16 U User profile u unununeneneneneeeeeeeenen 15 21 Username cece cece ce eeeeeeeaeaeaeaeceaeeeeneneaes 22 Ww Warranty ccd id eennee ceed ah ine 5 Work area in 15 Administrator Manual Cedex Version 1 6 1 37
76. enance of the measurement chamber flowcell the microscope optic and the 8 way valve This maintenance routine must be carried out only by an authorized service technician This chapter provides detailed instructions and guidelines for each individual cleaning and maintenance procedure 9 1 Cleaning and Maintenance Check List This check list shows when each cleaning and maintenance routine should be done Carry out these routines on a daily basis Control of the reagent container each time prior to use Filling status contamination LM ShutDown Always after the last measurement and before a Cedex system shut down e Standard Clean After the final measurement in a set of samples or before a longer pause Chamber Clean 2 At least twice a day Carry out this routines on a weekly basis Intensive Clean Change of Trypan Blue solution Cleaning the reagent containers and refilling them with fresh reagents Carry out this routine on a half monthly basis e Special Intensive Clean At least twice a month with 0 1 1 N HCL instead of von detergent see page 82 Carry out these routines on a regular basis e Removal and cleaning of the syringe Cleaning routines carried out by the Liquid Management Cleaning the external surface of the capillary tubes with a lint free cloth Calibration of the measurement system after every three months or after any change of position or vibration of the measurement
77. ennn 81 83 85 Syringe handling 83 System functions u enseesensnnnnee nennen 33 T Total assein Hein 45 Total cell count 20000 15 41 Total cell density 0 en 41 Traffic light symbol eee 32 Trypan Blue He 14 U Ultra Fast Clean 80 User identification area 31 User profile nee 75 Username aan an 75 Users menu uuuueeassnnesnnnnnnnnnnnnnnnnnnnnnn anne 31 V Valid image nesnnennnnnenennnnnennnn 68 Viability cerae 41 Viable cell density 2 00 41 Viable cells aas 14 View delete iiaii annaa 52 View load 0 0 eee cc cece eeeeceeeeeeeeeaeeaeeeeeeeeeaees 52 View SAVE ouar aa 52 Volume 80 Volume variations uuuuennneneeaannenennn 36 W Warranty usa 7 Waste container 80 Walter aussen een 23 Work afed a nein 23 Workbench uuu e enae 30 User Manual Cedex Version 1 6 1 97 Administrator Manual Cedex Version 1 6 1 e QUALITY THAT COUNTS Version Cedex 1 6 1 10 29 03 MM DD YY 2 Administrator Manual Cedex Version 1 6 1 Contents CONTENTS euere een 3 1 RIGHTS are 5 COP YN IG 1 NOPPREFIEEPEEEEPEERPEHLFRELEEEFFEPREHEFEFLEEFEPFEESEHREFFEURTTEFESDEEEEFEFESEEFTPLIUFEFFELREHERESEHEERETESLERFLFEEPREEFFEPRLELFEEPFER 5 DISGIALIMNGR GOPPPEPREEFRPPPEERFPFFPPREEPFEPPEEFERFEPFEELEPLPEEEFFPFEEFEPPEEPEEEEPEPLPEEFEFPFFEEEFPEEPEEFF
78. er certain circumstances it may be necessary to adjust this counting routine e g for cell cultures with an extraordinary appearance or for specific internal counting strategies i e if certain aggregates or cell nuclei are to be counted or not In this case the administrator can adjust the image processing and evaluation strategy of the Cell Type Standard Cells to meet specific requirements Adjusting the Cell Type Standard Cells is also known as creating a new Cell Type because a new Cell Type is created or an existing Cell Type is modified This can be done by changing certain parameters that are based on the Cell Type Standard Cells counting routine The Cell Type Standard Cells is based on seven parameters and one algorithm Each parameter directly influences the image processing and evaluation strategy and therefore the results The present software version has only one algorithm Operator Administrator Manual Cedex Version 1 6 1 23 Operator Y5 0 0 1 Operator Std Cells V 5 00 Algorithm Standard Cells 5 00 Living Cells Dead Cells Dead Cells Dead Cells Dead Cells Dead Cells Min Pollution Appearance Size Appearance Aggreg Min Aggreg Max Aggreg App Area 20 20 fo Revert 250 40 110 Revert 10 10 pi fo Revert 1500 5000 250 T 1500 I 360 2250 Revert Revert 10 Revert 30000 Cancel Fig
79. esults in the capillary tubes and 8 way valve being filled with the appropriate reagents All measurements are based on the capillary tubes having a precise amount of reagent volume in them Therefore not performing a Prime would lead to cell samples being incorrectly stained and consequently leading to false measurement results A Prime is not only performed when the Cedex system is set up for the first time but it should follow after every start of the measurement system i e at least once a day prior to the first measurement No measurement is possible without performing a Prime first because an error message appears Machine is not ready for measurement User Manual Cedex Version 1 6 1 33 Cedex Follow the instructions below to perform a Prime 1 Click on Liquid Management on the Cedex Workbench The Liquid Management window will appear This window is divided into several different functional areas see Maintenance of the Measurement System on page 79 Ensure that a clean and empty sample cup is correctly attached to the sample port Click on Prime The duration period of a Prime procedure is displayed in the Time to finish area The following message System ready appears when the Prime has been completed 4 Click on XJ to shut the window Choosing the Operating Option and Specifying Sample Data The Cedex Workbench offers three operating options 1 to carry out a new measurement 2 to view existing anal
80. evaluation of a dead cell aggregate cluster The cross sectional area of dead cell aggregates are automatically partitioned into single cells Increasing this value means that there are more single cells determined in the aggregate Minimum Pollution Area Range 5000 30000 Default 5400 Unit um Defines the minimum cross sectional area of the pollution size e g debris particles air bubbles etc Any pollution size larger than the specified minimum area results in the entire image being excluded ATTENTION The use of the sliders espectally in combination can lead to unexpected results Administrator Manual Cedex Version 1 6 1 25 Cedex Modifying a Cell Type An existing Cell Type can be modified although it is NOT advisable for data safety A new Cell Type based on an existing Cell Type should be created instead Follow the instructions below to modify an existing Cell Type not advisable 1 2 3 4 5 6 Click on the Setup menu on the Cedex Workbench Click on Cell Types Select Edit and the Choose Cell Type to edit window will appear Select the appropriate Cell Type Click on Edit Adjust the individual parameters as required by clicking on the H and E buttons to increase or decrease their values Click on Revert just below the individual parameters to delete the adjustments again Click on Save to confirm the adjustments Click on Cancel to shut the window without saving any changes Creating a New
81. g images with the calculator symbol Viewing Existing Analysis Results Modifying Measurement Parameters Existing data sets may be viewed and corrected in the archive mode which is available to all system users The options in the archive mode are however dependent on specific access rights of users Click on the floppy disk symbol H archive in the Data Source Identification area on the Cedex Workbench to access the archive mode 70 User Manual Cedex Version 1 6 1 The Analysis Results O The archive mode offers the following options Entering modifying or deleting a comment see Entering a Comment on page 35 Correction of the dilution see Defining the Measurement and Analysis Process Parameters on page 35 Re evaluation of cell images i e excluding or including images see Viewing and Selecting Cell Images on page 65 Viewing printing the analysis results see The Analysis Results on page 41 and Printing the Analysis Results on page 69 Follow the instructions below to work with the archive mode 8 Click on the operating option H archive Select the appropriate reactor ID from the Reactor ID drop down list Select the appropriate sample ID from the Sample ID drop down list If the data set has not yet been reprocessed then a selection in the Data Set area only Original Data available is not possible If the selected data has been reprocessed then a selection between the original data and the
82. g measurements 34 User Manual Cedex Version 1 6 1 Working with the Analysis System 5 Entering a Comment Entering a comment relating to a measurement is optional and only possible once the Data Source Identification area has been completely filled in and the traffic light symbol is green Enter the remarks in Comment on the Sample Properties area Defining the Measurement and Analysis Process Parameters The measurement and analysis parameters can be defined in the respective space of the Process Parameters area on the Cedex Workbench In principle all routine measurements should be carried out using the standardized default settings that are factory set Follow the instructions below to change the settings 1 Select the cell type from the Cell Type drop down list factory set only one option available Standard Cells 5 00 If a new Cell Type has already been created a selection would have been possible at this point Detailed information on this is found under Creating a New Cell Type on page 77 2 Choose the respective dilution from the Dilution drop down list or enter a separate dilution e g 1 3 1 6 1 7 etc When a dilution of 1 2 is prepared then 1 part cell suspension is added to 1 part dilution medium A dilution of 1 3 means that 1 part cell suspension is added to 2 parts of a dilution medium etc It may be necessary to dilute samples with very high cell densities gt 1x 10 Cells mL manua
83. g measurements due to debris particles or air bubbles is automatically recognized as invalid and excluded from all calculations by the Cedex analysis software Users can also manually exclude individual images from the calculations or include images that were automatically excluded by the software The Cedex analysis software enables a close view of the single image or an enlarged display for evaluation Viewing Individual Cell Images Individual images can be viewed on the Cell Image area Follow the instructions below Select the appropriate image using the arrows in the Cell Image area Only one a is displayed but an overview of all cell images can be displayed by clicking on sss User Manual Cedex Version 1 6 1 65 Cedex Enlarging a Cell Image Follow the instructions below 1 Select the appropriate image using the arrows in the Cell Image area 2 Move the cursor on the indicated image and click on the left mouse button to display an enlarged image click on the left mouse button again to return it to normal size Click on the right mouse button to open the ResultViewer see page 67 and display cells that were identified as either viable or dead as described in the next section Viewing a Cell Image Using the IP ResultViewer The IP ResultViewer contrary to the standard display marks all viable and dead cells of a cell image The user then has an idea of how abnormal cell shapes and clusters are evaluated by the software
84. ge 29 To start the Cedex analysis software additional steps are necessary Starting the Analysis Software Follow the instructions below 1 Start the operating system by pressing the keys lt Ctrl gt lt Alt gt lt Delete gt to login 2 Enter Administrator as username and Administrator as password or Cedex as username and Cedex as password the password is case sensitive and confirm by pressing lt Enter gt The Windows surface will appear 3 Start the Cedex program by double clicking on the corresponding icon Configuring the Analysis Software After successfully carrying out the instructions above the analysis software must now be configured and prepared for routine operations This involves four steps 1 Securing access rights and defining the administrator password 2 Creating a user profile and delegating access rights Administrator Manual Cedex Version 1 6 1 13 3 Defining target directories for the analysis data 4 Calibrating the measurement system Securing Access Rights and Defining the Cedex Administrator Password The Cedex analysis software and its graphical user interface known as the Cedex Workbench are at first only configured for the administrator The administrator password has not yet been set which means that free access to the software and all connecting data is possible Please read chapter 5 2 Access Control at the Cedex Level page 29 to get more information A Cedex Workbench
85. ge module with 8 way valve sample port with sample vial 2 5 mL syringe switch ON OFF tray with 5 reagent containers from left to right Trypan Blue detergent 70 alcohol distilled or deionized water waste Figure 1 Components of the measurement system 3 1 The Measurement System The measurement system consists of two main components the hardware to capture microscopic images and the liquid management system The measurement chamber flowcell is highly sensitive and especially developed to analyze cell images in a defined volume of cell suspension The Liquid Management undertakes the sample staining as well as the filling and cleaning of the measurement chamber including the cleaning of the entire measurement system The precise volume of sample and dye as well as specific cleaning solutions required for each measurement is transported through the syringe module with the 8 way distribution valve The individual components of the liquid management system are easily accessible to allow for trouble free maintenance and control User Manual Cedex Version 1 6 1 13 Cedex 3 2 The Measurement Procedure The determination of viable cells cells mL and cell viability of cell suspensions the percentage of viable cells compared to the total number of cells is a fully automated and highly standardized procedure based on the manual Trypan Blue exclusion method A defined volume of Trypan Blue is aspired by the syringe to the
86. h detergent and distilled water for approx 60 seconds The entire liquid management system is rinsed with distilled water then with detergent and finally with distilled water Use this routine in between measurements to remove dirt particles and air bubbles quickly from the measurement chamber flowcell Standard Clean with alcohol detergent and distilled water for approx 90 seconds The entire liquid management system is rinsed with distilled water alcohol detergent and finally with distilled water again This routine removes dirt particles and is used after measuring a set of samples or before a longer pause Intensive Clean with alcohol detergent and distilled water for approx 20 minutes The measurement chamber flowcell is thoroughly rinsed with distilled water alcohol distilled water detergent and finally distilled water The detergent remains in the measurement chamber for a longer period to react on resistant particles This routine is used on weekly basis to remove heavy and resistant contamination of the measurement chamber capillaries tubes syringe and 8 way valve Even the tiniest dirt particle attached to the sensitive measurement chamber can cause the glass wall to become uneven and result in aggregates building These particles are not necessarily visible in the microscopic image Cell Image but have a major impact on measurement results If measurement values within a sample measurement show large varia
87. he cross sectional area of dead cell aggregates are automatically partitioned into single cells Increasing this value means that there are more single cells determined in the aggregate Minimum Pollution Area Range 5000 30000 Default 5400 Unit um Defines the minimum cross sectional area of the pollution size e g debris particles air bubbles etc Any pollution size larger than the specified minimum area results in the entire image being excluded Detailed instructions of Modifying a Cell Type e Creating a New Cell Type Deleting a Cell Type can be found in the Cedex Administrator Manual User Manual Cedex Version 1 6 1 77 Cedex 8 3 Saving the Measurement Data Only Cedex user which have full access to the Windows desktop can start the burning software to save the mesurement data on CD Detailed instructions can be found in the Cedex Administrator Manual 78 User Manual Cedex Version 1 6 1 Maintenance of the Measuremment System D 9 Maintenance of the Measurement System The measurement system should be regularly serviced to guarantee optimal performance and maximize the life span of the Cedex system This includes the following maintenance procedures e Recurrent manual maintenance and cleaning routines if required e Cleaning routines with the Liquid Management e Syringe cleaning and maintenance Maintenance of the capillary tubes Maintenance of the syringe guide and spindle Maint
88. he y axis The combination of parameters is only possible for a maximum of two different units since only two ordinates left and right are available The values for the cell densities Cells mL are thereby always displayed on the left ordinate If display for viability is combined with diameter or compactness values for the viability appear on the left ordinate and those for the diameter or the compactness on the right ordinate respectively If diameter and compactness are combined together the left ordinate displays the diameter value and the right one the compactness value respectively User Manual Cedex Version 1 6 1 51 The View Area The measurement series grouped by the user for the Cultivation Time Chart creating a View including further information can be stored All information about this View is then displayed in the View area Name Name of the View user defined Creator User name of the creator Designed Date of creation of this View Modified Date of the last modification of this View The Save Load and Delete buttons can save load or delete a View grouping of the measurement series Saving a View The Save button opens the Save a View window in which the name of the View the grouping of the measurement series saved is set Name creator Modified 03 07 2002 17 58 My Special View Save Figure 20 The Save a View window In the entry row the user can enter a new name for the View or sele
89. hrough the measurement chamber and then pressed back again with compressed air Use this routine twice daily to remove resistant protein aggregates fluff particles or other obstructions from the measurement chamber The Syringe Handling Area The syringe plunger position is set here for the manual cleaning process see Syringe Cleaning and Maintenance on page 85 Carrying out a Cleaning Routine The different cleaning routines always have the same starting procedure Ensure that there is always a sample cup attached to the sample port Follow the instructions below 1 Click on Liquid Management on the Cedex Workbench and the Liquid Management window will appear 2 Click on the appropriate button e g Fast Clean to automatically start the cleaning routine The Cedex system is blocked during the cleaning routine and the duration is continually displayed on the Time to finish field User Manual Cedex Version 1 6 1 83 Interrupting Terminating a Cleaning Routine Use the EMERGENCY STOP button to interrupt the cleaning routine immediately if any leaks are detected in the liquid management system even in the case of the Ultra Fast Clean that is carried out automatically The Intensive Clean procedure can be interrupted to shorten the long detergent phase in the measurement chamber flowcell Click on the Cancel button in the Liquid Management window which is only activated during the long detergent phase IMPORTANT NOTICE
90. iameter jim 11 77 10 40 Aggregate Rate 16 9 10 1 Standard Deviation 10 2 2 42 Aggregate Histogram Diameter Histogram Measurement Chart Compact Histogram Print results O Neno Figure 9 Cedex Workbench with the final results Cultivation Time Chart Liquid Management User Manual Cedex Version 1 6 1 37 Cedex End of Measurement The cleaning procedure begins immediately after the last image is captured and displayed on the Process Parameters area The entire liquid management system is rinsed during this procedure Ultra Fast Clean Once this procedure has been completed the Process Parameters area displays Ready and the floppy disk symbol indicates that the next measurement can begin If the next measurement belongs to the previous measurements i e the same Reactor ID then the consecutive Sample ID will be automatically entered into the Sample ID area by the analysis software without further entries being necessary Click on the microscope symbol for the next measurement to begin When the final measurement in a set of samples has been completed or if there is a longer pause before the next measurement then the user should carry out a Standard Clean in the liquid management system besides the Ultra Fast Clean procedure that is automatically done Logout of the Cedex system once the cleaning procedure has been completed to allow the next user access to the Cedex system see Maintenance of the Measurement Sy
91. ide of each slider The borders of the first interval are defined by the minimum value and the first numerical value entered The next two values define the upper border ofthe second and third interval respectively The fourth interval is defined by the upper border of the third interval and the maximum value The amount of viable cells their percentage of the total viable cell number and the viable cell densities x10 Cells mL are listed in a table for each of the individual intervals respectively To obtain full details on viewing the individual cell images see Viewing Individual Analysis Results after Completion of the Measurement Process on page 42 48 User Manual Cedex Version 1 6 1 The Cultivation Time Chart The Analysis Results O Access to the Cultivation Time Chart takes place on the Cedex Workbench through the Cultivation Time Chart button In the Cultivation Time Chart cultivation progress result diagram growth or culture progresses of cell cultures can be graphically evaluated Besides there is the possibility of exporting and printing out the data After clicking on the button the Cultivation Time Chart window through which corresponding buttons call all the functions Cultivation Progress Diagram Export and Print opens 4 Cultivation Time Chart x 3 207 Data Selection nN 8 8 Cell Density 106 Cells mL Be 8 8 V Viable Cell Density in lin T Dead Cell Density
92. ids to close the reagent containers to avoid dirt particles getting into contact with the reagents 4 Ensure that the silicone tube and capillary tubes in the waste container only reach about 1cm into the container remove the lid NOTE THE TUBES SHOULD NOT BE SHORTENED UNDER ANY CIRCUMSTANCES 5 Fill 0 2 Trypan Blue solution on the recommendation of innovatis AG or 0 4 Trypan Blue solution in the respective reagent container refer to Figure 1 on page 13 If necessary dilute a more concentrated Trypan Blue solution with buffer Phosphate Buffered Saline PBS pH 7 4 Estimate the number of measurements to be determined before preparing the Trypan Blue volume To prevent the dye from being contaminated with micro organisms which could lead to false measurement results discard any left over dye solution after five days and rinse the container well afterwards see Cleaning the Reagent Containers on page 22 Dispose of any remaining Trypan Blue dye according to regulations For longer pauses of measurement store the Trypan Blue solution in a refrigerator NOTE ENSURE THAT THE APPLIED TRYPAN BLUE SOLUTION IS FREE OF CONTAMINATION OTHERWISE FILTER BEFORE USE 6 Fill the Cedex Detergent in the respective container Only the Cedex Detergent or a detergent with identical specifications should be used for the cleaning procedures to prevent damage to the sensitive surface of the Liquid Management components see Suppliers on p
93. ion Time Chart window in various ways One can thereby choose between the DIF and the CSV formats and in addition set further details such as the size and structure of the data These export settings can be stored as templates so that frequently used settings can quickly be used again User Manual Cedex Version 1 6 1 59 4 Export r Output File Name b Export dif Start Export r Output Format DIF Data Interchange Format i E Excel CSV Comma Separated Values r Additional Information V Add headline column names M Add creation information Hide r Data Series Mixing r CSV Options r Export Template Single file for each data series V Use IBM OEM Charset Name Common file for all data series T Use LF instead of CR LF Creator Arrangement Separator Designed One complete list C Comma Modified C List for each data series side by side risemicolonte List for each data series among each other C Tabulator Space Load Save Delete r Export Structure Available Data Export Data p Date Time Format Date Time BULL fal Dead Cells Total Cell Density 1045 cells mL T Day Month Device ID Viable Cell Density 1045 cells mL Move Up go Group Identifier Dead Cell Density 1045 cells mL e G 23 Operator Name Std Deviation 1045 cells mL Veer Bias RE Reactor ID 22 lAvg Compactness Mace Deer Yea
94. ions below 1 2 3 4 Ensure that the sample cup is correctly attached to the sample port Ensure that the reagent containers are sufficiently filled and in their correct positions Ensure that the waste container is empty Click on Yes to begin the measurement Every stage of the measurement procedure is continually displayed in the Process Parameters area The Cell Image area displays in succession the cell images captured by the camera including the corresponding analyzed results Image and the average values of the images analyzed so far of the sample Overall in the Result Data area see The Analysis Results on page 41 1 Cedex Workbench Eile Special Setup Users Help User Identification Username Administrator Password Shut down Logout Data Source Identification Sample ID Data Set Reprocess 1 gt Sample Properties Comment 5 FCS F Date Time 24 10 2003 17 40 Creator Administrator ioj xi innovatis AG 24 10 2003 17 41 20 r Process Parameters Cell Image 20 20 Cell Type sta Cells V5 00 z Dilution N one F Images 20 r Processtype Standard F Status Ready Audit Trail ERIC valia image Result Data Used images 20 Image Overall Viable Cell Density x10 13 88 15 63 Total Cell Density x10 18 59 21 19 Viability P6 74 6 PaT Total Cell Count 71 1619 Average Compactness 1 091 1 068 Average D
95. ivered e 1 x Measurement system 1x PC incl manual Windows 2000 and installed Cedex software and CD burner e 1 x Keyboard 1 x Mouse 1 x Mouse pad e 1 x Monitor incl manual e 1 x USB hardware key Dongle Cords 1 x VGA cord not attached to the monitor 2 x Serial cord to connect syringe focus control to the PC 1 x Video cable to connect the measurement system to the PC 3 x Connecting cable 1 x User Manual incl License agreement 1 x Administrator Manual 5 x Reagent container 250 mL each e 1 x 1000 mL Cedex Detergent Cat No see Suppliers page 93 1000 x Sample vial 4 mL Cat No see Suppliers page 93 e Cedex Calibration Beads for the calibration of focus and brightness Cat No see Suppliers page 93 IMPORTANT NOTICE PLEASE NOTIFY THE DISTRIBUTOR IMMEDIATELY IF THE SERIAL NUMBER ON THE CEDEX ANALYZER AND THE COMPUTER DO NOT MATCH MATCHING SERIAL NUMBERS ARE NECESSARY AS THEY ARE ESSENTIAL CRITERIA IN THE CALCULATIONS OF THE CEDEX SOFTWARE NOTE THE SOFTWARE AND HARDWARE VERSION OF YOUR CEDEX ANALYSIS SYSTEM CAN DIFFER FROM THE INFORMATION DESCRIBED HERE BUT SHOULD NOT MAKE A DIFFERENCE TO THE OPERATIONAL FUNCTIONS IN GENERAL NOTE A PRINTER IS NOT INCLUDED IN THE CEDEX SYSTEM FOLLOW THE INSTRUCTIONS TO INSTALL A PRINTER IN THE CEDEX ADMINISTRATOR MANUAL User Manual Cedex Version 1 6 1 19 Choosing a Suitable Location for the Cedex System The Cedex system is highly sensitive an
96. ld PF wn gt Enter a password optional for the user profile in the second and third fields or the new user can set up a password later 5 Define the access rights for the user profile and simply choose an existing user profile from the drop down list as a template This user profile can then be customized as shown below 6 Click on Create User to confirm the new user profile A message will appear to confirm its success Follow the same procedure later when adding modifying or deleting any user profiles Once a user profile has been successfully created further access rights can be included in the following areas Sample data Reprocess data Available work areas e Default work area Cell Type e Users System Administrator Manual Cedex Version 1 6 1 15 Cedex Description of Access Rights Each user profile can have specific access rights as described below Sample Data Access rights to sample data create view change delete e Reprocess Data Access rights to recalculate recorded measurements create view change delete allow changing work area Access rights to change the target directory Cell Types Access rights to evaluate Cell Types create view change delete Users Access rights to administer user profiles create view change delete System Access rights to specific system functions shut down the Cedex system shut down the Liquid Management change the
97. legated access rights The Specials Menu Normally only available to the administrator The individual menu items have the following functions Show IP Result Each cell recognized by the measurement system is marked as either dead red cross or viable yellow circle see Viewing a Cell Image Using the IP ResultViewer on page 66 Calibrate system Calibration of the analysis system see Viewing a Cell Image Using the IP ResultViewer on page 66 30 User Manual Cedex Version 1 6 1 Working with the Analysis System 5 The Setup Menu The menu items have the following functions Language To define the Cedex system language only English at present Change my Password The user password can be modified Show my Rights Displays the access rights of the present user Cell Types The analysis mode can be modified to suit specific cell types Normally only available to the administrator with possible viewing rights for the user see Cedex Administrator Manual Defaults To define default measurement parameters see Cedex Administrator Manual The Users Menu Normally only available to the administrator The individual menu items have the following functions Add Add user profile Delete Delete user profile Rights Edit user s access rights Change Name Change username Delete Password Delete user password The Help Menu It provides general information on the anal
98. limits are designed to provide reasonable protection against harmful interference when the equipment is operated in a commercial environment This equipment generates uses and can radiate frequency energy and if not installed and used in accordance with the instruction manual may cause harmful interference to radio communications Operating this equipment in a residential area is likely to cause harmful interference in which case the user will be required to correct the interference at his own expense Safety Instructions WARNING IT IS IMPORTANT THAT ALL SAFETY INSTRUCTIONS AND GUIDELINES ARE OBSERVED TO ENSURE SAFE AND RELIABLE OPERATIONS OF THE CEDEX SYSTEM e Place the device on a dry and sturdy surface free of vibrations Prevent the device from getting wet Protect the device from dust and smoke Do not expose the device to direct sunlight e Ensure there is sufficient air circulation of the Cedex system during operation particularly around the computer and monitor e Operate within a constant temperature range of 50 F 95 F 10 C 35 C e The Cedex system should only be used with the supplied devices measurement system computer monitor and software Do not install any additional software on the Cedex computer e Only trained personnel in a controlled laboratory environment are to operate the Cedex system User Manual Cedex Version 1 6 1 9 Cedex e All services and repairs should only be perfo
99. lly prior to measuring NOTE THE DILUTION RATIO ENTERED INDIVIDUALLY OR CHOSEN FROM THE DROP DOWN LIST ONLY RELATES TO THE RESPECTIVE SAMPLE WHICH WAS DILUTED MANUALLY AND IS ABOUT TO BE MEASURED DILUTIONS THAT ARE INTEGRATED IN THE MEASUREMENT PROCESS AND TAKEN INTO ACCOUNT BY THE CEDEX SYSTEM ARE CARRIED OUT AUTOMATICALLY AND HAVE NOTHING TO DO WITH THE ENTERED DILUTIONS 3 Select the number of images to be taken of the sample from the Images drop down list The default setting is for 20 images 20 images are equal to 10 manual measurements using the Neubauer hemocytometer The high amount of random checks allows for a small margin of deviation and minimizes measurement inaccuracies High cell density can reduce the number of cell images taken to 10 The maximum number of images that can be taken is 30 4 Choose the analysis process type from the Processtype drop down list only one option available at the moment i e Standard Should another process type be available then it can be selected from here 5 Click on the traffic light symbol to confirm the entries or press lt Return gt The traffic light symbol will then change to green When the traffic light symbol turns to green in the Process Parameter area the measurement procedure starts automatically by first checking the measurement chamber flowcell including the syringe The actual measurement begins after the respective message is confirmed User Manual Cedex Version
100. ment Offset window The measurement series offset can be manually shifted in this window Measurement Offset o 1 2 3 4 5 6 7 8 3 fo ma 5 Offset EA ihl tal th Cancel Ok Figure 25 The Measurement Offset window The offset can be separatelly entered for each of the five measurement series by clicking the button with the corresponding number of the measurement series Big time differences can be entered in days d small ones in hours h The selection is effected by the activation of the individual buttons h d The offset that is set causes an immediate shift of the measurement series in the diagram such that whether an appropriate position in the diagram was reached can be directly controlled The offset can be set by shifting the slider or by directly entering a numerical value in the Offset entry field 56 User Manual Cedex Version 1 6 1 The Analysis Results O The smallest time unit that can be set is 0 25 hours 15 Minutes The Ok button adopts values set in the graphics If the Cancel button is actuated the original offset values are re set The Details Button The Details button opens the Selection of Measurements window in order to view or change the grouping of the corresponding measurement series Grouping Measurement Series The Selection of Data Series Window The New or the Details button in the Data Series area opens the Selection of Data Series window Here measurement series can be grouped t
101. method 5 x 10 1x 107 cells per mL Material and Diameter of the capillaries Teflon 765 um 10 C 35 C 60 F 95 F Operating humidity 20 80 relative humidity non condensing Dimensions Height 660 mm 26 0 inches Width 300 mm 12 0 inches Depth 600 mm 23 6 inches Weight 27 0 kg 60 0 Ibs Energy requirements 100 250 VAC 50 60 Hz Energy consumption Automatic Sampler AS20 Number of samples Average measurement period for 20 samples Dimensions Height 170 mm 6 7 inches Width 175 mm 6 9 inches Depth 160 mm 6 3 inches Weight 2 6 kg 5 7 Ibs nergy requirements 12 VDC nergy consumption Power supply Input 100 240 VAC 47 62 Hz Output 12 VDC 2 0 A Cedex Computer Newest PC generation incl Windows 2000 Framegrabber card Network card 10 100 Mbit 10BaseT onboard Interface card 2 x serial D sub 9 1 x parallel CD ROM drive CD RW Maximum hard disk storage capacity 40 GB corresponds to 30 000 measurements Cedex Monitor 15 TFT Flatscreen Agency Approvals CE FCC A 36 Administrator Manual Cedex Version 1 6 1 Index A Access control uuueseeeeneeeeeeenee 13 29 Access rights Henn 14 16 Administrator 23 Administrator functions 21 Administrator password 14 Analysis data 24s een 16 Analysis software sen 13 AuditTrail ens
102. mple The average count of unstained viable cells per square 37 5 The total cell count viable dead cells per square 45 cells Calculate cell viability as follows 37 5 x 5x 10 viable cells per mL 1 875 x 10 viable cells per mL 45 x 5 x 10 total cell count per mL 2 250 x 10 total cell count per mL 1 875 x 10 viable cells per mL 2 250 x 10 total cell count per mL x 100 83 3 viability User Manual Cedex Version 1 6 1 15 A 4 1mm corner square y Figure 2 Diagram of a chamber of the Neubauer hemocytometer O counted cells uncounted cells Figure 3 Corner square containing 16 minor squares in a chamber of a Neubauer hemocytometer o lo O nn So o o fr Q zf ji 0 o SSS GS 0 Tole lo gt of Figure 4 Direction and strategy of cell counts 16 User Manual Cedex Version 1 6 1 Description of the Cedex System 3 3 Data Analysis with the Analysis Software The analysis software and its graphical user interface known as the Cedex Workbench is installed and supplied with the computer as part of the Cedex system The analysis software digitizes and analyzes the two dimensional images captured by the measurement system The analysis software recognizes structures identifies particles blends out background disorders and differentiates cells from cell debris and dirt particles Viable and dead cells are differentiated according to their degree
103. mptied regularly depending as the frequently of use Clean the container thoroughly and replace it remove lid so that the silicone tube and capillary tubes reach about 1 cm into the container Cleaning Routines Carried out by the Liquid Management The Liquid Management plays a key role in the Cedex system It is necessary in the beginning for a Prime and at the end for an LM ShutDown prior to shut down Click on the Liquid Management button on the Cedex Workbench to activate the procedure The Liquid Management is automatically activated after every sample measurement for the Ultra Fast Clean routine During the Ultra Fast Clean approx 30 seconds the entire liquid management system including capillary tubes measurement chamber flowcell and sample port are rinsed out thoroughly 80 User Manual Cedex Version 1 6 1 Maintenance of the Measuremment System o Besides the LM ShutDown and Prime routines the Liquid Management offers a range of different cleaning routines and prepares the syringe for removal Daily events LM ShutDown Prime 1 Reset Initialize Cleaning Fast Clean Standard Clean Intensive Clean Chamber Clean 1 Chamber Clean 2 Time to finish gt Cancel 00 00 min sec estimated Live image of the chamber Status System ready Figure 32 Liquid Management window The Liquid Management window is divided into the follo
104. ne other Cell Type is available besides the initial Cell Type Standard Cells see Adaptation of the Counting Routine on page 75 A new data set is available once the evaluation has been completed and all the changes can be retraced in the Audit Trail file and reviewed later see Viewing and Evaluating the Data Audit Trail File on page 69 The reprocess mode is activated by clicking on the calculator button u in the Data Source Identification area on the Cedex Workbench Follow the instructions below to reprocess existing data sets with changed analysis parameters 1 2 3 4 8 Click on the operating option u reprocess Select the appropriate reactor ID from the Reactor ID drop down list Select the appropriate sample ID from the Sample ID drop down list Click on the traffic light symbol to confirm Enter a name for the new data set on request reprocess 1 reprocess 2 etc Enter a new name for the data set the suggested name can be changed Should confirmed entries need to be changed again then click on the traffic light symbol to switch to red Make the necessary changes Select the required Cell Type in the Process Parameters area Start by clicking on the traffic light symbol in the Process Parameters area The next steps of the reprocessing procedure are the same as for a new measurement A new data set is available once the evaluation has been completed All the changes can be retraced in the Audit Trail
105. neeeennnenneeennen 32 B Beads solution sn 17 Blacklist 2 nen 30 Brightness n enden 17 Brightness histogram 18 Cc Calibrate the system 17 Calibrating nn 17 21 Calibrating solution eee 18 GD BUN Gi anni 28 Gell Type nuan nee ea 26 Checking brightness 18 Contact details eeaeee 35 Contents of the delivery 11 GOpYNINl ueennse anime 5 Counting routine enia 23 Cross sectional area nn 25 D Dead cells aggregate appearance 25 Dead cells aggregate maximum 25 Dead cells aggregate minimum 25 Dead cells appearance 25 Dead cells size nn 25 DONGI6 wicca arsenate eens 12 F FOCUS anna nenne et 17 18 Focus histogram se nennen 18 H Hardware components 12 I Installation serisi 11 L License agreement unenn 5 M Main power supply 13 Minimum pollution area 25 O OperatOn airera a E 23 P Password characteristics 31 Password management 29 Preparing test liquid 18 Printer 33 R Rights 222204s nn iiaa 5 S Safety instructions 0 ereen nenne 7 Standard C
106. neration incl Windows 2000 Framegrabber card Network card 10 100 Mbit 10BaseT onboard Interface card 2 x serial D sub 9 1 x parallel CD ROM drive CD RW Maximum hard disk storage capacity 40 GB corresponds to 30 000 measurements Cedex Monitor 15 TFT Flatscreen Agency Approvals CE FCC A 92 User Manual Cedex Version 1 6 1 Appendix Suppliers Cedex Detergent 3 Mucasol solution available from innovatis AG Germany 1 L Cedex Detergent solution for use Cat No 760 00 00 23 Mucasol concentrate available from Fisher Scientific Germany Mucasol Cat No 619 325 01 Omnilab Germany Mucasol Cat No 9 192 501 BrandTec USA Connecticut Mucasol Cat No 44003B Concentrated Mucasol is available in 2 liter bottles 5 liter containers and 60 liter barrels Sample cups available from innovatis AG Germany 1000 cups 4 mL Cat No 6 00 05 00 00 Fisher Scientific Germany 1000 cups 4 mL Cat No 9 401 531 Omnilab Germany 1000 cups 4 mL Cat No 9 401 531 IMPORTANT NOTICE The quality of these sample cups is controlled regularly by innovatis AG Sample cups with other catalogue numbers cause irreparable damage to the device Innovatis AG is not liable or responsible for any damage caused by the use of sample cups other than those specified above Trypan Blue available from Sigma Aldrich Germany 6 x 50 mL 0 4 Trypan Blue solution Cat No 93595 Cedex Calibration Beads to c
107. new user with identical access rights only the name of the existing user needs to be changed Follow the instructions below Click on the Users menu on the Cedex Workbench Click on Change name and the Select User to change list will appear Select the user profile to be changed Click Change and the Change user data area will appear Enter the user s present password in the Actual password field if applicable Enter the new username in the New user field Click on Change name and you will be requested to confirm the entry Oo NO F WN gt Click on Yes to confirm NOTE PLEASE PAY ATTENTION TO CHAPTER RE ISSUING USER NAMES PAGE 31 Deleting a User Password Follow the instructions below Click on the Users menu on the Cedex Workbench Click on Delete password and the Select User to delete password list will appear Select the appropriate user profile Click on Delete password and you will be requested to confirm the entry Click on Yes to confirm the entry and a message will confirm the entry Click on OK to shut the window oa F Wr gt Automatic Log out In version 1 50 and higher the Administrator can set a time limit after which a user is automatically logged out if the keyboard or the mouse button are not being used The log out also takes place during a current measurement The measurement however will thereby not be compromised In Cedex version 1 5 0 the default setting for Auto logout is 600 se
108. nge focus control to the PC 1 x Video cable to connect the measurement system to the PC 3 x Connecting cable 1 x Operating Instructions 1 x Administrator Manual 5 x Reagent container 250 mL each e 1 x 1000 mL Cedex Detergent 1000 x Sample vial 4 mL e Cedex Calibration Beads for the calibration of focus and brightness 1 x License agreement included in the operating instructions IMPORTANT NOTICE PLEASE NOTIFY THE DISTRIBUTOR IMMEDIATELY IF THE SERIAL NUMBER ON THE CEDEX ANALYZER AND THE COMPUTER DO NOT MATCH MATCHING SERIAL NUMBERS ARE NECESSARY AS THEY ARE ESSENTIAL CRITERIA IN THE CALCULATIONS OF THE CEDEX SOFTWARE NOTE A PRINTER IS NOT INCLUDED IN THE CEDEX SYSTEM FOLLOW THE INSTRUCTIONS TO INSTALL A PRINTER VERY CAREFULLY Administrator Manual Cedex Version 1 6 1 11 Choosing a Suitable Location for the Cedex System The Cedex system is highly sensitive and consists of top quality electronic and optical components To guarantee optimal and reliable performance of the Cedex system select the appropriate location according to the safety regulations described in the first section Connecting Hardware Components and Hardware Key Dongle See Figure 1 and Figure 2 Follow the instructions below to connect the hardware components Connect monitor keyboard and mouse to their respective ports on the PC Connect one end of the video cable to the Video port of the measurement system Connect the other end of the
109. nnanannnnnnnnn 69 Viewing and Evaluating the Data Audit Trail File uunnssnnnennnsnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnn 69 Editing Existing Analysis Results Data Sets unsuunsenssnnnnnnnnnnnnnnnanannnnnnnnnnnnnnanannnnnnnnn 70 Viewing Existing Analysis Results Modifying Measurement Parameters 70 Reprocessing Existing Data Sets 0u0444444nn4nnsnennnnnannnnnnnsnennnnnnnnnnnnnnnannnnnnn nn 71 GENERAL FUNCTIONS OF THE MEASUREMENT SYSTEM 73 Specifying Target Directories for the Analysis Data nsnssnresnnnsnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnn 73 Changing your own Password uussuersenssnannnnnnnnnnnnnnnannnnnnnnnnnnnnnnanmnnnnnnnnnnnnnanmnnnnnnnnnnnnnn anne 73 Viewing your own Access Rights uunsssnnneennnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnn namen 74 Data Entry Fields for the Data Source Identification Area 2222224000000nnnnnnnnannnnnnnnnnnn 74 Specifying your own Default Measurement Parameters uursunnssssnnsnnnnnnnnnnnnnnnannnnnn 74 ADMINISTRATOR FUNCTIONS sscssssstssssssscssnsssasnccsscntasnsssnsesdsasntasassenteansnance 75 Administration of User Profiles ccccccceseesseenee cece eeeeeeeeeeeeeeeeeeeaeeaneneeseeeseseeeaneeseseeeeeeeees 75 Adaptation of the Counting Routine unssssnennnnnnnennnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnennnnannennnnannnnn 75 Description of the Individual Parameters
110. nnn nennen 32 The Emergency Stop Bullon au ansehen 32 The Traffic Light Symbol 2 a caine ee nun 32 Description of Access Rights 4 4202444404 544442440040400040000000000000 ERARAS ANANA RANTE AA RAAR 33 Carrying out a Measurement zuursrresnnsnrnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnsnnnnnrnsnnnnnnnsnnnnnrnsnnnnnrnsnnn 33 Performing a PIME cererea ees active eed bed ne ee alee 33 Choosing the Operating Option and Specifying Sample Data cc eceeeeeeeeeeeeeneeeeeeeee 34 Enteringa Comment rue en ee reellen 35 Defining the Measurement and Analysis Process Parameters 224444 seen 35 Preparing the Sample and Attaching it to the Sample Port 4444s0nsnnnne nennen 36 TO Start Measuring u2 ee EE E 37 End of Measurement ae ea 38 Interrupting a Measurement uss4u444200nannnnnnnnnnnnnnnnannnnnnnnnnnnnnnannnnnnnnnnnnnnnnannnnnnnnn nen 38 Switching off the Cedex System unsnnnsnsnsnannnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnrnnn nenn nennen 39 Carrying out an LM ShutDown uuuurs2seesessnnnnenssnnnnnnnnnnnnnnsnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnn nn 39 THE ANALYSIS RESULTS 442222420000000000000000nnnnnnn nn nnnnnnn nenn nun 41 Description of the Analysis Results unensersesnnssennnnnnennnnnnnnnnnnnnnnnnnnnnnnnnnnnennennnnnnnnnn nennen 41 The Viable Cell Density Parameter 4sneeennnnnnnnnnnnnnn
111. nnnnnnnnnennnnnnnnnnnnnnnn nn 41 The Total Cell Density Parameter 4ssneeennnnnnnnnnnnnnnennnnnnnnnnnnnnnnnnnnnnnnennn nenn nn 41 The Viability Parameter u 44444 nenn nnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnn nn 41 The Total Cell Count Parameter 44nnnneenennnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnn nenn nnnn nenn 41 The Average Compactness Parameter usneennnnennennnnnnnnnnnnnnnnennnnnnnnnnnnnnnennnn nenn 42 The Average Diameter Parameter unnnneenennnnnnnnnnnnnnnnnnnnnnnnennnnnnnennn nenn onnn nenn 42 The Aggregate Rate Parameter snneeennnnennennnnnnnennnnnnnnnnnnnnnennnnnnnnnnnnnnsnennn nenn 42 The Standard Deviation Parameter seennnnennnennnnnnnennnnnnnennnnnnnennnnnsnnennn nennen 42 Viewing Individual Analysis Results after Completion of the Measurement Process 42 Graphical Display of the Analysis Results unsnssnsesnnnsnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnsn nennen 42 Partial View of Diagrams 4 ssnnnesennnnnnnennnnnnnennnnnnnnennnnnnennnnnnnennnnnnnnnnnnnnnnnnnnnnnnnn nn 43 The Aggregate Histogram 4444srsnnennnennnnnnnnnnnnnnnnnnnnnnnnnnnnnsnnnnnnnnnnnnnnnnsnennnnnsnnnnnnnsnnnnnn 44 The Measurement Chart 244usnsnneennnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnsnnnnnnnsnennnnnsnnnnn nn nn 46 The Compactness Histogram Chatt ssrsnneeennnnnnnnnnnnnnnnn
112. nnnnnnnnnnnnnnnennnnnnnnnnn nenn 47 The Diameter Histogram Chart 444snnnnneennnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnsnnnnn nenn 48 The Cultivation Time Chart 44ss444neeeennnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnsnnnnnnnsnnn nn 49 Areas in the Cultivation Time Chart Window usessennesnnnnnnnnnnnnennnnnnennnnnnn nenn 50 The Cultivation Time Chart Diagram 44444444 44H ne nnnnennnnnnnnnennnnnnnennnnnnn nn 50 The Data Selection Area aden debi apensstadageaest agents aha ersehen 51 The VIOW AlG ai oi 5 oon cals ncctassanocicaasacnacctteiensandasehesetsstancganeany carsancnedeaadacersattanses caadcertaaanndcaens 52 Saving a VIOW aresta a opcteeen pads AAS fees scyedies sabta ede sath edbbessdaded sath qeequtateedeevitag 52 LOAGING A VOW seess rna 53 Deleting VieW erase ua E S 54 The Genaral Noa ecn ierni on A 54 Printing User defined Results 00 0 cece ceeeeeeeeeeeeeeeeeeeeeeeeteeeaeeeseneaeeeseneaeeeseeeaeeeseeaeees 54 Exporting the Results 41 2 see ale erden 54 Entering a Comment 1 2244 ee nie aaa 55 Shielding the Cultivation Time Chart Window u sssssesssnnnnnnnnnnnnnnnnnnnnnnnnnnn nenn 55 The Extras Area nn era EEEE EEE 55 The Data Series Areas ae a aa Laien 55 The List of Measurement Series in the Data Series Area nennen 55 The New Button ee ee ee nern nn nertrte nern 55 User Manual Cedex Version 1 6 1 6
113. nstalling a local printer Follow the instructions below to install a printer Connect the printer to the PC according to the instructions in the printer manual Switch on the PC Press lt Ctrl gt lt Alt gt lt Delete gt after the booting process has ended BQ NS Login as administrator with the password Administrator or with your individual administrator password 5 Install the printer to the Windows 2000 Workstation according to the instructions in the Windows manual or printer manual 6 2 Installing a network printer The installation of a network printer to the Cedex computer is quite involved making it necessary for the network administrator to adjust the settings The system administrator has to set up the necessary settings in the network and in the Cedex computer Factory setting for the Cedex computer is Windows 2000 with local access for the following users 1 Administrator with the password Administrator 2 Cedex with the password Cedex 3 CedexSA with the password CedexSA For data security only the user Cedex and the user Administrator have access to the Windows desktop and therefore access to applications of Windows 2000 The user CedexSA is restricted to the Cedex application For set up of a network printer on the Cedex computer the System administrator has to deactivate the security package in advance following the instructions below Start the Cedex computer and login as Administrator to have access t
114. ntenance Instructions 4444ssrsnnnnnnnnnnnnnnnnnnnnnnnnnnn nennen 85 REMOVING the Syringe u nau kein pinnacles nied EEE eideeeene 85 Cleaning the Syringe isis aciiee eine ek ee nee ie 86 ithe Case ot Mild Soiling u nee ties AEAEE 86 inthe Case Of Heavy Solling use ee een 86 Assembling and Installing the Syringe usrnnsssnnnnnnnnennnnnnnnnnnnnnnennnnnnnennennnnnnnn nn 87 9 4 Maintenance of the Capillary Tubes ccccecesseeeeeeeeeeeeeeeenseeeeseenseeeseanseeeseeesesesenseeeeeeans 87 9 5 Maintenance of the Syringe Guide and Spindle uurssrrssnansennnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnn nn 87 9 6 Maintenance of the Measurement Chamber Flowcell Microscope Optic 8 Way Valve and the remaining Hardware COMPONENMS cccssseecceeseeeeeeeeeeeeeeeseeneeeeseeeeeenneseensens 89 APPENDIX nennen 91 Contact Detalls SPERBLPPFEBERFPPCHERFEEPERELEFFEPFEEERFPEFRERFPEPFEEFEFEPERERFFEPFREEEFELCHEREFEPFEREFEFPEREREFPPFFEEFFFESBEREERPFFREEEEEFFEEEER 91 Technical Specification S serie aaaea aa aaa Ea Aaaa Aaaa Naaa Eiaa 92 SUPPIIETS 2 se ee near 93 Material Safety Data Sheets for the Substances used unensnnnnnennnnnnnennnnnnnnnnnnnnnnnnnnnnnnnnnnnannnnn 93 Declaration Of Conformance 2 uses AENEA 94 INDEX ene ee en nn err ete ore er err rrr rere emer ee 95 6 User Manual Cedex Version 1 6 1 1 Rights Copyright Cedex Operating Instructions Copyright
115. o the desktop Open the Support folder Deactivate the Security Pack with a double click on MakeUnsecure Boo NS Shut all the windows on the screen and logout Administrator Manual Cedex Version 1 6 1 33 Cedex 5 Let the network administrator install the network printer as your standard printer for the local user Administrator Cedex and or CedexSA The respective user has to be logged in to Windows 2000 6 To activate the security pack the Cedex administrator has to be logged in to Windows 2000 Open the Support folder Activate the Security Pack with a double click on MakeSecure Start the Cedex system as usual after logging out After installation the printer the Cedex result data can be printed out by pressing the button Print in the Result Data area of the Cedex program see Cedex User Manual Printing the Analyses Results 34 Administrator Manual Cedex Version 1 6 1 Appendix Appendix Contact Details Information and technical support innovatis AG Meisenstr 96 33607 Bielefeld Germany Phone 49 0 521 2997 295 Fax 49 0 521 2997 285 email sales innovatis com support innovatis com validation innovatis com qa innovatis com internet www innovatis com Administrator Manual Cedex Version 1 6 1 35 Cedex Technical Specifications Cedex Measurement System Method of measurement Digital image and pattern recognition Viable dead cell differentiation Trypan Blue exclusion
116. of Trypan Blue staining The image processing is integrated in the Cedex Workbench in such a way that it allows the user or administrator to check on its function in the Cedex system constantly The analysis process cannot be altered during data input to allow for consistent and objective measurement results The Cedex system can register several thousand cells a minute depending on cell density The rapid processing of image data allows for a significant increase in random checks which greatly improves the reliability of the statistical accuracy compared to the manual count This analysis method provides information on cell viability and diameter form User Manual Cedex Version 1 6 1 17 18 User Manual Cedex Version 1 6 1 Installation and Operation ofthe Cedex System D 4 installation and Operation of the Cedex System Follow the instructions to install the hardware components of the measurement system and prepare the analysis software in the order given below Both actions should be done by the system administrator All instructions have to be followed in chronological order Detailed information is found in the Cedex Administrator Manual 4 1 Installing the Cedex System Checking the Contents of the Delivery IMPORTANT NOTICE PLEASE NOTIFY THE DISTRIBUTOR IMMEDIATELY IF ANY OF THE SHOCK WATCHES INDICATE IMPROPER HANDLING DURING TRANSPORT OTHERWISE THE GUARANTEE IS INVALID Ensure that all the items listed below have been del
117. ogether from the measurements at disposal 4 Selection of Data Series x Select Source Sample ID poo1 Workarea D Cedex lmages P003 POI2 P021 P004 P0I3 P022 P005 POI4 P023 Reactor ID KVcho0l Bode POTS P007 POI6 DataSet Poos Po1 7 P009 P018 gt Add selected measurements to list v v v v v v Selected Measurements Table contains source data of Data Series m kv 187 101 Original data Hs 10 2001 11 55 KV187 101 P002 Original data 17 10 2001 09 13 E Belt 7 KV187 101 P003 Original data 7 17 10 2001 16 12 KV187 101 P004 Original data 18 10 2001 08 41 M KV 187 101 P005 Original data x 18 10 2001 17 34 Action for selected entry KV187 101 P006 Original data 7 19 10 2001 08 42 ATA 7 KV187 101 P007 Original data 19 10 2001 11 47 KV187 101 P008 Original data 22 10 2001 10 01 _Remove Measurement s 7 KV187 101 P009 Original data 22 10 2001 17 02 KV187 101 POIO Original data 23 10 2001 14 08 AT 7 KV187 101 POI Original data 24 10 2001 13 58 KV187 101 P012 Original data 25 10 2001 11 11 7 KV187 101 P013 Original data 26 10 2001 10 32 KV187 101 PO14 Original data 29 10 2001 10 41 7 KV187 101 POIS Original data 30 10 2001 11 53 KV187 101 POI6 Original data z 31 10 2001 09 27 xl Cancel Ok Figure 26 The Selection of Data Series window User Manual Cedex Version 1 6 1 57 Cedex The Select Source Area In the Select Source area
118. on A progress bar indicates the time course during a loading process Data processing in progress Please wait Collecting data of all selected measurements Figure 22 The Progress bar There is a possibility of interrupting the loading process using the Cancel button If the current user does not have all access rights to the grouped data in a View a message calling his attention to this fact follows He or she can then choose to load only the remaining data or cancel the entire loading process altogether After the loading the data in the View the Data Series and the Diagram areas is updated User Manual Cedex Version 1 6 1 53 Deleting a View The Delete button opens the Delete a View window that is constructed like the Save a View window see Figure 20 page 52 The Save button in this window was replaced by the Delete button The selection of the View to be deleted occurs in the list or by entering the name in the entry row Deleting the data in the View is effected by the Delete button and confirming the safety challenge The window can be quit by using the Cancel button The General Area In General area printing and exporting data can be controlled and comments entered viewed The Hide button fades out the Cultivation Time Chart window Printing User defined Results The Print button opens the Print Culivation Time Chart window print window that allows printing out the diagram with the corresponding dat
119. on Preview to display a preview of the print or click on Cancel to shut the window and cancel a print 6 6 Viewing and Evaluating the Data Audit Trail File The Cedex software automatically creates a Audit Trail log file for each sample and records all relevant information relating to the sample allowing the measurement details to be reviewed and printed The Audit Trail file consists of at least the following details Manufacturer specifications on the device first 3 rows Measurement data and settings displays the date and time of measurements including the username as well as the Sample ID Reactor ID and parameters used for the sample analysis These entries correspond to data from the Sample Properties Data Source Identification and Process Parameters areas including a list of the Cell Type settings without any results User Manual Cedex Version 1 6 1 69 Cedex If the sample is modified or reprocessed the Audit Trail file will log these events in chronological order This allows all measurement data to be retraced including their modification reprocessing and to display the date time username and used parameters The Audit Trail file data can basically never be overwritten to ensure that all recorded analyzed and modified measurement data is documented NOTE SINCE THE AUDIT TRAIL IS SECURED TOGETHER WITH THE RESULT DATA DELETING THE RESULT DATA WILL RESULT IN A LOSS OF THE AUDIT TRAIL Click on Audit Trail in
120. on the x axis Figure 11 shows for example that 2 506 living and 666 dead cells of all identified cells respectively are available in aggregates build of two cells The percentages are displayed using the percent button 44 User Manual Cedex Version 1 6 1 The Analysis Results O z Aggregate Screen Aggregate Histogram B viable I dead 2 X 88 of total Cell Court SRD oa Figure 12 The Aggregate Histogram detailed view partitioned in living and dead cells here percentage distribution of the cells The percentage of cells of the Total Cell Count is displayed on the y axis The aggregate size in amount of cells per aggregate is given on the x axis Figure 12 shows for example that 21 6 of all living cells and 22 7 of all dead cells are available in aggregates build of two cells The Total Cell Count living and dead cells per aggregate is mapped in the histogram using the total button Figure 13 displays for example that 3 172 cells of the measurement are available in aggregates build of two cells E Aggregate Screen Aggregate Histogram EE Figure 13 The Aggregate Histogram total view sum of living and dead cells here number of cells The percentage of cells available in the aggregates in relation to the total cell count is displayed using the percent button User Manual Cedex Version 1 6 1 45 Aggregate Histogram oo S Geet rs vee ea i al Mel Riel E i ieee el ne
121. ons below to clean the syringe with a weak detergent 1 Fill a weak detergent Cedex Detergent in the syringe 2 Allow to stand for 30 minutes 3 Rinse the syringe at least 10 times with distilled or de ionized water Follow the instructions below to clean the syringe with an acid base combination Fill 0 1N NaOH in the syringe Allow to stand for at least 10 minutes Empty the syringe according to regulations Rinse the syringe with distilled or de ionized water Fill 0 1N HCl in the syringe Allow to stand for at least 10 minutes Empty the syringe according to regulations oN a F ON BS Rinse the syringe at least 10 times with distilled or de ionized water 86 User Manual Cedex Version 1 6 1 Maintenance of the Measuremment System D Assembling and Installing the Syringe Rinse a new syringe with 70 alcohol and then with distilled or de ionized water before installing it Follow the instructions below 1 Gently press the plunger into the syringe barrel 2 Insert the syringe back onto the syringe module 3 Secure the plunger to the syringe module with the screw at the bottom Ensure that the syringe barrel and plunger do not get jammed 4 Hold the syringe barrel by the Luer lock and turn it clockwise into the port of the 8 way valve 5 Should the Liquid Management window not have opened yet click on Liquid Management on the Cedex Workbench 6 Click on Reset Initialize in the Syringe handling area The
122. ouse pointer on the top left hand corner of the area to be enlarged 2 Hold the left mouse button pressed down and move it to the lower right hand corner of the area to be enlarged 3 Release the mouse button and the selected area will fill the complete window In order to display the diagram in its original size do the following 1 Place the mouse pointer anywhere on the diagram 2 Hold the left mouse button pressed down and form a box from the bottom right corner to the upper left corner 3 Release the mouse button The chart will then be in its original size Detailed Information Total Cell Density 22 34 x105 12 949 d Data Series Point 14 Figure 19 Detailed Information in the View area Information listed in the Detailed Information area e Symbol in the diagram corresponding to form and color Term of the selected evaluation Viable Cell Density Dead Cell Density Total Cell Density Avg Diameter Avg Compactness Viability e Value on the y axis and the appropriate unit e Value on the x axis and the appropriate unit Number of the measurement series in the Data Series table Number of the measurement Point in the Data Series table 50 User Manual Cedex Version 1 6 1 The Analysis Results O Variable axis calibration The cell density on the y axis of the diagram can be displayed in two variants Buttons 10 5 and 10 6 determine whether cell density in the unit 10 5 Cells mL or 1
123. plates for Excel By default export templates that take special features of respective Excel versions into account are put at the disposal of the user Administrator for various Excel variants User Manual Cedex Version 1 6 1 63 Cedex The Export Structure Area In this area individual data fields that are available in the Cultivation Time Chart can be selected for the export The left list Available Data shows all available data for the export Selected data from this list is transfered to the right list Export Data using Add Using Remove selected data from the Export Data list can again be removed Drag amp Drop can as well shift the data The Move up and Move down buttons move a selected entry in the Export Data list up or down respectively This has an effect on the structure sequence in the export file The following data fields are available for export Avg Compactness Avg Diameter um Comment Creator Data Series Number Data Set Date Time Dead Cell Density 10 5 Cells mL Dead Cell Density 10 6 Cells mL Dead Cells Device ID Dilution Group Identifier Operator Name Operator Parameter Reactor ID Sample ID Std Deviation 10 5 Cells mL Std Deviation 1046 Cells mL Total Cell Density 10 5 Cells mL Total Cell Density 106 Cells mL Total Cells Total Images Used Images Viability Viable Cell Density 10 5 Cells mL Viable Cell Density 10 6 Cells mL Viable Cells Workarea
124. position of the microscope stage The value displayed is named quality factor and based on the highest and lowest points of the brightness histogram 18 Administrator Manual Cedex Version 1 6 1 Installation and Operation of the Cedex System amp The calibration result is displayed in the window below Calibration results x Calibration completed Old brightness 73 New brightness 73 Old focus position 2343 New focus position 2343 Maximum focus path 9268 Figure 5 Calibration result The values of the previous calibration are also shown as a comparison Variations in brightness of 10 and of the focus position of 30 are allowed Please contact a service technician from innovatis AG if variations between the old and the new values remain constantly high A greater variation in brightness is a sign that the lamp s life cycle is nearing its end 6 Click on OK to confirm the result T Click on X to shut the window The analysis system can now start measurements Shut down the Cedex system if no measurements are to follow NOTE TO SHUT DOWN THE SYSTEM RESULTS IN AN AUTOMATIC BACK UP THE CURRENT CEDEX INI AND CEDEXST INI FILES FROM THE C CEDEX DIRECTORY IN THE D CEDEX DIRECTORY THE RESTARTED CEDEX PROGRAM APPLIES THE TEMPORALLY MORE UP TO DATE VERSIONS OF THE CEDEX INI AND CEDEXST INI Administrator Manual Cedex Version 1 6 1 19 20 Administrator Manual Cedex Version 1 6 1 General Administrator Fun
125. program The Windows administrator determines which of the two Cedex accounts will be used The account that should not be used must be disabled in the Cedex ini file To provide the CedexSA user with the ability to print or to use a network drive it is necessary to allow access to the desktop the Option Make Unsecure in the Support folder gives the user this access After he access is provided the rights have to be reset by the option Make Secure in the Support folder The security functions of the new password management for the Cedex program can only be administered over access to the desktop because entries for the Cedex ini are hereby undertaken Access to the desktop is allowed only for the Windows administrator Operating System Administrator or for the Cedex user If a Cedex or CedexSA user registers at the operating system level the default passwords are Cedex and CedexSA respectively 5 2 Access Control at the Cedex Level For the Cedex program only the Cedex administrator is installed per default no default password installed For security reasons the Cedex administrator should secure his her access to the Cedex program with a password The Cedex administrator installs the Cedex user and gives him her the necessary rights see Cedex User Manual in addition to the above mentioned Administrator Manual Cedex Version 1 6 1 29 Cedex 5 3 Administration of the New Password Management The new functions for the password
126. r XX 11 59 PM Sample ID Avg Diameter um B Total Cells Comment v Ex 31 12 2001 23 59 Total Images Remove Creator p R g p Used Images 4 Dilution Decimal Point String Terminators 4 Viable Cells Operator Parameter C Comma C Single Quote Workarea Data Series Number Dot Double Quote Dataset X C No Terminator Figure 27 The Export window The Output File Name Area In the entry field of the Output File Name area the path and the name under which the export file should be stored are entered The button El on the right of the entry field opens a window in which path or file can be directly selected The Output Format Area Here format for the export file can be selected DIF e g for Excel or CSV generally usable The files automatically and unchangeably receive dif or csv file name extensions The Data Series Mixing Area The distribution or structure of the measurement series for the export file are selected in this area Single file for each data series is used to create a file for each measurement series The file name is thereby automatically supplemented with the number of the current measurement series directly in front of the file name extension Common files for each data series is used to store all the measurement series in a common file Besides how the list should be structured can be additionally chosen 60 User Manual Cedex Version 1 6 1 The Analysis Results O One complet
127. ra Eredar cone 30 The File Menu 0442444444400nnnnnnnnnnnnnnnannnnnnnnnnnnnnannnnnnnnnnnnannnnnnennnnnnsnnnnnnnrennnnnnannn 30 The Specials Menu 4440ss4444444nnnnnanennnnnnnennnnnnannnnnnnnnnnnnanennnnennnennennnannnnr rennen 30 The Setup Menu 42444444snannnnnnnennnnnnnnennnnnnnennnnnnennnnnnnnnnnnnanennnnnrnnennennnennnnrrnnnn nn 31 The Users Menu 2444444404H0nnnnnnnennennnnennnnnnnnnnennnnennnnnrnnnnnnnnnnnnnernnnnesnnnennnnr rn namen 31 The Help Menu 44444440004H44Hnnnnnnanannnnnnnnnnnnnnannnnnnnnnnnnannnnnnennnnnnsnnnnnnnrnn nennen 31 The User Identification area 4424444444H0nnnnnnnnnnennnannnnnnnnennnnnnnannnnnnnennennnennnnrnnnennenn 31 The Data Source Identification area ss444444444nnnnnenennnnnnnnnnnnnennnnnnnennnnnnnnannnnnnnennenn 31 User Manual Cedex Version 1 6 1 3 Cedex 5 5 5 6 5 7 5 8 6 2 6 3 The Microscope Symbol puerta a weet atte yeeee tea eed 31 The Floppy Disk Symbol na la addon 31 The Calculator Symbol nedisan iai R aan 31 The Sample Properties Area 4444444400unnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnennnnnnnennnnnnnnnnnn nn 32 The Process Parameters Area 2a lad aha 32 The Cell mage Area nee nennen needs elec 32 The Result Data Area ae eds 32 The Liquid Management Button 444s4ssnneeennnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnennnnnnnennnnnnne
128. reprocessed data is possible Click on the traffic light symbol to confirm the entries Should confirmed entries need to be changed again then click on the traffic light symbol to switch to red Make the appropriate modifications Should a dilution be corrected or a cell image excluded or included in the evaluation then the traffic light symbol in the Process Parameters area will switch to red Click on the traffic light symbol to confirm Click on Yes to confirm the changes and an appropriate message will appear Click on No to delete all changes and return to the previous setting Click on OK to shut the window All changes can be retraced in the Audit Trail file and reviewed later see Viewing and Evaluating the Data Audit Trail File on page 69 Reprocessing Existing Data Sets Reprocessing existing measurement results data sets i e viewing a set of recorded image data and evaluating them with new analysis parameters can be done in the reprocess mode which is only accessible to authorized system users User Manual Cedex Version 1 6 1 71 Cedex The reprocess mode offers the following options Enter modify or delete a comment see Entering a Comment on page 35 Correction of the dilution see Defining the Measurement and Analysis Process Parameters on page 35 Change the Cell Type see Defining the Measurement and Analysis Process Parameters on page 35 The Cell Type can only be changed if at least o
129. rmed by authorized service technicians Always ensure that there are sufficient reagents in the reagent containers e Empty the waste container often to prevent it from overflowing Always observe the safety regulations when handling Trypan Blue detergent ethanol or acids and bases e Waste contaminated with tissue cells e g sample cups pipette tips etc must be disposed of according to the recommended rules for handling bio hazard materials Any leaks in the liquid management system must be repaired immediately Contact your distributor Always ensure that there is a sample cups attached to the sample port e Only use Cedex Detergent or a detergent with identical specifications for the cleaning process in the liquid management system see Suppliers on page 93 Only use sample cups from suppliers specified on page 93 for the conveyance of cell samples to the sample port Other sample cups with other catalogue numbers can cause irreparable damage to the Cedex e Always wear protective clothing when handling samples particularly gloves To prevent damage to the measurement chamber flowcell no particle larger than 80 um should enter the measurement system Do not use Microcarriers for this device Ensure that all utensils required for sample preparation e g sample cups pipette tips etc are stored in a dust free environment to guarantee precise measurement results Ensure that the Trypan Blue sol
130. s as described below Sample Data Access rights to sample data create view change delete Reprocess Data Access rights to recalculate recorded measurements create view change delete allow changing work area Access rights to change the target directory Cell Types Access rights to evaluate Cell Types create view change delete Users Access rights to administer user profiles create view change delete System Access rights to specific system functions shutting down the Cedex system shutting down the Liquid Management changing the syringe use of special functions 5 6 Carrying out a Measurement Carry out measurements only after having followed the instructions for Starting the Cedex System on page 27 IMPORTANT NOTICE AFTER STARTING THE CEDEX SYSTEM ALWAYS PERFORM A PRIME PRIOR TO ANY MEASUREMENTS A number of relevant parameters and entries must be specified on the Cedex Workbench prior to any measurements They are as follows e Choosing the operating option and specifying sample data e Entering a comment optional Defining the measurement and analysis process parameters e Preparing the sample and attaching it to the sample port Only when the steps above have been completed can the measurement procedure begin Performing a Prime The capillary tubes and syringe are filled with air prior to each start of the Cedex system or after its installation Performing a Prime r
131. s to be retraced see Viewing and Evaluating the Data Audit Trail File on page 69 The Cell Image Area The actual captured or analyzed cell image of a sample is displayed here tas This button shows the total amount of images in one view see Viewing and Selecting may Cell Images page 65 The Result Data Area The current measurement and analysis data are displayed in this area see The Analysis Results on page 41 The Liquid Management Button The maintenance and cleaning options in the Liquid Management are displayed It is also possible to perform a Prime or to display the actual image of the measurement chamber flowcell The Emergency Stop Button The measurement procedure can be interrupted in case of emergency and be resumed later IMPORTANT NOTICE ONLY USE THE EMERGENCY STOP BUTTON IN A CASE OF EMERGENCY BECAUSE THE ACTUAL MEASUREMENT DATA CAN OTHERWISE BE LOST RESULTING IN THE CEDEX PROGRAM MAYBE HAVING TO BE COMPLETELY RESTARTED The Traffic Light Symbol Clicking on the traffic light symbol confirms any entries made and only switches to green when all the necessary data are entered Entries can also be confirmed with lt Return gt Should confirmed entries need to be modified again click on the traffic light symbol to switch it back to red 32 User Manual Cedex Version 1 6 1 Working with the Analysis System 5 5 5 Description of Access Rights Each user profile can have specific access right
132. s44444244444HBHnennnnnnnnnnnnnennnnnnnnnnnnnnnnannnnnnnennenn 14 3 3 Data Analysis with the Analysis Software ussnnssnnssnnnsnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnn 17 4 INSTALLATION AND OPERATION OF THE CEDEX SYSTEM 19 4 1 Installing the Cedex System nunsnnnsennennnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnn nn 19 Checking the Contents of the Delivery 40su424440ennnsnnnennnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnn ana 19 Choosing a Suitable Location for the Cedex System u 22444ssssnnnennnnnnnnnnnnnnnnnnnnnn nenn 20 Connecting Hardware Components and Hardware Key Dongle een 20 Connecting the Cedex System to the Main Power Supply uussssnsnensssnnnnnnnsnnnnnnnnnnnnnnn nn 21 Preparing the Measurement System 0 cccceeceeeeee enn ee ee eeneeeee tees eeecaeeeeeeeaeeeseeaeeeeeenaeeeeseaas 22 Cleaning the Reagent Containers s esssessssssseennannsnnnatennannannantennaanananaatennaanannaatennaan annaa 22 Filling the Containers and Attaching the Sample Cup to the Sample Port 22 4 2 Preparation and Configuration of the Analysis Software by the Cedex Administrator 23 5 WORKING WITH THE ANALYSIS SYSTEM cccceeeeeeeeeeeeeeeeeeeeeeeees 25 5 1 Analysis System General Instructions nnsssnnssnnennnnennnnnnnnnnnnnnnnnnnnnnnnnnnnennnnnnnnnnnnnnnennnen 25 Switching on off the Cedex Sys
133. see may e only use the software on the PC supplied with the Cedex system only produce copies of the software for internal archiving and backup purposes i e on condition that all property and copyright notices shall be included on the copies only transfer the software on a permanent basis excluding leasing and renting in conjunction with a transfer of the entire Cedex system and on condition that the licensee retains no copies and the recipient accepts to be bound by the terms of this agreement The Licensee may not e use the software on more than one computer workstation or file server in a network or multiple user system except if an additional license for every additional computer or workstation is purchased from innovatis AG modify copy adapt reverse engineer decompile disassemble or create derivative works of the software or related documentation excluding a backup copy lease or rent the rights on the software and related documentation in any form whatsoever to a third party remove property and copyright notices and marks in or on the software documentation containers and devices All rights claims use and intellectual property rights relating to the software documentation and any copies made by the licensee remain with innovatis AG Unauthorized copying of the software or documentation or failure to comply with the limitations outlined above shall automatically terminate the license 6 Admini
134. sibilities for the Cedex usage The factory setting of the Cedex program has established a Cedex administrator no password in the factory setting The Cedex administrator is advised to secure his access by a password for reasons of safety Cedex user and their necessary rights are customized by the Cedex administrator Administrator Manual Cedex Version 1 6 1 9 10 Administrator Manual Cedex Version 1 6 1 Installation and Operation of the Cedex System amp 3 Installation and Operation of the Cedex System Follow the instructions to install the hardware components of the measurement system and prepare the analysis software in the order given below 3 1 Installing the Cedex System NOTE THE SOFTWARE AND HARDWARE VERSION OF YOUR CEDEX ANALYSIS SYSTEM CAN DIFFER FROM THE INFORMATION DESCRIBED HERE BUT SHOULD NOT MAKE A DIFFERENCE TO THE OPERATIONAL FUNCTIONS IN GENERAL Checking the Contents of the Delivery IMPORTANT NOTICE PLEASE NOTIFY THE DISTRIBUTOR IMMEDIATELY IF ANY OF THE SHOCK WATCHES INDICATE IMPROPER HANDLING DURING TRANSPORT OTHERWISE THE GUARANTEE IS INVALID Ensure that all the items listed below have been delivered e 1 x Measurement system 1x PC incl manual Windows 2000 and installed Cedex software and CD burner e 1 x Keyboard 1 x Mouse 1 x Mouse pad 1 x Monitor incl manual e 1 x Hardware key Dongle Cords 1 x VGA cord not attached to the monitor 2 x Serial cord to connect syri
135. stem on page 79 and The Logout on page 26 The sample cup of the last measurement should remain attached to the sample port during the complete cleaning procedure and only be removed for the next measurement When the Cedex system is to be shut down after a set of measurements or at the end of the day then the complete liquid management system should be cleaned with an LM ShutDown procedure to maximize the life span of the Cedex system and guarantee optimal performance The entire liquid management system is rinsed with distilled or deionized water during the LM ShutDown which is then pumped out see Carrying out an LM ShutDown on page 39 5 7 Interrupting a Measurement The EMERGENCY STOP button found at the bottom left corner of the Cedex Workbench can be activated during the measuring procedure to interrupt the analysis process Measurements can resume by clicking on the Run button This allows the user to react immediately in an emergency e g refilling reagents or emptying the waste container IMPORTANT NOTICE ONLY USE THE EMERGENCY STOP BUTTON IN A CASE OF EMERGENCY BECAUSE THE ACTUAL MEASUREMENT DATA CAN OTHERWISE BE LOST RESULTING IN THE CEDEX PROGRAM MAYBE HAVING TO BE COMPLETELY RESTARTED 38 User Manual Cedex Version 1 6 1 Working with the Analysis System 5 5 8 Switching off the Cedex System Switching off the Cedex system at the end of the day or for a longer pause should always be carried out in this order
136. strator Manual Cedex Version 1 6 1 General Administrator Functions O Dead Cells Size Range 40 250 Default 110 Unit um Defines the minimum cross sectional area of a particle in um Increasing this value means that less particles will be recognized as dead cells Dead Cells Appearance Range 10 10 Default 0 Unit absolute measure Dead cells have a variety of appearances under the microscope This can range from sharply outlined small dark dots to less distinct objects with varying brightness and contrast This parameter distinguishes large dark objects which are typical dead cells from less distinct objects Increasing this value means that there are more deviations from typical dead cells resulting in a higher count of dead cells Dead Cells Aggregate Minimum Range 250 1500 Default 360 Unit um Defines the minimum cross sectional area of a dead cell aggregate cluster When this size is reached it is then automatically partitioned into single cells Dead Cells Aggregate Maximum Range 1500 5000 Default 2250 Unit um Defines the maximum cross sectional area of a dead cell aggregate cluster The dead cell aggregate area within this range is automatically partitioned into single cells Any aggregate larger than the specified maximum area is not taken into account Dead Cells Aggregate Appearance Range 10 10 Default 0 Unit absolute measure Defines the automatic
137. strator Manual Cedex Version 1 6 1 General Provisions The licensee may terminate this license at any time by destroying all copies of the software or returning all copies to innovatis AG Innovatis AG may terminate this license if the licensee fails to comply with all terms and conditions of this license agreement In such an event the licensee agrees to destroy or return all copies of the software The licensee further agrees to be responsible for all taxes resulting from this license Any unauthorized leasing renting or transferring except as outlined above is illegal If any provisions of this license agreement are invalid for whatever reason this shall have no effect on the remaining terms hereof The license agreement is governed by the laws of the Federal Republic of Germany This is the only agreement between innovatis AG and the licensee and replaces any other negotiations or prior agreements relating to the software and documentation Cedex Reg U S Pat amp TM Off FCC Notification WARNING CHANGES OR MODIFICATIONS TO THIS UNIT NOT EXPRESSLY APPROVED BY THE PARTY RESPONSIBLE FOR COMPLIANCE MAY VOID THE USER S AUTHORITY TO OPERATE THE EQUIPMENT Note This equipment has been tested and found to comply with the limits for a Class A digital device pursuant to Part 15 of the FCC Rules These limits are designed to provide reasonable protection against harmful interference when the equipment is operated in a commercial en
138. system User Manual Cedex Version 1 6 1 79 Cedex 9 2 Daily Cleaning and Maintenance The following cleaning and maintenance routines should be carried out on a regular basis or whenever necessary Checking the Reagent Containers Volume Always check that the reagent containers are sufficiently filled with reagents to prevent the syringe and 8 way valve being used under dry conditions and getting damaged which would lead to incorrect measurement results Check the volume in the individual reagent containers and refill if necessary prior to each measurement see Filling the Containers and Attaching the Sample Cup to the Sample Port on page 22 Cleaning All reagent containers must be cleaned thoroughly with distilled or de ionized water when the measurement system is used for the first time and frequently recommended once a week during routine operations Avoid reagents being contaminated with dust fluff particles and especially micro organisms in the case of the Trypan Blue solution because this can lead to false measurement results If the Trypan Blue solution is contaminated intense areas that make image analysis difficult or even impossible can be visible in the image If not in use keep the Trypan Blue solution in a refrigerator A replacement of Trypan Blue after 5 days is recommended Detailed information see Cleaning the Reagent Containers on page 22 Emptying the Waste Container The waste container should be e
139. t 49 D Daily events area 40 nennen 81 Data analysis 42244 nennen 17 Data entry fields u4 eeeeeeeeeeee 74 Data save nennen 78 Data Series nennen 55 Data Series Mixing ne 60 Data sets nennen 70 Data source identification 31 Data source identification area 74 Dead cells aggregate appearance 77 Dead cells aggregate maximum 77 Dead cells aggregate minimum 77 Dead cells appearance uun 77 Dead Cells size nen 77 Declaration of Conformance 94 Default measurement parameters 74 Defaults a een 74 Deleting a View 54 Detalled u un anne 45 Detailed Information 50 Details button u0ss2222 een 57 Detergent ee nahen 22 Diameter nu nennen 36 Diameter histogram 48 Dilution isis telnet neen 35 Dongle en nenne 20 E Emergency Stop i 32 38 84 87 Ethanol ae 23 Excel au anne 65 Excluded images 4 nenne 68 EXPO Mi ni nn 54 Export Structure ueeeeeeneeneee 61 64 Export Template nenn 61 Export Window cecceeeeeeeeeeeenaeees 59 Export Start recaian 65 Exporting the data 54 5
140. tecsnsneceiss 29 5 1 Access Control at the Operating System Level ununnnensesnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnn 29 5 2 Access Control at the Cedex Level unnuunsnsnsnnnennnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnannnnnn 29 5 3 Administration of the New Password Management nunnensennnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnn 30 The Gedex Adiministr at r n en eat 30 The Blacklist cerci e alien leeren 30 Specifying Password CharacteristieS enesssrcimrrriiesiinirrriiiiinri rinn Anra aA 31 Re lssuing User Names sncseriscinironesiiiiiii ae nul EE 31 5 4 The System AuditTrail File sssrin aaaea ENA AEAEE RAKNA NAAA 32 Administrator Manual Cedex Version 1 6 1 3 6 INSTALLING A PRINTER ec nn 33 6 1 Installing a local printer uennesnnnenneennnennennnnennnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnennnnnn nenn 33 6 2 Installing a network printer uunnsennnesnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnn nenn 33 APPENDIX nenne ee 35 Contact Detalls RE RN ASRHRREEIEE INFOR R R 35 Technical Specifications 2H sc Nee ER 36 INDEX YRRRNERRNERENEEERE EEE EEEEEEIREIEE EDEN ER eira incer aineeton painei eguiii 37 4 Administrator Manual Cedex Version 1 6 1 1 Rights Copyright Cedex Manuals and Operating Instructions Copyright 2003 innovatis AG All rights reserved Operating instructions and manuals are protected by copyright No part of this document may be reproduced
141. tem unsessssnnsnnnnnnnnensnnnnnnnnnnnnnnonnnnnnnnnnnnnn namen nn 25 Starting the Analysis Software ccc creer nee ene e eerie ee erties erties ee teeeee teaser nennen nn 26 5 2 General instructions on working with the Cedex Workbench j cseccseseeeeeeseeeeeenneeneeens 26 TSS OU Ar i E E E aa oh pn ead yan eet reals He na ohana es 26 2 U 01 21 OUT seats ected E ade aa Pa cn pa eee eee ade Peeper ee 26 Automatic Log OUt uusrsnseeensnnnnnennnnnnnennnnnnnennnnnnnennnnnnnnennnnnnnennnnnnnnnnnnnnnnnnnnnnnnnennnnnnnnrsn ann 26 To Shut Down the Cedex Workbench s444444444444BHonennnnnennnnnnnennnnnnnnnnnnnnnnannnnnnnennen 27 5 3 Starting the Cedex System nnesnnssnnennnnennnnnnnnnnnnnnnennnnnnnennnnnnnnnnnnsnnnnnnnsnnnnnnnnennennnnennenn nennen 27 Switching on the Cedex System uuuessnneeesssnnnnnnensnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnn nn 27 Check the Measurement System and have Samples prepared 4ssennnnen 28 To Start the Cedex Workbench 444440osnsssnnnnennnnnennnnnnnnnnnnnnnannnnnnnennennnennnnnnn nennen 28 The Loader Program 4444nsnneennnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnennnnnnnennnnnsnennnnnsnnnnnnnsnnnnnnnnsnnnnn 29 5 4 Description of the Cedex Workbench 4424440000nnnnnnnnnnnnannnnnnnnnnnnnnannnnnnnnnnnnnnnnnn 30 Men Barci ernie een ana Laer coed En cote da coded sah Eee Cade easy mata
142. ter installation are not observed Software License Agreement Grant of License This is a license agreement between you the purchaser or licensee of the Cedex system and innovatis AG Innovatis AG grants the licensee the following license on condition that the licensee accepts all provisions of this agreement The licensee agrees to be bound by the terms of this agreement once commissioning or starting the Cedex system Administrator Manual Cedex Version 1 6 1 5 Cedex The Cedex software is based on the following e application software to control the hardware components analysis module for processing registered images and data analysis e data base module to archive and process obtained data graphical user interface for the complete software all technical descriptions and instructions on the use of the software innovatis AG grants the licensee a non transferable non exclusive sublicense except as outlined below to use a copy of the software and documentation belonging to the Cedex system in compliance with the terms of this agreement Any property and commercial protection rights regarding the software product shall remain with innovatis AG The software is only licensed not sold No property rights regarding the software product shall be transferred to the licensee The software product must be treated as any other material protected by copyright However this shall not apply to the following The Licen
143. ters Description of the Individual Parameters Living Cells Appearance Range 20 20 Default 0 Unit absolute measure Defines the sharpness and clarity of viable cells Increasing this value means that more cells will be determined as viable cells This parameter should be handled with care because it influences the determination of the cell density and the viability The following text describes a situation where a change of this parameter is necessary A cell line should be analyzed where dead cells absorb the Trypan Blue badly If the default parameter setting is taken for an analysis some of the dead cells will be recognized as living cells When the parameter Living Cells Appearance is changed towards negative values in special situations towards values lt 10 the darker objects are more and more recognized as dead cells Thus the measured viability decreases If the parameter is changed towards positive values the amount of objects counted as living cells will increase Thus the measured viability increases A change towards positive values could make sense if due to the condition of the cells or he process media the optical appearance has a deficient contrast ATTENTION In standard situations a change of this parameter results in labeling of debris objects In the range 10 10 the Std Cells 5 00 operator behaves similar to the Std Cells 3 0 operator 76 User Manual Cedex Version 1 6 1 Maintenance of th
144. the Process Parameters area to gain access to the Audit Trail file which is also available in the archive and reprocess mode The Audit Trail file is displayed when the floppy disk symbol is activated after the completion of a measurement It is available in the archive mode when the data set is recorded and the traffic light symbol is green in the Data Source Identification area It is available in the reprocess mode when a new data name is set and the traffic light symbol in the Data Source Identification area is green and a new name for the reprocessed data is specified NOTE PLEASE MAKE SURE THAT THE RESULT DATA AND THE DIRECTORY IS WRITABLE BY THE APPLICATION A READ ONLY SITUATION CAN OCCUR WHEN COPYING BACK SAVED RESULTS FROM CD OR NETWORK DRIVE IN THIS CASE A WARNING BOX WILL INFORM YOU THAT THE FILES ARE NOT WRITABLE SELECTING OK WILL LET YOU PROCEED WITHOUT UPDATING THE AUDIT TRAIL 6 7 Editing Existing Analysis Results Data Sets Besides the analysis of measurements the Cedex system also allows existing measurement results or data sets to be reprocessed and modify These changes are done on existing software data image and analysis results and therefore do not require any adjustments to the measurement system There are two options possible when making changes to the analysis results Viewing and possible correction of evaluated measurements and their corresponding results with the floppy disk symbol or reprocessing existin
145. the zero point of single measurement series to each other A window in which the data type that should be used is selected appears The following are for selection Viable Cell Density Dead Cell Density Total Cell Density By means of this function an automatic shifting of the zero point takes place on the x axis in order to be able to compare the measurement series to each other better The lowest common value of the measurement series is hereby automatically determined and the graphs in the diagram shifted to each other such that it falls in on the x axis in one point In doing so the leftmost point of the graph of all the measurement points is set at value 0 See also Measurement Series Offset Page 56 The Data Series Area The List of Measurement Series in the Data Series Area In the Data Series area general data of the measurement series that were selected for the display in the diagram is listed down in summary Up to five measurement series can be grouped together In the fields Reactor ID Sample ID Start and Start Date Time data from the first measurement of each measurement series is used and in the field Sample ID End data from the last measurement of each measurement series The New Button The New button opens the Selection of Data Series window in which new measurement series can be grouped see Grouping Measurement Series page 57 If measurements are already in the Data Series list the Sec
146. tically carries out a system check up Among others ports and drivers of various Cedex system components are thereby checked Components that are full proof are displayed as green in a list see Figure 7 The Loader Program closes automatically if no errors were discovered Figure 7 System check If the necessary components are not available or not correctly installed a corresponding error message appears In this case please contact your service technician The program can be quit by using the Exit Button User Manual Cedex Version 1 6 1 29 Cedex 5 4 Description of the Cedex Workbench The Cedex Workbench is divided into the following areas and elements Menu bar User Identification area e Data Source Identification area e Sample Properties area Process Parameters area Cell Image area e Result Data area Liquid Management button Emergency Stop button A description of the individual areas and elements is given below Menu Bar The menu bar at the top of the Cedex Workbench has three to five menu items depending on the user i e a normal user or administrator e File menu e Specials menu Setup menu Users menu e Help menu The File Menu The menu items have the following functions Logout To logout of the Workbench Shut down To shut down the Workbench Choose Workarea Choosing a target directories to save the analysis data It is only displayed if the user has been de
147. tions then this could indicate that the measurement chamber is soiled A special Intensive Clean is required twice a month for a thorough cleaning Therefore a solution of 0 1 1 N HCL is used instead of detergent Follow the instructions below 1 Remove the reagent container and replace it with an appropriate container with a 0 1 1 N HCL solution taking care that the capillary tube reaches far enough into the solution Start Prime Start Intensiv Clean after Prime has ended 82 User Manual Cedex Version 1 6 1 Maintenance of the Measuremment System D 4 Replace the HCL container with the detergent container at the end of the Intensive Clean process Start Prime again Start Standard Clean after Prime has ended The Chamber Clean 1 and Chamber Clean 2 cleaning routines are only required for the measurement chamber flowcell Chamber Clean 1 with water for approx 10 seconds The measurement chamber flowcell is rinsed at high pressure with distilled or de ionized water being drawn in through the syringe Use this routine to remove resistant protein aggregates fluff particles or other obstructions from the measurement chamber Chamber Clean 2 with water detergent and air for approx 30 seconds This routine intensifies the Chamber Clean 1 procedure Once the measurement chamber flowcell is rinsed with water detergent the remaining liquid in the capillary tubes in between the syringe and valve head is drawn t
148. to individual cell images To view results relating to individual cell images see Viewing Individual Analysis Results after Completion of the Measurement Process on page 42 mi ompactness Screen Figure 16 Compactness Histogram The ratio between the cell circumference and the cell area is based on an ideal sphere Ar which is equivalent to the value gt 1 lt on the abscissa of the histogram The extent of variation between the cell shapes and a sphere is reflected in the values being greater than 1 Ifa significant number of cells ordinate have a value higher than 1 then this could indicate improper handling of a sample Check the state of the cell culture i e if the stirring speed is too high or nutrient composition is incorrect User Manual Cedex Version 1 6 1 47 The Diameter Histogram Chart Click on the Diameter Histogram button to display a diagram of viable cell diameters based on the Average Diameter value of all images in the Overall column 1 Diameter Screen Cell Diameter Histogram Number of Cells a8 88853558 Figure 17 Diameter Histogram The Diameter Histogram provides three sliders to define separate diameter intervals This allows you to divide the Diameter Histogram into maximum four parts The sliders can be activated by clicking on the corresponding boxes and moved by sliding them You can also define the margins of the interval by entering the numerical values on the left hand s
149. ult PasswordMinNum X Passwords do not require a minimum number of digits Passwords must contain at least X digits PasswordReuse 0 default PasswordReuse X Passwords already used can immediately be reused Passwords already used can only be reused after X used different passwords entry of 1 to 25 possible NOTE THE CEDEX ADMINISTRATOR CAN ENTER INCORRECT PASSWORDS WITHOUT LIMITS AND AN EXTENDET TIME PERIOD AFTER RETRY OF ADDITIONAL 10 SEC EACH TIME Re Issuing User Names The following settings determine whether reuse of user names already deleted is permissible ReuseUsernames 1 default ReuseUsernames 0 Deleted user names can be re issued Deleted user names cannot be re issued Administrator Manual Cedex Version 1 6 1 31 Cedex 5 4 The System AuditTrail File All important actions carried out by the Cedex program are recorded in the AuditTrail_ lt year gt _ lt month gt log file found in the Cedex folder The file is drafted every month and indicates the year and the month in which these actions were carried out Example of listed actions Date Time ID User Action characters 03 06 2003 14 27 50 lt O gt system System started 03 06 2003 14 27 51 lt I gt ID of Security device 1896413025 03 06 2003 14 27 51 lt I gt DevicelD G007D1018 03 06 2003 14 27 51 lt I gt system System type Cedex Standard 03 06 2003 14 27 5
150. ure 6 Setting Cell Type Standard Cells parameters Description of the Individual Parameters Living Cells Appearance Range 20 20 Default 0 Unit absolute measure Defines the sharpness and clarity of viable cells Increasing this value means that more cells will be determined as viable cells This parameter should be handled with care because it influences the determination of the cell density and the viability The following text describes a situation where a change of this parameter is necessary A cell line should be analyzed where dead cells absorb the Trypan Blue badly If the default parameter setting is taken for an analysis some of the dead cells will be recognized as living cells When the parameter Living Cells Appearance is changed towards negative values in special situations towards values lt 10 the darker objects are more and more recognized as dead cells Thus the measured viability decreases If the parameter is changed towards positive values the amount of objects counted as living cells will increase Thus the measured viability increases A change towards positive values could make sense if due to the condition of the cells or he process media the optical appearance has a deficient contrast ATTENTION In standard situations a change of this parameter results in labeling of debris objects In the range 10 10 the Std Cells 5 00 operator behaves similar to the Std Cells 3 0 operator 24 Admini
151. uremernt nennen 33 Measurement chamber 13 89 Measurement chatt 46 Measurement Offset 56 Measurement parameter 70 Measurement procedure 14 Measurement Series in the diagram 56 Measurement Series offset 56 Measurement system 13 22 Menu Dal 0cccccccceceeeseeeeeeeeeeeseeeeeeeeeees 30 Micro organisms u een 22 96 User Manual Cedex Version 1 6 1 Microscope Optic u nennen 89 Microscope symbol uuseneennee 31 Minimum pollution area 77 N NaOH ecciesia enia S 86 Neubauer hemocytometer 14 16 INOW vessecdeadeercncetathaned dgarancccxshancdectesansansesesaes 55 Number of Cells nen 44 O OSE tirienn a 56 Operating option 31 34 Operator a 11 75 Output File Name 0 60 Output Format nenenn 60 P Parameter cccccseeeeeeeeeeees 32 35 71 72 Password siiis erkannten ira 73 PSren espn nnen aa 44 PrevieW ninen eanna aaa 54 Primera een 33 Print nl ia 54 Print c lored u schnee 54 Printing ea 69 71 Printing User defined Results 54 PrintOptions nassen 54 Process paramelters nn 32 Progress B r u un seenaendaeke 53 Protein aggregate nuueesnnneennnenn 83
152. urity Question window in which information on whether all or only the selected measurement series should be deleted must be entered appears when the New button is actuated After selection and confirmation with Yes new measurement series can then be selected in the Selection of Data Series window The newly selected measurement series can then be adopted in the corresponding row of the Data Series table User Manual Cedex Version 1 6 1 55 Security question Do you really want to remove the following data series from the list Figure 24 The Security Question window Display ofthe Measurement Series in the Diagram Once a data set has been constructed the display of the very measurement series in the diagram can be switched on On or switched off by activating or deactivating the corresponding checkbox in the on off column respectively Measurement Series Offset In the diagram measurement series are automatically displayed such that all measurement series begin at time 0 The measurement series can however then be well compared to each other only when the beginning of the exponential growth phase log Phase sets in at the same time That s why it is possible to shift the curves in the diagram Behind every measurement series that is displayed in the Data Series list a button in the Offset column displays the offset set by the user the button displays 0 h if offset was not set Clicking on this button opens the Measure
153. ution is free of contamination otherwise filter prior to use Check that the sample cup is free of contamination before use Only clean sample cups can guarantee an optimal function of the Cedex If not in use keep the Trypan Blue solution in a refrigerator Secure your measurement data frequently recommended approx every after 250 measurements NOTES EVERY CEDEX SYSTEM IS THOROUGHLY TESTED IN THE END CONTROL ALL RESULTS OF THIS CONTROL TESTING IS SAVED IN THE SYSTEM FOR POSSIBLE USE LATER THE SOFTWARE AND HARDWARE VERSION OF YOUR CEDEX ANALYSIS SYSTEM CAN DIFFER FROM THE INFORMATION DESCRIBED HERE BUT SHOULD NOT MAKE A DIFFERENCE TO THE OPERATIONAL FUNCTIONS IN GENERAL IMPORTANT NOTICE THE MANUFACTURER IS NOT RESPONSIBLE OR LIABLE FOR ANY PERSONAL OR OTHER DAMAGE RESULTING FROM IMPROPER USE OR FAILURE TO COMPLY WITH THE REGULATIONS IN THE OPERATING INSTRUCTIONS 10 User Manual Cedex Version 1 6 1 Introduction 2 introduction The Cedex system determines the cell density and viability of cell suspensions fully automatic The analysis is based on the Trypan Blue exclusion method whereby cell suspensions are stained and analyzed The obtained data allows the evaluation of the overall state of the cell culture including the calculation of specific consumption needs and proliferation rate as well as the documentation of the cultivation process The complete procedure from sample preparation to optical recognition un
154. vailable work areas area and a window similar to that of Windows Explorer will appear Select the appropriate target directory Click on Add to include the directory to the list of available target directories Click on Close to shut the selection window Select a default target directory from the Default work area drop down list Click on Save to store the changes Follow the same procedure any time later when adding or deleting target directories Calibrating the Measurement System The Cedex system must be calibrated when it is set up for the first time or after its position is changed The brightness light intensity and focus are optimally set for the objects to be analyzed Calibration is carried out with the supplied Cedex Calibration Beads or a highly concentrated viable cell suspension and takes approximately 15 minutes Follow the instructions below to calibrate the measurement system Transfer 1 mL of the well mixed beads solution Cedex Calibration Beads or the highly concentrated viable cell culture in the sample cup Attach the sample cup to the sample port Click on the Special menu on the Cedex Workbench If user profiles have already been created then the Special menu is only available to the Cedex administrator Click on Calibrate the system and the Calibration window will appear Click on Start calibration Administrator Manual Cedex Version 1 6 1 17 E Calibration Pile E3 20 40 60 60 100 120 140 160 1
155. vironment This equipment generates uses and can radiate frequency energy and if not installed and used in accordance with the instruction manual may cause harmful interference to radio communications Operating this equipment in a residential area is likely to cause harmful interference in which case the user will be required to correct the interference at his own expense Safety Instructions WARNING IT IS IMPORTANT THAT ALL SAFETY INSTRUCTIONS AND GUIDELINES ARE OBSERVED TO ENSURE SAFE AND RELIABLE OPERATIONS OF THE CEDEX SYSTEM e Place the device on a dry and sturdy surface free of vibrations Prevent the device from getting wet Protect the device from dust and smoke Do not expose the device to direct sunlight Ensure there is sufficient air circulation of the Cedex system during operation particularly around the computer and monitor e Operate within a constant temperature range of 50 F 95 F 10 C 35 C The Cedex system should only be used with the supplied devices measurement system computer monitor and software Do not install any additional software on the Cedex computer Only trained personnel in a controlled laboratory environment are to operate the Cedex system All services and repairs should only be performed by authorized service technicians Administrator Manual Cedex Version 1 6 1 7 Cedex e Always ensure that there are sufficient reagents in the reagent containers
156. wing areas Daily events Daily cleaning routines with Prime before starting with measurements and LM ShutDown before turning off the system Cleaning Additional cleaning routines Syringe handling Syringe maintenance e Time to finish Duration of cleaning routine The Daily Events Area The Prime and LM ShutDown can be activated here see Performing a Prime on page 33 Carry out an LM ShutDown prior to every shut down of the Cedex system The liquid management system is then prepared for an extended pause i e overnight or weekend An LM ShutDown is automatically preceded by a Standard Clean routine The 8 way valve is then rinsed with distilled or de ionized water to prevent crystallization Compressed air finally presses the Trypan Blue solution back into its container see Carrying out an LM ShutDown on page 39 Remove the syringe clean and re install it if there is a longer pause see Syringe Cleaning and Maintenance on page 85 User Manual Cedex Version 1 6 1 81 Cedex The Cleaning Area The following cleaning routines are offered e Fast Clean e Standard Clean Intensive Clean e Chamber Clean 1 Chamber Clean 2 Fast Clean and Standard Clean are carried out in the entire liquid management system including capillary tubes measurement chamber flowcell and sample port therefore always ensure that there is a sample cup attached to the sample port to prevent flooding Fast Clean wit
157. wing the instructions below 1 Place the mouse pointer on the desired area in the diagram 2 Hold the right mouse button pressed down and move the chart as desired Return to the original size of the chart by following the instructions below 1 Place the mouse pointer anywhere on the diagram 2 Hold the left mouse button pressed down and form a box from the bottom right corner to the upper left corner 3 Release the mouse button The chart will then be in its original size User Manual Cedex Version 1 6 1 43 The Aggregate Histogram The Cedex system offers a throughout evaluation of cell cultures in aggregates Clusters of up to 15 cells are displayed The Aggregate Histogram button in the Cedex Workbench is used to call the corresponding histogram The Aggregate Histogram shows the amount of cells of the cell culture available in aggregates and how many cells the aggregate is composed of Both the cell amount as well as their percentage distribution can be displayed EP Aggregate Screen Aggregate Histogram F viable H dead EE En ee a ed ee ee I et ee Se BR ees aio eee oe eee ie ee a kan ae a eae nae eae eet aaa eae eee eee aan ae Be eS Se Oe Pee Se E oD eT ae PE er Figure 11 Aggregate Histogram Detailed View partitioned in living and dead cells here number of cells The identified Number of Cells is displayed on the y axis The Aggregate Size in the amount of cells per aggregate is given
158. ysis results and measurement data modify measurement parameters and 3 to reprocess recalculate measurements on the basis of existing cell images The relevant entry and selection areas for choosing the operating option and specifying the sample data can be found in the Data Source Identification area on the Cedex Workbench Follow the detailed instructions to select the desired operating option and specify the sample data Click on carrying out a measurement 2 Enter a clearly defined reactor ID to describe the sample e g the name of the cell culture in the Reactor ID area You may either choose a reactor ID from the drop down list or enter a new reactor ID A new entry is defined by 25 alphanumerical symbols 3 Enter a clearly defined sample ID in the Sample ID area e g P001 The sample ID can be freely defined Ifthe intended measurement is not the first in a row of measurements i e if it has the same reactor ID as a preceding measurement then the analysis software automatically enters the consecutive sample ID into the Sample ID area e g P001 P002 It is not necessary to enter the sample ID manually 4 Click on the traffic light symbol to confirm or press lt Return gt The traffic light symbol then changes to green This method of entry defines the sample very clearly and automatically allows the Cedex system to create an appropriate directory structure on the hard disk to save sample data and images durin
159. ysis software and information needed by the support The User Identification area The username continues to be displayed in this area after the login and the user has two options to choose from to logout of the Cedex system Logout or to shut down the analysis software Shut down The Data Source Identification area All relevant sample data is entered into this area Reactor ID and Sample ID including the selected operating mode The Data Set drop down list is only relevant when editing the analysis data see Choosing the Operating Option and Specifying Sample Data on page 34 The Microscope Symbol Click here to start anew measurement The Floppy Disk Symbol H Click here to review measurement data or correct measurement parameters dilution comment valid images etc The Calculator Symbol Click here to reprocess existing measurement data User Manual Cedex Version 1 6 1 31 Cedex The Sample Properties Area The Cedex system automatically enters the information in this area e g username date time Entering a comment is optional see Entering a Comment on page 35 The Process Parameters Area All measurement and analysis parameters are entered in this area Cell Type Dilution Images number of images and the Processtype processing mode see Defining the Measurement and Analysis Process Parameters on page 35 The Audit Trail button allows the analysis results and their modifications or change

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