Instructions for Use. COULTER EPICS XL Flow Cytometer COULTER EPICS XL-MCL Flow Cytometer SYSTEM II Software. PN CA (September 2010)

Transcription

1 READY ON FS SS AUX FL1 FL2 FL3 FL4 TM COULTER EPICS XL Flow Cytometer COULTER EPICS XL-MCL Flow Cytometer SYSTEM II Software Instructions for Use CYTOMETER LASER SAMPLE FLOW LOW MED HIGH (September 2010) Beckman Coulter, Inc. 250 S. Kraemer Blvd. Brea, CA 92821

2 WARNINGS AND PRECAUTIONS READ ALL PRODUCT MANUALS AND CONSULT WITH BECKMAN COULTER-TRAINED PERSONNEL BEFORE ATTEMPTING TO OPERATE INSTRUMENT. DO NOT ATTEMPT TO PERFORM ANY PROCEDURE BEFORE CAREFULLY READING ALL INSTRUCTIONS. ALWAYS FOLLOW PRODUCT LABELING AND MANUFACTURER S RECOMMENDATIONS. IF IN DOUBT AS TO HOW TO PROCEED IN ANY SITUATION, CONTACT YOUR BECKMAN COULTER REPRESENTATIVE. HAZARDS AND OPERATIONAL PRECAUTIONS AND LIMITATIONS WARNINGS, CAUTIONS, and IMPORTANTS alert you as follows: WARNING - Can cause injury. CAUTION - Can cause damage to the instrument. IMPORTANT - Can cause misleading results. BECKMAN COULTER, INC. URGES ITS CUSTOMERS TO COMPLY WITH ALL NATIONAL HEALTH AND SAFETY STANDARDS SUCH AS THE USE OF BARRIER PROTECTION. THIS MAY INCLUDE, BUT IT IS NOT LIMITED TO, PROTECTIVE EYEWEAR, GLOVES, AND SUITABLE LABORATORY ATTIRE WHEN OPERATING OR MAINTAINING THIS OR ANY OTHER AUTOMATED LABORATORY ANALYZER. WARNING Risk of operator injury if: r All doors, covers and panels are not closed and secured in place prior to and during instrument operation. r The integrity of safety interlocks and sensors is compromised. r Instrument alarms and error messages are not acknowledged and acted upon. r You contact moving parts. r You mishandle broken parts. r Doors, covers and panels are not opened, closed, removed and/or replaced with care. r Improper tools are used for troubleshooting. To avoid injury: r Keep doors, covers and panels closed and secured in place while the instrument is in use. r Take full advantage of the safety features of the instrument. Do not defeat safety interlocks and sensors. r Acknowledge and act upon instrument alarms and error messages. r Keep away from moving parts. r Report any broken parts to your Beckman Coulter Representative. r Open/remove and close/replace doors, covers and panels with care. r Use the proper tools when troubleshooting. CAUTION System integrity might be compromised and operational failures might occur if: r This equipment is used in a manner other than specified. Operate the instrument as instructed in the Product Manuals. r You introduce software that is not authorized by Beckman Coulter into your computer. Only operate your system s computer with software authorized by Beckman Coulter. r You install software that is not an original copyrighted version. Only use software that is an original copyrighted version to prevent virus contamination. IMPORTANT If you purchased this product from anyone other than Beckman Coulter or an authorized Beckman Coulter distributor, and, if it is not presently under a Beckman Coulter service maintenance agreement, Beckman Coulter cannot guarantee that the product is fitted with the most current mandatory engineering revisions or that you will receive the most current information bulletins concerning the product. If you purchased this product from a third party and would like further information concerning this topic, call your Beckman Coulter Representative.

3 REVISION STATUS Issue A, 1/96 SYSTEM II Software Version 1.0. Initial issue for customer distribution. Issue B, 12/98 Complete revision. SYSTEM II Software, Version 3.0. Includes new quality control features, patient report export feature, and user-defined run time reports. Issue C, 6/03 Changes were made to, r r comply with the EU IVD Directive (98/79/EC). change the company name from Coulter Corporation to Beckman Coulter Inc. Issue CA, 9/10 Updates were made to the company corporate address. Note: Changes that are part of the most recent revision are indicated in text by a bar in the margin of the amended page. This document applies to the latest software listed and higher versions. When a subsequent software version changes the information in this document, a new issue will be released to the Beckman Coulter website. For labeling updates, go to and download the most recent manual or system help for your instrument. iii

4 REVISION STATUS iv

5 CONTENTS REVISION STATUS, iii INTRODUCTION, xi USING YOUR XL AND XL-MCL FLOW CYTOMETER MANUALS, xi ABOUT THIS MANUAL, xi CONVENTIONS, xii Description of Reporting Units, xiii ICONS, xiii 1 STARTUP, BEFORE YOU BEGIN, DAILY STARTUP PROCEDURE, QUALITY CONTROL, QC MATERIALS, MANUAL QC VS. AUTOSTANDARDIZATION, MANUAL QC, 2-3 Running Flow-Check Fluorospheres, 2-3 Running Flow-Set Fluorospheres, 2-6 Running CYTO-COMP Cells and Reagents, 2-7 Running Immuno-Trol Cells or CYTO-TROL Control Cells, AUTOSTANDARDIZATION, 2-12 Protocol Naming Conventions, 2-12 _A protocols, 2-12 _C protocols, 2-12 _Q protocols, 2-12 Reviewing QC Data From Autostandardization Panels, RUNNING AUTOSTANDARDIZATION PANELS, 2-13 Prepare QC Materials, 2-13 Running Autostandardization Panel or Protocol for Flow-Check Fluorospheres, 2-14 Running Autostandardization Panel for Immunophenotyping, SETTING UP AUTOSTANDARDIZATION PROTOCOLS, 2-19 Setting Up a Flow-Check Fluorospheres Protocol, 2-19 Setting Up _Axyz Protocols, 2-22 Setting Up _C Protocols, 2-25 Setting Up _QxyzABCD Protocols, CREATING AUTOSTANDARDIZATION PANELS, 2-31 Creating a Flow-Check Fluorospheres Panel, 2-31 Creating an Immunophenotyping Panel, 2-32 v

6 CONTENTS 3 CREATING A WORKLIST, OVERVIEW, ENTERING SPECIMEN ID, 3-2 Using the Keyboard, 3-2 Using the Hand-Held Bar-Code Scanner, ENTERING PATIENT INFORMATION, 3-3 Go to the Patient Pop-Up Box, 3-3 Enter the Patient Name, 3-3 Enter the Patient ID, 3-4 Enter the DOB (Date of Birth), 3-4 Enter Additional Information, SAVING INFORMATION TO THE DATABASE, 3-5 Retrieving Records, ASSIGNING PANELS TO A WORKLIST, 3-6 Copying Panels on the Worklist Screen, 3-7 Moving Lines on the Worklist Screen, 3-8 Erasing Worklist Information, ASSIGNING WORKLIST TUBE ID, 3-9 Before Assigning Tube ID Bar-Codes, 3-10 Assigning the Bar Codes, 3-10 Using the Keyboard or Hand Held Scanner, 3-10 Using the Assign BC Button, 3-11 Erasing Tube ID, SAVING THE WORKLIST, RUNNING SAMPLES, BEFORE RUNNING SAMPLES, RUNNING A SAMPLE IN THE MANUAL MODE (XL OR XL-MCL FLOW CYTOMETER), RUNNING SAMPLES IN THE AUTO MODE (XL-MCL FLOW CYTOMETER ONLY), 4-3 Interrupting the Auto Mode Cycle, 4-6 Finishing the Tube, 4-6 Aborting the Tube, 4-6 Resuming the Auto Mode Cycle After You Interrupt a Panel, 4-7 What the Completion Codes in the Carousel Summary Mean, DATA ACQUISITION FUNCTIONS, 4-8 PRIME Button, 4-8 Cytometer Setting Adjustments, 4-9 Using Fast Set, 4-9 vi

7 CONTENTS Using Fast Comp, 4-10 Restarting the Cytometer after Making Changes, 4-10 Acquisition Buttons and Keys, 4-10 Editing Regions During Acquisition, 4-11 Creating, Erasing, Copying, or Redrawing Regions During Acquisition, 4-11 Alt Save Button, 4-12 Histogram Display Keys, DATA ANALYSIS, LISTMODE ANALYSIS, REPLAYING INDIVIDUAL LISTMODE FILES, Selecting the Files, Reordering the List Queue, Saving the List Queue, Choosing Output Options, Choosing the Replay Mode, Replaying The Files, Editing Regions, REPLAYING LISTMODE FILES AS A PANEL, Selecting the Application, Choosing the Replay Mode, Selecting the Panel, Selecting the Files, Reordering the List Queue, Saving the List Queue, Choosing Output Options, Replaying The Files, Editing Regions, LISTMODE BATCH ANALYSIS, Creating a Listmode Queue, Loading a Listmode Queue, Choosing Output Options, Choosing the Replay Mode, Adjusting Regions, Replaying Batch, Stopping Batch Listmode Replay, HISTOGRAM ANALYSIS WITH MULTIGRAPH, Selecting the Files, Reordering the Hist Queue, Saving the Hist Queue, Printing and Saving in Multigraph, Choosing the Analysis Mode, Loading and Passing Regions, Editing Regions, Displaying Next Set and Next Test, 5-14 vii

8 CONTENTS 5.6 HISTOGRAM BATCH ANALYSIS, Creating a Hist Queue, Loading a Histogram Queue, Printing and Saving in Multigraph, Choosing the Analysis Mode, Loading and Passing Regions, Editing Regions, Analyzing the Queue, Stopping Batch Multigraph Replay, HISTOGRAM DISPLAYS, 5-17 Displaying Single Overlay Histograms, 5-17 Dual Overlay Histogram Displays, 5-18 Hist Data Screen, PRINTING AND EXPORTING PATIENT REPORTS, INTRODUCTION, PRINTING PATIENT REPORTS, EXPORTING PATIENT REPORTS (*.PXP) FILES, 6-3 Overview, 6-3 Exporting from the Acquisition Application, 6-3 Exporting from the Listmode Application, 6-5 Exporting from the Reports Data Entry Screen, 6-7 Viewing a *.PXP File, REVIEWING QC DATA, OVERVIEW, 7-1 Relationship of Protocols to QC Templates and QC Data Files, LOADING A QC TEMPLATE, 7-1 Overview, 7-1 Procedure, RELATIONSHIP BETWEEN THE LEVEY-JENNINGS GRAPHS AND THE QC DATA TABLE, 7-4 Data Colors, 7-4 Lot Numbers, REVIEWING LEVEY-JENNINGS GRAPHS, REVIEWING THE QC DATA TABLE, COMPARING TWO QC FILES, WORKING WITH THE LOT INFORMATION POP-UP BOX, 7-13 viii

9 CONTENTS 8 CLEANING AND SHUTDOWN, 8-1 INDEX 8.1 CLEANING AND SHUTDOWN FREQUENCY, 8-1 When to Clean the Sampling System, 8-1 When to Shut Down the Cytometer, ROUTINE CLEANING PROCEDURE, 8-1 Manual Mode Procedure (XL and XL-MCL Flow Cytometers), 8-2 Auto Mode Procedure (XL-MCL Flow Cytometer Only), 8-4 Testing for Residual Stain, SAMPLE HEAD AND PROBE CLEANING PROCEDURE, 8-5 Manual Mode Procedure (XL and XL-MCL Flow Cytometers), 8-5 Auto Mode Procedure (XL-MCL Flow Cytometer Only), VACUUM LINE CLEANING PROCEDURE, 8-7 Auto Mode Procedure (XL-MCL Flow Cytometer Only), 8-7 Manual Mode Procedure (XL and XL-MCL Flow Cytometers), SHUTDOWN PROCEDURE, 8-9 TRADEMARKS ix

10 CONTENTS x

11 INTRODUCTION This introductory section contains the following topics: r r r r Using your COULTER EPICS XL and XL-MCL Flow Cytometer manuals About this manual Conventions Icons. USING YOUR XL AND XL-MCL FLOW CYTOMETER MANUALS Use the Reference manual for in-depth information on the principles of flow cytometry, information about what your instrument does, the methods it uses, its specifications, and information on installation, safety, and system options. Use the Getting Started manual to become familiar with the controls and indicators for your system and to learn about protocols, regions, panels, and the basic skills you need to operate the system. This manual also has an overview of the software. Use the Operator s Guide for the day-to-day running of your instrument. Go through the detailed step-by-step procedures of startup, quality control (QC), running samples, analyzing data, printing reports, reviewing QC data, and shutdown. Use the Data Management manual for instructions on how to export, save, copy, move, archive, and delete files. It also has information about the types of files your system creates and uses, instructions for working with QC features, and instructions for setting up the report template that you use to create your patient reports. Use the Special Procedures and Troubleshooting manual to clean, replace, or adjust a component of the instrument. The Troubleshooting tables and error messages are at the back of the manual. Use the Operating Summary as a quick reference for basic procedures. Use the Master Index to easily locate a topic in any of your manuals. Use the User's Comment Card in the Reference manual to give us your comments about the manual and ways to improve it. ABOUT THIS MANUAL Your EPICS XL and XL-MCL Flow Cytometer Operator s Guide describes the day-to-day running of your instrument. Read the controls and indicators chapter in the Getting Started manual to become familiar with the different parts of your system. Then go through the detailed step-by-step procedures in each chapter of this manual. This information is organized as follows: s s Chapter 1, Startup Contains instructions for the daily startup routine, checking system pressure and vacuum. Chapter 2, Quality Control Contains instructions for the Quality Control (QC) procedures using COULTER Flow-Check Fluorospheres, Flow-Set Fluorospheres, CYTO-COMP Cells, xi

12 INTRODUCTION CONVENTIONS s s s s s s s CYTO-COMP Reagents, Immuno-Trol Cells, Flow-Count Fluorospheres, and CYTO-TROL Control Cells. Chapter 3, Creating a Worklist Contains instructions for creating and using a worklist to run samples and save patient information to the system database. Chapter 4, Running Samples Contains instructions for running samples in the manual and automatic modes and instructions for Post Run Time Functions. Chapter 5, Data Analysis Contains instructions for analyzing samples using Listmode and MultiGraph applications. Chapter 6, Printing and Exporting Patient Reports Contains instructions for printing patient reports after data acquisition procedures. Contains instructions for exporting patient reports in the acquisition and listmode applications. Chapter 7, Reviewing QC Data Contains instructions for reviewing QC data. Chapter 8, Cleaning and Shutdown Contains instructions for the daily shutdown and cleaning procedures. Index Use the Index to easily locate specific information in this manual. CONVENTIONS This manual uses the following conventions: Throughout this manual your EPICS XL or XL-MCL flow cytometer is referred to as the system. Italics indicate screen messages. Bold indicates a menu item. Courier font indicates text you have to type using the keyboard. ë indicates a key (such as Û). ë+ë indicates that the two keys listed (such as Þ+Ê) are linked for a specific function and must be pressed in this sequence: 1. Press down on the first key listed and while continuing to press it, press down on the second key listed. 2. Release both keys at the same time. ë ë indicates to press and release the first key listed then press and release the next key listed. For example, Y Û. File tt Save Select the Save item on the File menu. [OKAY] Use the mouse to click on the screen button labeled [OKAY]. É through Ô Special function keys. xii

13 INTRODUCTION ICONS Description of Reporting Units Unless otherwise stated, all parameter units are shown in the US unit format (cells/µl) throughout the manuals. ICONS Press and release the left mouse button. Press and release the right mouse button. Move the mouse without pressing either button. Press and hold the right mouse button while moving the mouse. Turn off the system at the computer. Press and hold the left mouse button while moving the mouse. Operational check. Turn on the system at the computer. Power Supply. No fluid. Close the Power Supply door.open the Power Supply door. Open the Power Supply door. Call your Coulter Representative. xiii

14 INTRODUCTION ICONS xiv

15 1STARTUP BEFORE YOU BEGIN This chapter explains the daily startup procedures. Before doing these procedures: 1. Read the Reference manual. Using your system is easier if you have a general understanding of how it works. 2. Read the Getting Started manual. It contains instructions for the Controls and Indicators and Learning the Basics for using your system. You use these basic techniques in the other chapters. 3. Read each procedure entirely. 1.2 DAILY STARTUP PROCEDURE Perform the following steps to start up the system. If you did the daily shutdown procedure in the Shutdown chapter, a test tube of deionized water is still on the sample stage, which is up. 1 Turn on the system at the computer. 2 If you have a FlowCentre Workstation and it is configured to start from an XL icon: r Double click the XL icon to start the software. 3 During system startup, the following series of Cytometer status messages are displayed. The startup cycle includes a prime cycle. Startup in Process System Verification Run Initialization Please Wait... Processing. 1-1

16 STARTUP DAILY STARTUP PROCEDURE 4 If you have not specified a startup protocol or panel on the Utilities Configuration screen, perform the following steps to select one: Protocol, Panel, or Worklist. A protocol, panel, or worklist you want to use. [Okay]. 5 The message Insert Sample Tube appears. 6 Open the Power Supply door. 7 Check the WATER TRAP, AIR FILTER, and VACuum FILTER. Call your Coulter Representative if: r r The TRAP is more than one-third full. The FILTERS have any fluid. VAC FILTER VAC TRAP AIR FILTER SYS VAC WATER TRAP SYS PRESS PRESS ADJ 30 PSI MCL P A 1-2

17 STARTUP DAILY STARTUP PROCEDURE 1 8 Check the SYStem VACuum gauge. If it reads less than 17 in. Hg, call your Coulter Representative. 17" Hg SYS VAC B 9 Check the SYStem PRESSure gauge. If it does not read between 28 and 32 psi, do the following: 30 ±2 PSI SYS PRESS B a. Pull the collar around the PRESSure ADJuster out toward you A 1-3

18 STARTUP DAILY STARTUP PROCEDURE b. Adjust the pressure. (+) (-) C c. Push in on the collar to lock it into place A 1-4

19 STARTUP DAILY STARTUP PROCEDURE 1 10 Check the VACuum TRAP. If it is more than 1/4 full of fluid, clean it as explained in the Special Procedures and Troubleshooting manual. VAC FILTER VAC TRAP AIR FILTER WATER TRAP 30 PSI SYS VAC SYS PRESS MCL P LESS THAN 1/4 FULL PRESS ADJ A 11 Close the power supply door. 12 Remove the tube from the Sample Stage A 1-5

20 STARTUP DAILY STARTUP PROCEDURE 13 Record the startup checks on the QC Maintenance screen: Applications tt QC. Screen tt Maintenance. Type in the actual readings in the System Vacuum and System Pressure boxes. Each of the remaining boxes related to today s daily startup checks. This enters your operator ID color and symbol. If you need to enter your operator ID into the system: The operator ID field in the upper left portion of the screen and enter your operator ID. See the Special Procedures and Troubleshooting manual for detailed instructions on the QC Maintenance screen. 14 Refer to the manuals that came with your Printer to: r Perform Printer diagnostics. r Check ink cartridges if you have a color Printer and replace if necessary. 1-6

21 2QUALITY CONTROL 2 Perform the following quality control checks to ensure that your system is working accurately and precisely. The protocols needed for these quality control (QC) procedures are included with your system software. You can run the QC protocols manually. Instructions are listed under heading 2.3, Manual QC, in these subheadings: r r r Running Flow-Check Fluorospheres Running Flow-Set Fluorospheres Running CYTO-COMP Cells stained with CYTO-COMP Reagents, or other compensation reagents r Running Immuno-Trol Cells or CYTO-TROL Control Cells. You can also use heading 2.4, Autostandardization, to create Autostandardization panels that allow you to automate the QC procedures. Heading 2.2, Manual QC vs. Autostandardization, describes the differences between the two methods of performing QC. 2.1 QC MATERIALS Use the following QC materials. COULTER Flow-Check Fluorospheres COULTER Flow-Set Fluorospheres COULTER CYTO-COMP Cells stained with CYTO-COMP Reagents Fluorospheres used to check the stability of the optical and fluidics systems. Fluorospheres used to standardize the light scatter intensity and fluorescence intensity. CYTO-COMP Cells stained with CYTO-COMP Reagents are used to adjust color compensation settings for multicolor analysis. OR Other compensation reagents COULTER Immuno-Trol Cells OR COULTER CYTO-TROL Control Cells A normal representative specimen stained with mutually exclusive cell surface markers. Stabilized erythrocytes and leukocytes with a known quantity of surface antigens. Used to verify monoclonal antibody performance as well as verify the process of sample staining, lysing, and analysis.v Lyophilized lymphocytes with a known quantity of surface antigens. Used to verify monoclonal antibody performance. The package inserts for these quality control materials have instructions for establishing your laboratory's normal ranges for daily use. You must re-establish your laboratory's ranges: r When you use a new lot of fluorospheres. 2-1

22 QUALITY CONTROL MANUAL QC VS. AUTOSTANDARDIZATION r Whenever a major part of the system has been serviced or replaced (for example, laser alignment or replacement, PMT replacement). In addition to doing the daily quality control procedure in this chapter, you should make a quality control check for the specific applications you are running. 2.2 MANUAL QC VS. AUTOSTANDARDIZATION The chart below shows the differences between performing daily QC manually or by using the Autostandardization protocols and panels. QC Process Manual Method Autostandardization Method Verification of fluidics and alignment Adjustment of high voltage and gain for a given application Adjustment of color compensation for a given application Verification of correct settings with an application Control Update all required protocols with new settings Update Levey-Jennings graphs for QC monitoring. Flow-Check Fluorospheres. Verify HPCV versus expected value. Record settings manually. Flow-Set Fluorospheres. Ascertain target mean position based upon application and adjust high voltage and gain daily to that target. Record settings manually. CYTO-COMP Cells stained with CYTO-COMP Reagents, or whole blood stained with mutually exclusive markers. Adjust color compensation to within defined difference in mean intensity between the positive and negative population. Record settings. Update the control protocol with the settings derived from above. Run a biological control equivalent to the application, such as Immuno-Trol Cells, CYTO-TROL Control Cells, or a normal whole blood. Compare values to the control assigned levels. The patient report generator can be used to flag low and high values. Determine whether the Cytometer is set correctly for the application. User selects all appropriate protocols and manually updates each with the Cytometer settings. User manually maintains Levey-Jennings graphs. Flow-Check Fluorospheres. SYSTEM II software compares HPCV to value entered. Flags are Pass, Population % or Fail. Results are stored in a QC file. Flow-Set Fluorospheres. SYSTEM II software adjusts high voltage and gain to a defined target mean position based upon application. Settings are stored in a QC file. CYTO-COMP Cells stained with CYTO-COMP Reagents, or whole blood stained with mutually exclusive markers. SYSTEM II software adjusts color compensation to within a defined difference based upon application. Adjustment is flagged as pass or fail. Settings are stored in a QC file. SYSTEM II software updates the verification control protocol with the settings derived for the application. Immuno-Trol Cells or CYTO-TROL Control Cells are run and compared to the assigned levels. The patient report generator can be used to flag low and high values. User determines whether the Cytometer is set correctly for the application. User accepts or rejects new settings. SYSTEM II software updates all identified protocols from the QC file. SYSTEM II software maintains Levey-Jennings quality control graphs for specified parameters. 2-2

23 QUALITY CONTROL MANUAL QC MANUAL QC Manual QC consists of running: r r r r Flow-Check Fluorospheres Flow-Set Fluorospheres CYTO-COMP Cells stained with CYTO-COMP Reagents, or other compensation reagents Immuno-Trol Cells, CYTO-TROL Control Cells, or normal sample. Running Flow-Check Fluorospheres Run Flow-Check Fluorospheres to check the stability of your instrument s optical and fluidics systems Check that the daily startup procedure (heading 1.2) was performed. Allow about 30 minutes to warm-up the system. 3. Check the Cytometer status message. Start this procedure when the Cytometer status message Insert Sample Tube appears. Record any error messages in your instrument s logbook. You can, however, use the software while waiting for the message to appear. For example, you can create protocols. 4. Protocol tt Select. The Flow-Check protocol. [Okay]. 2-3

24 QUALITY CONTROL MANUAL QC 5. If the [Save Cyto] button is on (green), change it to off (blue). This prevents overwriting the Cytometer settings in the Flow-Check protocol. 6. Put 15 to 20 drops ( 0.5 ml) of Flow-Check Fluorospheres in a test tube. (Follow the package insert instructions for mixing and handling these fluorospheres.) C B 7. Put the tube on the sample stage. A B B 8. After data acquisition starts: Histogram 1. Ensure that the region includes only the main population. Edit the region if it does not fully enclose the population. 9. [Next Hist] button. 2-4

25 QUALITY CONTROL MANUAL QC Wait for data to be acquired and acquisition to stop. 11. Follow the directions in the Flow-Check Fluorospheres package insert to establish expected range values for your laboratory. If you change the Cytometer settings in this protocol, you must re-establish your laboratory s expected ranges. Check that the values for Peak Position, and the half-peak coefficient of variation (HPCV) for all parameters are within your laboratory's acceptance limits and record them in a logbook. 12. After acquisition stops and the sample stage lowers, remove the tube A 2-5

26 QUALITY CONTROL MANUAL QC Running Flow-Set Fluorospheres Run Flow-Set Fluorospheres to standardize the light scatter and fluorescence intensity. 1. Check that the Cytometer is within your laboratory's acceptance limits for Flow-Check Fluorospheres. 2. Protocol tt Select. The Flow-Set protocol. [Okay]. 3. Follow the directions in the Flow-Set Fluorospheres package insert to establish expected range values for your laboratory. Check that the [Save Cyto] button is on (green). 4. Put 15 to 20 drops ( 0.5 ml) of Flow-Set Fluorospheres in a test tube and load onto the sample stage. (Follow the package insert instructions for mixing and handling these fluorospheres.) C B 5. After data acquisition starts: Histogram 1. Ensure that the peak is within the region. If the PK Position is not within the region, adjust the FS voltage as needed to place the population within the acceptance limits. 2-6

27 QUALITY CONTROL MANUAL QC 2 6. Wait for data to be acquired on 10,000 events and acquisition to stop. 7. Check that the peak position for each parameter is within your laboratory's acceptance limits and record them in a logbook. If the peak positions are not within the acceptance limits, adjust PMT high voltage to position the peaks within the acceptance limits Running CYTO-COMP Cells and Reagents Run the CYTO-COMP Cells stained with CYTO-COMP Reagents or run other compensation reagents to calibrate your system s color compensation Check that the Cytometer has been standardized with Flow-Set Fluorospheres. Protocol tt Select. The Flow-Set protocol. [Okay]. 3. Check that the [Save Cyto] button is on (green). 2-7

28 QUALITY CONTROL MANUAL QC 4. Update the light scatter and fluorescence Cytosettings to match those for Flow-Set Fluorospheres. 5. Prepare the CYTO-COMP Cells and stain them with the CYTO-COMP Reagents as instructed in their package inserts. OR Prepare the compensation reagents. Use a normal representative specimen stained with mutually exclusive cell surface markers. 6. Load the test tube onto the sample stage. 7. After data acquisition starts: Histogram Check that the cells of interest are within the gate region. [Next Hist] button. Wait for data to be acquired on 1,000 events. 9. Adjust color compensation for each dual parameter histogram so that: r The mean intensity of Q1 = mean intensity of Q3 for X-axis. r The mean intensity of Q4 = mean intensity of Q3 for Y-axis. Press [Restart] after each time you change a color compensation setting. If the differences are not within your laboratory s acceptance limits, adjust the color compensation again. Record the results in a logbook. 2-8

29 QUALITY CONTROL MANUAL QC 2 Running Immuno-Trol Cells or CYTO-TROL Control Cells 1. Check that the Cytometer has been standardized with Flow-Set Fluorospheres and the color compensation has been calibrated with compensation reagents. 2. Use the appropriate amount of Immuno-Trol Cells according to the instructions on the package insert or reconstitute the CYTO-TROL Control Cells according to the instructions on the package insert. 3. Stain the cells with the monoclonal antibodies you use for the protocol or panel. 4. Note: If you want to obtain absolute counts, add the appropriate amount of Flow-Count Fluorospheres to the Immuno-Trol Cells. Panel or Protocol tt Select. The appropriate Immuno-Trol or CYTO-TROL protocol or panel. [Okay]. 2-9

30 QUALITY CONTROL MANUAL QC 5. Update the light scatter and fluorescence high voltage and gain settings to match the results from the analysis of Flow-Set Fluorospheres. 6. Update the color compensation settings to match the results from the analysis of compensation reagents. 7. Place the first stained tube (with Immuno-Trol Cells or CYTO-TROL Control Cells) on the system. Pause the system and create or edit an FS SS region to include only lymphocytes. Immuno-Trol Cells CYTO-TROL Control Cells 2-10

31 QUALITY CONTROL MANUAL QC 2 8. Use this lymphocyte region to gate the remaining histograms. Acquire 5,000 gated events. 9. Restart acquisition and print the protocol results. 10. Check that the percentages for all parameters are within the limits given on the package insert. If not, troubleshoot your sample preparation. Immuno-Trol Cells CYTO-TROL Control Cells 2-11

32 QUALITY CONTROL AUTOSTANDARDIZATION 2.4 AUTOSTANDARDIZATION The Autostandardization protocols use specific naming conventions depending upon the function of the protocol. You must run these protocols sequentially in a panel to standardize the Cytometer and pass the settings on to the test protocols in the panel. Protocol Naming Conventions _A protocols r Automatically standardize high voltage and gain settings. r r Must be assigned as Primary protocols so that the settings are passed to the other protocols in a panel. Generate and/or update *.QCS files that contain the voltages, gains, and compensation values for daily QC monitoring. _C protocols r Standardize fluorescent (color) compensation. r Must be assigned as Secondary protocols in a panel. r Update the compensation values in the *.QCS file generated by the _A protocol run in the panel. _Q protocols r Verify the Cytometer setup after you have run the _A and _C protocols. r r r Generate and/or update *.QCC files that contain the QC statistical data for all QC regions. When used alone (as in a Flow-Check panel), a _Q protocol must be assigned as a Primary protocol. When used with _A and _C protocols, _Q protocols must be assigned as Secondary or Control protocols at the end of the Autostandardization panel. Reviewing QC Data From Autostandardization Panels See Chapter 7, Reviewing QC Data, for additional information on reviewing QC data produced by the Autostandardization Panels. 2-12

33 QUALITY CONTROL RUNNING AUTOSTANDARDIZATION PANELS RUNNING AUTOSTANDARDIZATION PANELS If you are running Autostandardization for the first time or need to make changes to your Autostandardization protocols or panels see heading 2.6, Setting Up Autostandardization Protocols, and heading 2.7, Creating Autostandardization Panels. Prepare QC Materials 1. Put 15 to 20 drops ( 0.5 ml) of Flow-Check Fluorospheres in a test tube. 2. Put 15 to 20 drops ( 0.5 ml) of Flow-Set Fluorospheres in a test tube. C B 3. Prepare the compensation reagents. Use CYTO-COMP Cells stained with CYTO-COMP Reagents or a normal representative specimen stained with mutually exclusive cell surface markers. 4. Prepare a sample from a specimen with known assay values (Immuno-Trol Cells, CYTO-TROL Control Cells, or normal whole blood) to use for verification. Note: Add Flow-Count Fluorospheres to Immuno-Trol Cells if you want absolute count measurements. 2-13

34 QUALITY CONTROL RUNNING AUTOSTANDARDIZATION PANELS Running Autostandardization Panel or Protocol for Flow-Check Fluorospheres r Do not access the Cytosettings screen while running an Autostandardization panel. r Do not run Autostandardization panels or _A, _C, or _Q protocols in a Worklist. IMPORTANT Acquisition data is lost if you use Ò to Pause and Continue when you are running an Autostandardization panel. If you need to change settings in an Autostandardization panel or protocol, stop acquisition, make the changes and restart the panel from the first tube. 1. Panel The _Qxyz Flow-Check protocol or panel you want to use. [Okay]. 4. Load the tube of Flow-Check Fluorospheres onto the sample stage and acquire data. 2-14

35 QUALITY CONTROL RUNNING AUTOSTANDARDIZATION PANELS 2 5. Check that the peak position of each detector is within the established range. If it is not within range: r [Erase Entry] so that the results of the last run protocol are not saved to a QC file. You can also delete the results from the QC Data Table. r Run the _Axyz Flow-Check protocol with the matching xyz identifier to automatically adjust the detection voltage. r Reanalyze the _Qxyz Flow-Check protocol. 6. When acquisition of the Flow-Check Fluorospheres stops, remove the tube and check that the histograms with HPCV check regions display CV PASS above the histogram. If the HPCVs pass, continue with running the Autostandardization Panel. 2-15

36 QUALITY CONTROL RUNNING AUTOSTANDARDIZATION PANELS If the HPCVs are not within limits you may need to: [Erase Entry] so that the results of the last run protocol are not saved to a QC file. You can also delete the results from the QC Data Table. r r r Pause the system. Prime the system. Refer to the Special Procedures and Troubleshooting manual to troubleshoot. Running Autostandardization Panel for Immunophenotyping r Do not access the Cytosettings screen while running an Autostandardization panel. r Do not run Autostandardization panels or _A, _C, or _Q protocols in a Worklist. IMPORTANT Acquisition data is lost if you use Ò to Pause and Continue when you are running an Autostandardization panel. If you need to change settings in an Autostandardization panel or protocol, stop acquisition, make the changes and restart the panel from the first tube. 1. Panel. The Autostandardization panel you want to use. [Okay]. 2. Load the tube of Flow-Set Fluorospheres onto the sample stage. The _Axyz Flow-Set protocol should be highlighted. 2-16

37 QUALITY CONTROL RUNNING AUTOSTANDARDIZATION PANELS 2 3. When acquisition of the Flow-Set Fluorospheres stops, check the peak positions to ensure they meet preset values. Remove the Flow-Set Fluorospheres tube. If the values are out of range, you may need to: [Erase Entry] so that the results of the last run protocol are not saved to a QC file. Load the appropriate CYTO-COMP reagent tube onto the Sample Stage. The first _C protocol should be highlighted. 4. When acquisition of the compensation reagent stops, remove the compensation reagent tube and check that the histograms with DIFF<n.nn region names display COMP PASS above the histograms. If the region names display COMP FAIL above the histogram, you may need to: Erase Entry] so that the results of the last run protocol are not saved to a QC file. Repeat this step for all of the compensation reagent tubes in the panel. 5. Load the Immuno-Trol or CYTO-TROL con trol sample tube onto the sample stage. The _Qxyz protocol should be highlighted. Note: [PAUSE] is disabled during _Q protocols. 2-17

38 QUALITY CONTROL RUNNING AUTOSTANDARDIZATION PANELS 6. When acquisition of the control sample stops, check that the percentages for all the parameters are within limits for the assay values. Immuno-Trol Cells If they are not within limits: CYTO-TROL Control Cells [Erase Entry] so that the results of the last run protocol are not saved to a QC file. You can also delete the results from the QC Data Table. Troubleshoot the system and run Autostandardization again. 2-18

39 QUALITY CONTROL SETTING UP AUTOSTANDARDIZATION PROTOCOLS SETTING UP AUTOSTANDARDIZATION PROTOCOLS Setting Up a Flow-Check Fluorospheres Protocol _Qxyz Flow-Check is a protocol set up to measure HPCV. The linear regions on each histogram should be named CV<n.nn where n is the acceptable HPCV for instrument operation. The _Qxyz Flow-Check protocol contains histograms to monitor FS, FL1, FL2, FL3, and FL4. If you have a 3 PMT system, delete the FL4 histogram from the protocol. The xyz nomenclature of the _Qxyz Flow-Check protocol retrieves the matching baseline file (nxyz.qcs) containing the stored voltage and gain settings for your instrument. The statistical data from any regions named QC in this _Q protocol are stored into a QC data file named nxyzabcd.qcc where n is the instrument node ID. 1. Setup Screen tt Protocol File tt Select. The _Qxyz Flow-Check protocol you want to use. [Okay]. 2. Edit the parameter user names if you want to change them to the parameters you are investigating. 3. Erase any parameters that you do not need. 2-19

40 QUALITY CONTROL SETTING UP AUTOSTANDARDIZATION PROTOCOLS 4. Check the histogram titled AUTOPRIME to ensure the parameters assigned to it identify the population being monitored. If not: a. Assign parameters that you are investigating to the Autoprime histogram. b. Setup Screen tt Run/ Regions. c. Gate the histogram so as to identify the population of interest. d. Recreate the rectilinear region and name it PRIME. 5. Edit the linear region name on each histogram to the acceptable HPCV for your application. When you run the _Qxyz Flow-Check protocol, the HPCV of each region is compared to the region name. If the HPCV is lower or equal to the region name, the message CV < n.nn PASS is displayed above the histogram. If the HPCV is higher than the region name, then the message CV > n.nn FAIL is displayed on the histogram. If the region percent is <90%, the message CV > n.nn Population% is displayed on the histogram instead of FAIL. 2-20

41 QUALITY CONTROL SETTING UP AUTOSTANDARDIZATION PROTOCOLS 2 If FAIL appears: r r Prime the system. Rerun the sample. Refer to the Special Procedures and Troubleshooting manual to troubleshoot. 6. Create a lot number region (rectilinear or amorphous) to save the lot number of the Flow-Check Fluorospheres with the run s statistical QC data. The lot number and statistical data are posted in the *.QCC file. Name the region LOT#=ABCD1234, where ABCD1234 is the lot number of the Flow-Check Fluorospheres. 7. Create any rectilinear, linear, or amorphous QC regions whose statistics you want to monitor. Be sure to include QC in the region name. Note: You must put QC at the start of the QC region name if you also set up the region to display CV PASS or CV FAIL as in step

42 QUALITY CONTROL SETTING UP AUTOSTANDARDIZATION PROTOCOLS 8. Setup Screen tt Protocol. File tt Save As to save the protocol with a new name for use in your Autostandardization panel. Before running samples, you must load the new protocol into the system so the settings are used from the correct baseline file. Be sure to use _Q plus an xyz identifier as the first five characters of the _Qxyz protocol name. To load the cytosettings appropriate to your application, the QC software uses the xyz nomenclature to obtain your system s baseline file settings. Setting Up _Axyz Protocols The software includes _Axyz protocols. Use _Axyz [3] for three-color immunofluorescence and _Axyz [4] for four-color immunofluorescence. The xyz prefix in the protocol name is a three-letter identifier that you specify. The software uses this three-letter combination as the name of a QC baseline file (nxyz.qcs). The software saves the high voltage and gain settings in the QC baseline file. You use the same three letters at the beginning of any other protocol that you want updated with the settings from this QC baseline file. For example: _A protocols use two regions to locate and move the peak of Flow-Set Fluorospheres to a specific channel: r r r _A3CL Flow-Set the AutoStandardization Flow-Set protocol for three-color immunofluorescence. This protocol creates the n3cl.qcs baseline file. r 3CL Three-Color a generic analysis protocol for a three-color immunofluorescence panel. It uses the n3cl.qcs baseline file. r _Q3CL trichrome _Q3CL CD45/4/3 protocols used to obtain QC statistical data. These protocols use the n3cl.qcs baseline file. A large capture region that locates the population at its current position. A small target region that identifies the location where to move the peak of the population. To ensure that the data is correct when acquired and that the Cytometer adjustments are valid, a check is performed on the HPCV of the Flow-Set Fluorospheres. If the CV check fails, 2-22

43 QUALITY CONTROL SETTING UP AUTOSTANDARDIZATION PROTOCOLS 2 an automatic PRIME is performed and acquisition restarts. If the CV check continues to fail, the _A protocol stops without continuing the panel. 1. Setup Screen tt Protocol File tt Select. The _Axyz protocol you want to use. [Okay]. 2. Edit the parameter user names if you want to change them to the parameters you are investigating. 3. Erase any parameters that you do not need. 2-23

44 QUALITY CONTROL SETTING UP AUTOSTANDARDIZATION PROTOCOLS 4. Check the histogram titled AUTOPRIME to ensure the parameters assigned to it are parameters you are investigating. If not: a. Assign parameters you are investigating to the Autoprime histogram. b. Setup Screen tt Run/ Regions. c. Gate the histogram on the population of interest. d. Recreate the rectilinear region and name it PRIME. 5. Edit the small target regions to place them in the peak position according to: r r r The instructions in the Flow-Set Fluorospheres package insert. OR The instructions in the reagent application s System Guide. OR Your laboratory s predetermined values A TARGET REGIONS 2-24

45 QUALITY CONTROL SETTING UP AUTOSTANDARDIZATION PROTOCOLS 2 6. Edit one of the larger region names on one of the single-parameter histograms to enable the CV check. The region name should be CV<n.nn where n.nn is an acceptable CV value for Flow-Set Fluorospheres. 7. Create a lot number region (rectilinear or amorphous) to save the lot number of the Flow-Set Fluorospheres with the run s voltage and gains values. The lot number, voltages, and gain settings are posted in the related nxyz.qcs file. 8. Name the region LOT#=ABCD1234, where ABCD1234 is the lot number of the Flow-Set Fluorospheres. Setup Screen tt Protocol. File tt Save As to save the protocol with a new name for use in your Autostandardization panel. Before running samples, you must load the new protocol into the system so the settings are used from the correct baseline file. Be sure to use _A and an xyz identifier as the first five characters of the protocol name. Setting Up _C Protocols CYTO-COMP Cells stained with CYTO-COMP Reagents or other compensation reagents and specific regions are used to automatically adjust the quad-stat color compensation regions. Compensation settings from the _C protocol are posted into the nxyz.qcs data file generated by the _Axyz protocol. 2-25

46 QUALITY CONTROL SETTING UP AUTOSTANDARDIZATION PROTOCOLS Each quad-stat region is named DIFF<n.nn for flagging purposes. The difference in the mean intensity of the positive population versus the negative population for X and Y is compared to the quad-stat region name. After compensation adjustment, if the difference is less than or equal to the region label then the message COMP PASS is displayed above the respective histogram. If the difference is more than the region label then the message COMP FAIL is displayed above the respective histogram. If COMP FAIL appears: 1) prime the system, 2) check the order of the sample tubes, 3) prepare the samples again, and 4) rerun the samples. 1. Protocol tt Select. The _C protocol of your choice, such as _C1 FITC/RD1 or _C2 RD1/ECD. [Okay]. 2. Edit the parameter user names if you want to change them to the parameters you are investigating. 3. Erase any parameters that you do not need. 2-26

47 QUALITY CONTROL SETTING UP AUTOSTANDARDIZATION PROTOCOLS 2 4. Edit the quad-stat region names to the acceptable difference between the positive and negative populations X and Y. 5. Check the histogram titled AUTOPRIME to ensure the parameters assigned to it are parameters you are investigating. If not: a. Assign parameters you are investigating to the Autoprime histogram. b. Setup Screen tt Run/ Regions. c. Gate the histogram on the population of interest. d. Recreate the rectilinear region and name it PRIME. 2-27

48 QUALITY CONTROL SETTING UP AUTOSTANDARDIZATION PROTOCOLS 6. Create a lot number region (rectilinear or amorphous) to save the lot number of the CYTO-COMP reagent with the run s compensation values. The lot number is posted with the compensation values in the nxyz.qcs file created. Name the region LOT#=ABCD1234, where ABCD1234 is the lot number of the CYTO-COMP Reagent or Reagent Kit. If using multiple _C protocols in the panel, you only need to create a lot number region the last _C protocol and enter the lot number of the CYTO-COMP Reagent Kit. 7. Setup Screen tt Protocol. File tt Save As to save the protocol with a new name for use in your Autostandardization panel. Before running samples, you must load the new protocol into the system so the settings are used from the correct baseline file. Be sure to use _C as the first two characters of the protocol name. You do not need the xyz prefix in the name of the _C protocol. Setting Up _QxyzABCD Protocols The xyz prefix in the _QxyzABCD protocol name is a three-letter identifier that you enter to match the corresponding _Axyz protocol. This xyz prefix allows the system to retrieve the gains and voltage settings from the matching QC baseline file (nxyz.qcs) and link them to the nxyzabcd.qcc file generated by the _Q protocol. The software saves the statistical QC data for all regions with QC in their names from the _QxyzABCD protocol into the nxyzabcd.qcc file. Multiple _Qxyz protocols can be set up. Use different ABCD identifiers after the _Qxyz portion of the protocol name to distinguish each _QxyzABCD protocol. For example, these are for a three-color panel on a 3PMT system: _A3CL Flow-Set _C1FITC/RD1 CYTO-COMP 2-28

49 QUALITY CONTROL SETTING UP AUTOSTANDARDIZATION PROTOCOLS 2 _C3 RD1/PC5 CYTO-COMP [3CL-3PMT] _Q3CL 0 45-FITC/G1-RD1/G1-PC5 [3] _Q3CL 2 45-FITC/4-RD1/3-PC5 [3] _Q3CL 4 45-FITC/8-RD1/3-PC5 [3] _Q3CL 6 45-FITC/19-RD1/3-PC5 [3] _Q3CL 8 45-FITC/56-RD1/3-PC5 [3] For multicolor immunofluorescence, use Immuno-Trol Cells or CYTO-TROL Control Cells to acquire statistical QC data. 1. Protocol tt Select. a _QxyzABCD protocol. [Okay]. 2. Edit the parameter user names if you want to change them to the parameters you are investigating. 2-29

50 QUALITY CONTROL SETTING UP AUTOSTANDARDIZATION PROTOCOLS 3. Assign the parameters to the histograms. 4. Create or edit analysis regions appropriate for the control material you are using. 5. Create any rectilinear, linear, or amorphous QC regions whose statistics you want to monitor. Be sure to include QC in the region name. 6. Create a lot number region (rectilinear or amorphous) to save the lot number of the Immuno-Trol or CYTO-TROL Cells with the run s statistical QC data. The lot number and statistical data are posted in the nxyzabcd.qcc file. Name the region LOT#=ABCD1234, where ABCD1234 is the lot number of the Immuno-Trol or CYTO-TROL Cells. 2-30

51 QUALITY CONTROL CREATING AUTOSTANDARDIZATION PANELS 2 7. File» Save As to save the protocol with a new name for use in your AutoStandardization panel. Before running samples, you must load the new protocol into the system so the settings are used from the correct baseline file. Repeat as needed to define as many _QxyzABCD protocols as needed. Be sure to use _Q and the identifier xyz as the first five characters of the name. Follow these five characters with an ABCD identifier to distinguish among multiple _QxyzABCD protocols. 2.7 CREATING AUTOSTANDARDIZATION PANELS Creating a Flow-Check Fluorospheres Panel Create an Autostandardization panel with the _Qxyz Flow-Check protocol you have set up to check your instrument. 1. Protocol tt Select. The _Qxyz Flow-Check protocol to use in the panel. 2. [Okay]. Setup Screen tt Panels. 2-31

52 QUALITY CONTROL CREATING AUTOSTANDARDIZATION PANELS 3. File tt Save As to save the panel with a new name. 4. Type a name for the panel and press Û. Creating an Immunophenotyping Panel Create an Autostandardization panel with the _A, _C, and _Q protocols you have set up for the type of samples you are investigating. 1. Protocol tt Select. The protocols to use in the panel. [Okay]. 2. Setup Screen tt Panels. Assign: r r r _A protocol as the Primary protocol _C protocols as Secondary protocols _Q protocols as Secondary or Control protocols. 2-32

53 QUALITY CONTROL CREATING AUTOSTANDARDIZATION PANELS 2 3. File tt Save As to save the panel with a new name. Type a name for the panel and press Û. 4. Setup Screen tt Run to return to the Acquisition Run screen. 2-33

54 QUALITY CONTROL CREATING AUTOSTANDARDIZATION PANELS 2-34

55 3CREATING A WORKLIST OVERVIEW Create a Worklist to: r Specify the order of specimen and sample analysis. r Input specimen information for up to 32 tubes. A worklist can consist of a panel or panels that contain protocols to be run consecutively. r Minimum requirements for a Worklist are one specimen ID and one panel. r A single protocol can only be used in a worklist by saving that protocol as a panel. r Autostandardization panels or _A, _C, or _Q protocols cannot be run in a Worklist. To create a worklist: r Enter the Specimen ID. r Enter the Patient Name and Patient ID (these are automatically entered if they are in the database). r Save the Specimen and Patient information to the database. r Assign the Panels to use for each specimen. r Assign the tube IDs. r Save the Worklist file. Note: You can leave blank lines between entries on a Worklist. You do not need to leave a corresponding space in the XL-MCL flow cytometer carousel. Tubes in the carousel are processed in the order of the panels listed ignoring blank lines. 3-1

56 CREATING A WORKLIST ENTERING SPECIMEN ID 3.2 ENTERING SPECIMEN ID Specimen ID can be entered into the Worklist by: r r r Manually entering the information using the keyboard and mouse. Scanning the bar-code label on the specimen tube using the optional hand-held bar-code scanner. Scanning the bar-code label on the test request form using the optional hand-held bar-code scanner. Using the Keyboard 1. An empty SPECIMEN ID field on the Worklist screen. The software prompts you for the specimen ID. 2. Type the SPECIMEN ID and press Û Using the Hand-Held Bar-Code Scanner 1. An empty SPECIMEN ID field on the Worklist screen. The software prompts you for the specimen ID. 2. Hold the specimen tube or test request form so that the bar-code label is facing the scanner and is about 1 in. away from it. Press the trigger on the scanner A 3-2

57 CREATING A WORKLIST ENTERING PATIENT INFORMATION ENTERING PATIENT INFORMATION To print the Patient Name on your patient reports, you have to enter a Patient Name and Patient ID on the worklist by entering them in the Patient pop-up box and saving them to the database. You can enter other patient demographic information on this screen and save the information as a record in the system database to be used for patient reports. Refer to the Data Management manual for detailed information on using all the fields in this screen for patient reports and creating patient report templates in the Reports application. Go to the Patient Pop-Up Box Patient Name field on the Worklist screen. The software sends you to the Patient pop-up box. Note: Changing the Specimen ID clears all the other fields. If you need to change the Specimen ID, do so first and then update the other fields. Enter the Patient Name 1. Last Name on the Patient pop-up box. 2. Type the name and press Û. 3-3

58 CREATING A WORKLIST ENTERING PATIENT INFORMATION 3. At First Name on the Patient pop-up box, type the name and press Û. Note: If the patient is 100 years old or older, you might want to add the age with the name entry as a comment. See subheading Enter DOB below. Enter the Patient ID 1. Patient ID on the Patient pop-up box. 2. Type the Patient ID and press Û. Enter the DOB (Date of Birth) 1. DOB on the Patient pop-up box. 2. Type the patient s date of birth and press Û. Note: Since the year portion has only two digits, if the year entered is: r r Equal to or less than the current year, the system assumes the year is in the current century. (Example: If it is currently 1998 and you enter a DOB of 21JUN91, the system assumes the year is 1991.) Greater than the current year, the system assumes the previous century. (Example: If it is currently 1998 and you enter a DOB of 13JUL99, the system assumes the year is 1899.) 3-4

59 CREATING A WORKLIST SAVING INFORMATION TO THE DATABASE 3 Enter Additional Information 1. Any other field where you want to enter information. 2. Type the information and press Û. If you enter any hematology information, you must include a WBC or RBC. Note: Do not use a comma when entering large numbers, such as for platelets. 3. Press È after you complete all the prompt boxes you need. 3.4 SAVING INFORMATION TO THE DATABASE [Save Record] and then [Done] to save the information to the system database and go back to the worklist. Retrieving Records You can use Retrieve Record and a Specimen ID to retrieve a specimen record from the database. If the Specimen ID has been entered on the worklist: 1. [Spec Info] and any line on the Worklist screen. 2. The software sends you to the Patient pop-up box and searches the database for the specimen record. [Done] to go back to the worklist. The information from the retrieved record is entered into the worklist. 3-5

60 CREATING A WORKLIST ASSIGNING PANELS TO A WORKLIST If the Specimen ID has NOT been entered on the worklist: 1. An empty PATIENT NAME field or [Spec Info] and any line on the Worklist screen. 2. Specimen ID on the Patient pop-up box. 3. Type the Specimen ID number you want to retrieve and press Û [Retrieve Record] [Done] to go back to the worklist and the information from the retrieved record is entered on the worklist. 3.5 ASSIGNING PANELS TO A WORKLIST 1. Setup Screen tt Worklist. 2. The Panel column. Note: The panels and their protocols must all be in either the local or off-line directories as specified in the Utilities Configuration screen. 3. The first panel for the worklist. 4. [Okay]. Note: Do not use Autostandardization protocols or panels in a worklist. 5. The desired panel. [Okay]. 3-6

61 CREATING A WORKLIST ASSIGNING PANELS TO A WORKLIST 3 The panel and protocols are assigned to the worklist. Now you can: r r r Enter Specimen ID, Patient Name and Patient ID if you have finished assigning panels. OR Repeat steps 2 through 4 to assign more panels. OR Use the [Copy] button to copy the panels. See instructions below for copying panels. Copying Panels on the Worklist Screen 1. [Copy]. 2. The panels you want to copy (you can copy more than one panel). Note: Only panels and protocols are copied. Specimen ID, Patient Name, and Tube ID are not copied. 3. An empty line in the Panel column. The panel is copied starting on the line you selected. 4. Repeat steps 2 and 3 to continue copying the panel to the worklist. [Copy] when finished. 3-7

62 CREATING A WORKLIST ASSIGNING PANELS TO A WORKLIST Moving Lines on the Worklist Screen [Move]. The line you want to move An empty line. The whole line is moved, starting on the line you selected. Repeat steps 2 and 3 to continue moving lines on the Worklist. [Move] when finished. Erasing Worklist Information To erase an entire worklist: 1. [New]. 2. Press Y and then press Û. 3-8

63 CREATING A WORKLIST ASSIGNING WORKLIST TUBE ID 3 To erase a panel from the worklist: 1. [Erase]. 2. The panel you want to erase. 3. Press Y and then press Û. 3.6 ASSIGNING WORKLIST TUBE ID Assigning the Tube ID is optional. You can: r r Use the keyboard to type in a Tube ID. Use the Worklist screen [Assign BC] (bar-code) button to have the system assign a bar-code Tube ID to some or all of the tubes. r Use the hand-held bar-code reader to scan in the Tube ID from a bar-code label on the tube. r Choose not to use the Tube ID. If you use Tube ID, you can: r Print bar-code labels from the Worklist screen if you have the optional bar-code label printer. OR r Use your laboratory s preprinted bar-code labels (see the heading Bar-Code Specifications in the Reference manual for additional information). When running the Worklist, the system compares the Tube ID to the bar-code labels on the tubes and alerts you if they do not match. 3-9

64 CREATING A WORKLIST ASSIGNING WORKLIST TUBE ID Before Assigning Tube ID Bar-Codes Before assigning Tube ID you may need to determine what characters to use, 16 numeric or 8 alphanumeric. 1. Applications tt Utilities Assigning the Bar Codes The box next to Worklist Barcodes to change it to the type of characters you want to use. Applications tt Acquisition. Setup Screen tt Worklist. Using the Keyboard or Hand Held Scanner 1. A field in the Tube ID column. 2. Type or scan in using hand held scanner up to 16 numeric or 8 alphanumeric characters and press Û. 3-10

65 CREATING A WORKLIST ASSIGNING WORKLIST TUBE ID 3 Using the Assign BC Button To have the system assign the Tube ID: 1. [ Assign BC]. A field in the Tube ID column to assign Tube IDs one tube at a time. OR Erasing Tube ID The Tube ID header to assign Tube IDs to all of the tubes. The bar-code the system assigns consists of the date, time, and line number on the worklist. To erase all the Tube IDs: 1. [Erase]. 2. The Tube ID header. To erase the Tube ID for a single tube: [Erase]. The line for the tube ID you want to erase. 3-11

66 CREATING A WORKLIST SAVING THE WORKLIST 3.7 SAVING THE WORKLIST File tt Save As. Type a name for the worklist and press Û. 3-12

67 4RUNNING SAMPLES BEFORE RUNNING SAMPLES 1. Check that the daily startup and quality control procedures were done (Chapters 1 and 2). 2. Turn the [Save Cyto] button on (green) if you want changes to the Cytometer settings and regions to be updated in the protocol at the end of a sample cycle. If not, turn it off (blue). 3. Check that the [Output Options] are set correctly for the samples you are running. 4. Check drive C to ensure sufficient space is available for the samples to be processed. 5. Check that there is enough sheath fluid and printer paper for the samples to be analyzed. 6. Check that the waste container is empty. If the waste container fills completely, acquisition stops. You must empty the waste container to continue. 4.2 RUNNING A SAMPLE IN THE MANUAL MODE (XL OR XL-MCL FLOW CYTOMETER) 1. Select a worklist, protocol, or panel. a. Application tt Acquisition. b. To select a worklist: Worklist. Type a name or just press Û. The desired worklist. [Okay]. c. To select a protocol: Protocol tt Select. The desired protocol. [Okay]. d. To select a panel: Panel. The desired panel. [Okay]. 4-1

68 RUNNING SAMPLES RUNNING A SAMPLE IN THE MANUAL MODE (XL OR XL-MCL FLOW CYTOMETER) 2. Check that the Cytometer status message reads Insert Sample Tube. CAUTION Possible flow cell damage. To avoid clogging the sample probe, sample tubing or flow cell, ensure that 12 mm x 75 mm test tubes are free of debris before you use them. 3. Put the tube on the sample stage. Use the Acquisition Run Screen Cytometer Control Window to adjust settings during data acquisition if necessary. 1 During the sample cycle, the indicator in the Cytometer RUN button is orange and the following series of Cytometer status messages appears. 2 Sample Tube Detected Acquiring Data Sample Wash Cycle In Process Please Wait... Processing Insert Sample Tube 4. When data acquisition ends: A a. When you are not running a worklist you are prompted for the specimen ID. When the ID is entered, it appears in the upper right of the screen. b. Remove the tube A 4-2

69 RUNNING SAMPLES RUNNING SAMPLES IN THE AUTO MODE (XL-MCL FLOW CYTOMETER ONLY) 4 c. The Output Cycle window shows the progress of printing and file saving. Data is saved and printed as specified in the Output Options. Note: If the boxes appear red instead of green, the destination disk is not present or there is not enough available disk space. 4.3 RUNNING SAMPLES IN THE AUTO MODE (XL-MCL FLOW CYTOMETER ONLY) 1. Select a worklist: a. Application tt Acquisition. b. Worklist. Type a name or just press Û. c. The desired worklist. 2. d. [Okay]. You can also run a protocol or panel in the Auto Mode. See heading 4.2, Running a Sample in the Manual Mode, for instructions on selecting a protocol or panel. Check that the Cytometer status message reads Insert Sample Tube. CAUTION Possible flow cell damage. To avoid clogging the sample probe, sample tubing or flow cell, ensure that 12 mm x 75 mm test tubes are free of debris before you use them. 4-3

70 RUNNING SAMPLES RUNNING SAMPLES IN THE AUTO MODE (XL-MCL FLOW CYTOMETER ONLY) 3. Put the loaded carousel in the MCL. Carousel is in home position when handle points toward back Press the AUTO button (or select [Run] on the Aquisition screen). a. The system prompts Start a new carousel? Y/N. b. If you are not running a worklist, you are prompted Do you want to enter specimen ID? Y/N IMPORTANT Even if you respond Yes, you must not pause for more than 10 seconds during data entry, or the system continues the analysis, using the carousel number and tube number as the specimen ID. During the sample cycle, the indicator in the Cytometer RUN button is orange and the following series of Cytometer status messages appears. Sample Tube Detected...MCL Acquiring Data MCL Sample Wash Cycle In Process MCL Insert Sample Tube MCL Note: While running a worklist, if the Tube ID in the worklist does not match the bar code on the tube, a system message appears. After you acknowledge the message, the mismatched tube is aborted and the processing stops for the rest of the carousel. 4-4

71 RUNNING SAMPLES RUNNING SAMPLES IN THE AUTO MODE (XL-MCL FLOW CYTOMETER ONLY) 4 When the MCL acquisition is completed: r r r r The AUTO button indicator turns off. The Output Cycle window shows the progress of printing and file saving. Note: If the boxes appear red instead of green, the destination disk is not present or there is not enough available disk space. The carousel report is printed and displayed if it was enabled on the Output Options box. r To return to the Acquisition Run screen, press any key. 5. Remove the carousel and close the MCL A If the Printer runs out of paper while running a carousel, the sample data is stored in a listmode file, even if not specified to do so in the protocol. If a system error is detected while the last sample in a carousel is being analyzed, the carousel summary is not printed. 4-5

72 RUNNING SAMPLES RUNNING SAMPLES IN THE AUTO MODE (XL-MCL FLOW CYTOMETER ONLY) To print the carousel summary: [Alt Save]. [Print MCL Summary]. [OK]. To print the individual runtime histograms in either case, select: [Alt Save]. [Print MCL Report]. [OK]. Interrupting the Auto Mode Cycle You can interrupt the Auto mode cycle to run samples manually (for example, to run a Stat sample). Pressing AUTO at the sample stage or clicking [Abort] on the Acquisition Run screen interrupts the Auto mode cycle. How you interrupt the Auto mode cycle depends on whether or not you want the MCL to finish the cycle for the current sample tube. Finishing the Tube Press the AUTO button on the Cytometer. The indicator in the AUTO button turns off and the Cytometer returns to the Manual mode after acquisition of the current tube is complete A Aborting the Tube Selecting [Abort] stops acquisition without saving, printing, or exporting data as specified in the protocol. The sample or run results are discarded and the output options are not applied. If you edit a panel or worklist after selecting [Abort], you cannot resume the Auto mode cycle with the panel or worklist. You must run the panel or worklist over again from the beginning. 4-6

73 RUNNING SAMPLES RUNNING SAMPLES IN THE AUTO MODE (XL-MCL FLOW CYTOMETER ONLY) 4 1. [Abort]. 2. Press Y and then Û to discard the entire carousel. The indicator in the AUTO button turns off and the Cytometer returns to the Manual mode. Press n and then Û to discard the current tube only and continue with the rest of the carousel. Resuming the Auto Mode Cycle After You Interrupt a Panel To resume the Auto mode cycle after interrupting it. 1. Press the AUTO button (or select [Run] on the Aquisition screen). The indicator in the button glows green and you are prompted: Are you starting a new carousel? Y/N. Respond N for no. The Auto mode cycle resumes A 2. How Auto mode resumes depends upon whether or not you finished the tube before interrupting the panel and how you used the software since interrupting the panel. a. If you displayed the worklist, panel, or protocol screen (from the Setup Screen menu), or selected another worklist, panel, or protocol and selected [Okay], or ran a sample tube in the manual mode after you interrupted the Auto mode cycle, then The Auto mode cycle begins the panel over again from the Primary. This ensures that the correct Cytometer settings and regions are still passed on. b. If you finished the tube when you interrupted the cycle, and have not selected another worklist, panel, or protocol and have not displayed the worklist, panel, or protocol screen, or have not run a sample tube in the manual mode after interrupting the panel, then The Auto cycle continues with the next tube in the carousel. 4-7

74 RUNNING SAMPLES DATA ACQUISITION FUNCTIONS c. If you aborted the tube when you interrupted the cycle and have not selected another worklist, panel, or protocol and have not displayed the worklist, panel, or protocol screen, or have not run a sample tube in the manual mode, then The Cytometer resumes the cycle with the aborted tube in the current panel. What the Completion Codes in the Carousel Summary Mean Final column shows one of the following completion codes: P Primary C Control S Secondary * Last in panel A Abort test E Total events limit F File size limit H Cytometer problem M Manual stop 'n' nth hist stop count O Out of target abort (prime unsuccessful) R Min counter satisfied T Time stop V Volume stop X Memory limit Y Fluid level stop Z Worklist Tube ID & Barcode do NOT match W Patient Info could NOT be printed. G Autogating Failed A listing of the codes is included at the bottom of the printed report B 4.4 DATA ACQUISITION FUNCTIONS PRIME Button If no data points are displayed on the screen within a few seconds of the Cytometer status message Acquiring Data, you may need to perform a Prime cycle. r r r Press the PRIME button. Wait for the cycle to finish. Restart the acquisition with the [Restart] button or Ñ B 4-8

75 RUNNING SAMPLES DATA ACQUISITION FUNCTIONS 4 Cytometer Setting Adjustments If you have followed the Quality Control procedures in Chapter 2, you do not need to make further changes for cell surface marker analysis. If you are running an unusual sample type, you may need to adjust Cytometer settings to see the populations of interest more clearly. You can use the Cytometer Control Window on the Acquisition Run screen to make changes while the system acquires data. Remember to set [Save Cyto] to on (green) if you want to update the protocol. Using Fast Set Use [Fast Set] to move a population and automatically update high voltage settings. While the system is acquiring data: 1. The single-parameter or dual-parameter histogram with the population you want to move. 2. [Display Options] to display the Signal box in the Cytometer Control Window. 3. [Fast Set]. 4. The population and drag it to a new position on the histogram. When you release the mouse button, the population is anchored at the new position and the high voltage settings are updated. 4-9

76 RUNNING SAMPLES DATA ACQUISITION FUNCTIONS Using Fast Comp Use [Fast Comp] to move a population and automatically update compensation settings. While the system is acquiring data: 1. The single-parameter or dual-parameter histogram with the population you want to move. 2. [Display Options] to display the Compensation box in the Cytometer Control Window. 3. [Fast Comp]. 4. The population and drag it to a new position on the histogram. When you release the mouse button, the population is anchored at the new position and the compensation settings are updated. Restarting the Cytometer after Making Changes IMPORTANT The data you have just acquired is invalidated if you change any of the settings except flow rate. Select [Restart] or press Ñ to clear the stored data and start acquiring again. Acquisition Buttons and Keys You can override stop counters, if you wish, with the screen acquisition buttons or their keyboard equivalents. [Run] Ñ [Stop] Ò [Pause] Ó [Abort] Ô 4-10

77 RUNNING SAMPLES DATA ACQUISITION FUNCTIONS 4 Editing Regions During Acquisition You can review region statistics and edit regions on any histogram during acquisition without using the [Region] button. To edit a region during acquisition: The histogram containing the region and edit the regions as required. If you are unable to edit a region, you may have changed Cytosettings without restarting. [Restart] or press Ñ and try again. When you edit a region during acquisition, the software automatically replays the stored data and then continues data acquisition. Creating, Erasing, Copying, or Redrawing Regions During Acquisition To create, erase, copy, or redraw regions during acquisition: [Pause]. Create, erase, copy, or redraw regions as explained in the Using Regions chapter of the Getting Started manual. [Cont]. 4-11

78 RUNNING SAMPLES DATA ACQUISITION FUNCTIONS Alt Save Button You can also print or save results that were not designated to be printed or saved in the protocol: [Alt Save]. The items you want to save or print. [Okay]. Histogram Display Keys Use Ì to rescale one- or two-parameter histograms. Note: Í, Î, Ï, and Ð apply only to two-parameter histograms. Press Í to change to contour plots. Press again to change to color contour plots. Press Î to change to dot density plots. Press again to change to color dot density plots. Press Ï to view isometric display. Press again to change the isometric viewing angle. Press Ð to turn projection display on or off. 4-12

79 RUNNING SAMPLES DATA ACQUISITION FUNCTIONS 4 Isometric F 5 F 6 F 7 F 8 Density Projection A 4-13

80 RUNNING SAMPLES DATA ACQUISITION FUNCTIONS 4-14

81 5DATA ANALYSIS LISTMODE ANALYSIS To replay files in the Listmode Application: r r r r r r r Select files for replay. Reorder and save the list of files to a List Queue, if desired. Choose output options. Use Runtime Protocol to replay files with the acquisition protocol exactly as it was acquired. Use New Pnl/Pro to replay files with a different panel and a different protocol and to export data. Replay the files. Assign, create, or edit regions. The Listmode application has two screens that are not in the Acquisition application. r The File Info screen displays information about the listmode file. r The List Queue screen is used to order listmode data files for batch analysis. 5-1

82 DATA ANALYSIS REPLAYING INDIVIDUAL LISTMODE FILES 5.2 REPLAYING INDIVIDUAL LISTMODE FILES 1. Selecting the Files To select individual listmode files for replay: Applications tt Listmode. File tt Select. Listmode files to replay. [Okay]. When you select more than one listmode file to analyze, you have created a List Queue. 2. Reordering the List Queue a. To change the order the files are analyzed: Setup tt List Queue. [Next]. The first file to analyze. [Order]. Remaining files in order. [Order]. 5-2

83 DATA ANALYSIS REPLAYING INDIVIDUAL LISTMODE FILES 5 b. To delete listmode files from the queue: 3. Saving the List Queue Create, to change it to Erase. Listmode files to delete. The listmode file remains on disk; this just deletes the file from the queue. Note: If you need to add files to the queue, you must reselect the files. See step 1 above. If you need to analyze the same set of files later, save them to a List Queue. If not, skip to step 4. File» Save As. Type a name for the list of files and press Û. Setup» Analysis. 4. Choosing Output Options To choose which output options to enable: [Output Options]. The items you want to save, print, or enable. [Ok]. 5-3

84 DATA ANALYSIS REPLAYING INDIVIDUAL LISTMODE FILES 5. Choosing the Replay Mode The software uses the Runtime Protocol (histograms and regions from the acquisition protocol) as the default mode. Proceed to step 6, Replaying the Files, if you want to use the Runtime Protocol. If you want to replay files with a panel or a different protocol, change to the New Pnl/Pro mode. Note: The new panel or protocol must already exist before selecting [New Pnl/Pro]. You cannot use this button to create new panels or protocols. [Reset Queue]. [Runtime Protocol] to change the mode to [New Pnl/Pro]. Panel or Protocol tt Select. The panel or protocol you want to use. [Okay]. The system uses as much of the protocol as is consistent with the data file. If the protocol has histograms with parameters that are not stored in the listmode file, then those histograms are not shown. When you replay a listmode file in Listmode with a new protocol or panel, the Cytometer settings (signals and compensation values) for the new protocol appear in the Listmode File Info screen but do not affect the data. The original Cytometer settings saved with the protocol used when the data was acquired are also used when the protocol is replayed. 6. Replaying The Files [Next File] steps through the file list without replaying data. [Play List] replays the current file in the queue. [Play Next] replays the next file in the queue. [Batch] replays all the files in the queue without stopping after each one. 5-4

85 DATA ANALYSIS REPLAYING LISTMODE FILES AS A PANEL 5 7. Editing Regions IMPORTANT Misleading results can be reported if you edit, erase, or reassign a gating equation without replaying the data. If you edit, erase, or reassign a gating equation, you must replay the data to update statistics. To edit or reassign regions: [Region]. Edit the regions as needed. Reassign regions as needed. See the heading Assigning Regions in the Getting Started manual. [Region]. Play List] to update the histogram display and statistics with any new gate or region assignments. 5.3 REPLAYING LISTMODE FILES AS A PANEL 1. Selecting the Application Applications tt Listmode. 2. Choosing the Replay Mode To replay the panel, change to the New Pnl/Pro mode. When analyzing a series of specimens in Listmode using the same New Panel/Protocol for each specimen, adjust the gate for each specimen if these gates were adjusted manually during acquisition. Note: The new panel must already exist before selecting [NEW PNL/PRO]. You cannot use this button to create new panels or protocols. [RUNTIME PROTOCOL] to change the mode to [NEW PNL/PRO]. 5-5

86 DATA ANALYSIS REPLAYING LISTMODE FILES AS A PANEL 3. Selecting the Panel Panel tt Select. The panel you want to use. [Okay]. Review the protocols listed in the panel. The system uses as much of the protocol as is consistent with the data file. If the protocol has histograms with parameters that are not stored in the listmode file, then those histograms are not shown. When you replay a listmode file in Listmode with a new protocol or panel, the Cytometer settings (signals and compensation values) for the new protocol appear in the Listmode File Info screen but do not affect the data. 4. Selecting the Files To select individual listmode files for replay: Applications tt Listmode. File tt Select. Listmode files to replay. [Okay]. When you select more than one listmode file to analyze, you have created a List Queue. 5-6

87 DATA ANALYSIS REPLAYING LISTMODE FILES AS A PANEL 5 5. Reordering the List Queue a. To change the order the files are analyzed: Setup tt List Queue. [Next]. The first file to analyze. [Order]. Remaining files in order. [Order]. b. To delete listmode files from the queue: 6. Saving the List Queue Create, to change it to Erase. Listmode files to delete. The listmode file remains on disk; this just deletes the file from the queue. Note: If you need to add files to the queue, you must reselect the files. See step 4 above. If you need to analyze the same set of files later, save them to a List Queue. If not, skip to step 7. File tt Save As. Type a name for the list of files and press Û. Setup tt Analysis. 5-7

88 DATA ANALYSIS REPLAYING LISTMODE FILES AS A PANEL 7. Choosing Output Options To choose which output options to enable: [Output Options]. The items you want to save, print, or enable. [Ok]. 8. Replaying The Files [Next File] steps through the file list without replaying data. [Play List] replays the current file in the queue. [Play Next] replays the next file in the queue. [Batch] replays all the files in the queue without stopping after each one. 5-8

89 DATA ANALYSIS LISTMODE BATCH ANALYSIS 5 9. Editing Regions IMPORTANT Misleading results can be reported if you edit, erase, or reassign a gating equation without replaying the data. If you edit, erase, or reassign a gating equation, you must replay the data to update statistics. [Region]. Edit the regions as needed. Reassign regions as needed. See the heading Assigning Regions in the Getting Started manual. [Region]. [Play List] to update the histogram display and statistics with any new gate or region assignments. [Play Next] and edit the regions for each remaining file. 5.4 LISTMODE BATCH ANALYSIS Listmode batch replay can be used to replay listmode files with a panel or protocol. You can replay an existing Listmode Queue or create one. In batch replay, the system replays the listmode files with the protocols in order, starting with the first file in each queue. If you have more listmode files than protocols, the system repeats the protocol queue until all of the listmode files are replayed. 1. Creating a Listmode Queue Perform steps 1 through 3 in heading 5.2, Replaying Individual Listmode Files, to create a Listmode Queue and then skip to step 3. If you have already saved the Listmode Queue you want to analyze, skip to step 2, Loading a Listmode Queue. 2. Loading a Listmode Queue To recall an arrangement of listmode files analyzed previously: Setup tt List Queue. File tt Select. Listmode queue name. [Okay]. 5-9

90 DATA ANALYSIS LISTMODE BATCH ANALYSIS 3. Choosing Output Options To choose which output options to enable: [Output Options]. The items you want to save, print, or enable under the Listmode column. [Ok]. 4. Choosing the Replay Mode The software uses the Runtime Protocol (histograms and regions from the acquisition protocol) as the default mode. Proceed to step 6, Replaying Batch, if you want to use the Runtime Protocol. If you want to replay files with a panel or a different protocol, change to the New Pnl/Pro mode. Note: The new panel or protocol must already exist before selecting [New Pnl/Pro]. You cannot use this button to create new panels or protocols. [Runtime Protocol] to change the mode to [New Pnl/Pro]. Panel or Protocol tt Select. Panel or protocol to use. [Okay]. The system uses as much of the protocol as is consistent with the data file. If the protocol has histograms with parameters that are not stored in the listmode file, then those histograms are not shown. 5. Adjusting Regions [Play Next] to load and replay the first file in the queue. Edit the regions in the new protocol so that they are around the populations of interest. 5-10

91 DATA ANALYSIS HISTOGRAM ANALYSIS WITH MULTIGRAPH 5 6. Replaying Batch Replay tt Batch. Histograms are displayed, printed, and saved as specified in the protocol and the output options. The system replays all the listmode files until the List Queue is finished. Note: If the error message [Error! Printer Not Ready. Waiting For Printer!] is displayed while replaying a batch, check that the Printer has paper. Otherwise the error message means the Printer is busy. 7. Stopping Batch Listmode Replay a. Press È and hold it down for a few seconds. The batch stops at the end of the current file being replayed. The [BATCH] button changes color. b. To restart the batch on the next file in the queue, press [BATCH]. 5.5 HISTOGRAM ANALYSIS WITH MULTIGRAPH 1. Selecting the Files Applications tt Multigraph. File tt Select. [Clear All]. Histograms for analysis. [Okay]. When you select more than one histogram file to analyze, you have created a Hist Queue. The software reassigns new region ID letters to the histogram regions as they are loaded to avoid duplicate region ID letters. The region labels are the same as they were in Acquisition. 5-11

92 DATA ANALYSIS HISTOGRAM ANALYSIS WITH MULTIGRAPH 2. Reordering the Hist Queue a. Screen tt Hist Queue. [Next]. 3. Saving the Hist Queue First histogram to analyze. [Order]. Remaining histograms in order. [Order]. b. To delete histograms from the queue, Create to change it to Erase. Histograms to delete. The histogram file remains on disk; this just deletes it from the queue. Note: If you need to add files to the queue, you must reselect the files. See step 1 above. If you want to analyze this same set of histograms later, save the queue. Otherwise skip to step 4. File tt Save As. Type a name for the queue and press Û. 5-12

93 DATA ANALYSIS HISTOGRAM ANALYSIS WITH MULTIGRAPH 5 4. Printing and Saving in Multigraph When enabled, histograms automatically print whenever you select Next Set in Hist Mode or Next Test or Batch in Test Mode. [Save Disabled] or [Print Disabled] to change to [Save Enabled] or [Print Enabled] if you want to save or print histograms. Note: To print Multigraph overlays, the Color option must be turned ON in the Utilities Configuration screen. 5. Choosing the Analysis Mode Hist mode is the default and displays the histograms in groups of eight. Test mode allows you to display the histograms grouped by run number. Test mode also allows you to edit regions and then pass the new region on to the other histograms in the next run number group. [Hist Mode] to change to [Test Mode] if you want to display the histograms grouped by run number. IMPORTANT Misleading results can be reported if you edit or delete regions used for gating. Histogram displays do not reflect the changes made from when you edit or delete regions. Do not edit or delete regions used for gating. 5-13

94 DATA ANALYSIS HISTOGRAM ANALYSIS WITH MULTIGRAPH 6. Loading and Passing Regions The system default is Load Regions. Regions saved with the histograms are loaded with the histogram. When you are in Hist Mode, Load Regions is the only option. Note: Region statistics from histograms acquired on all of the older EPICS flow cytometers will not exactly match when analyzed using the SYSTEM II software. This is due to a difference in rounding conventions on older systems. In Test Mode, Pass Regions allows you to edit a region in Multigraph and pass that new region on to the other histograms you are analyzing. If you want to pass regions in Test Mode: [Load Regions] to change it to [Pass Regions]. 7. Editing Regions The histogram containing the region to edit. The region ID letter to edit. OR [Edit] to change it to erase or create. Create, edit, or erase analysis regions only. Press Ì through Ð to adjust histogram displays. Press Ë to print the results. [8 Hist] to display the other histograms or [Display Next] to display the next histogram. 8. Displaying Next Set and Next Test [Next Set] to display the next set of eight selected histograms, and then repeat step 3. [Next Test] to display the next set of histograms in Test Mode, and then repeat step

95 DATA ANALYSIS HISTOGRAM BATCH ANALYSIS HISTOGRAM BATCH ANALYSIS Batch analysis allows you to create or load a Histogram Queue and then let the system analyze all the histograms in the queue. 1. Creating a Hist Queue Perform steps 1 through 3 in heading 5.5, Histogram Analysis with Multigraph, to create a Histogram Queue and then skip to step 3 of this procedure. If you have already saved the Histogram Queue you want to analyze, skip to step 2, Loading a Histogram Queue. 2. Loading a Histogram Queue To recall an arrangement of histograms analyzed previously: Screen tt Hist Queue. File tt Select. The queue to use (select only one). [Okay]. 3. Printing and Saving in Multigraph When enabled, histograms are automatically printed whenever you select Next Test or Batch. [Save Disabled] or [Print Disabled] to change to [Save Enabled] or [Print Enabled] if you want to automatically save or print histograms. Note: To print Multigraph overlays, the Color option must be turned ON in the Utilities Configuration screen. 5-15

96 DATA ANALYSIS HISTOGRAM BATCH ANALYSIS 4. Choosing the Analysis Mode Batch is not available in Hist Mode. Change to Test Mode to use batch analysis. Test mode allows you to display the histograms grouped by run number. In Test Mode you can edit regions and then pass the new regions on to the other histograms in the run. [Hist Mode] to change to [Test Mode]. IMPORTANT Misleading results can be reported if you edit or delete regions used for gating. Histogram displays will not reflect the changes made from when you edit or delete regions. Do not edit or delete regions used for gating. 5. Loading and Passing Regions The system default is Load Regions. Regions saved with the histograms are loaded with the histogram. When you are in Hist Mode, Load Regions is the only option. In Test Mode, Pass Regions allows you to edit a region in Multigraph and pass that new region on to the other histograms you are analyzing. The Pass regions option is not available in Hist Mode. If you want to pass regions while in Test Mode: 6. Editing Regions [Load Regions] to change it to [Pass Regions]. The histogram containing the region to edit. The region to edit. OR [Edit] to change it to Erase or Create. Create, edit, or erase the analysis regions only. 5-16

97 DATA ANALYSIS HISTOGRAM DISPLAYS 5 7. Analyzing the Queue [Batch] to analyze all the selected histograms in Test Mode. 8. Stopping Batch Multigraph Replay a. Press È and hold it down for a few seconds. The batch stops at the end of the current file being replayed. The [BATCH] button changes color. b. To restart the batch on the next file in the queue, press [BATCH]. 5.7 HISTOGRAM DISPLAYS Displaying Single Overlay Histograms To display the single-parameter histograms superimposed on one another: Screen tt Single Overlay. The single-parameter histograms in the queue are listed in the Histograms box in blue. If you want to remove or add histograms: A run number in the Histogram box. [Remove] or [Add]. The color of the files listed in the Overlay Info box corresponds to the histogram display color. 5-17

98 DATA ANALYSIS HISTOGRAM DISPLAYS Dual Overlay Histogram Displays To display the dual-parameter histograms superimposed on one another: Screen tt Dual Overlay. The dual-parameter histograms in the queue are listed in the Histograms box in blue. If you want to remove or add histograms: A run number in the Histogram box. [Remove] or [Add]. The color of the files listed in the Overlay Info box correspond to the histogram display color. 5-18

99 DATA ANALYSIS HISTOGRAM DISPLAYS 5 Hist Data Screen The Hist Data screen displays histogram information that was saved with the histogram when it was acquired. To display the Hist Data screen: Screen tt Hist Data. 5-19

100 DATA ANALYSIS HISTOGRAM DISPLAYS 5-20

101 7REVIEWING QC DATA OVERVIEW The quality control data obtained from running _A, _C, and _Q protocols (usually run in Autostandization panels) is placed into the *.QCS and *.QCC files of the related protocols. The QC data can be: r r r Accessed by either loading a protocol and its QC template at the QC Template screen or by manually selecting the individual QC files at the Levey-Jennings or QC Data Table screens. Viewed in Data Table and Levey-Jennings graph format. Visually reviewed for trends, shifts, outliers, and violations of your laboratory s QC guidelines. r Checked by the system against Westgard rules. r Compared to a reference file. Refer to the Data Management manual for detailed information on creating and defining QC templates, Levey-Jennings graphs, and Data Tables. Relationship of Protocols to QC Templates and QC Data Files To load or create a QC template, you must first select the related protocol at the QC Template screen. The QC template is stored in the same directory as its releated protocol. When the QC template is loaded or created at the QC Template screen, you can then view the QC data files (*.QCS and *.QCC) linked to the protocol at the Levey-Jennings and QC Data Table screens. The QC data available for viewing when the QC template is loaded depends upon the protocol selected. Copying, moving, or deleting any protocol from any screen where these functions are available, copies, moves, or deletes its corresponding QC template file. Protocol Selected _Axyz _C _QxyzABCD QC Data Files Available Through the QC Template Corresponding nxyz.qcs file with voltages, gains, and compenstion values. nxyz.qcs file where the compensation values from this _C protocol were posted. Corresponding nxyzabcd.qcc file with the statistical data for all QC regions defined for the protocol and, if it exists, the related nxyz.qcs file and its voltages, gains, and compenstion values data. 7.2 LOADING A QC TEMPLATE Overview You load a QC template by selecting a _A, _C, or _Q protocol (that was run as an individual protocol or in an Autostandardization panel) at the QC Template screen. This QC template allows you to access the QC data you want from the protocol s corresponding *.QCS or *.QCC data files. 7-1

102 REVIEWING QC DATA LOADING A QC TEMPLATE Note: If no QC data files appear after you load a QC template that should contain QC data files, check if the instrument node identifier was changed. If the node was changed you can still manually select these QC files at the Levey-Jennings, QC Data Table, or QC Compare screens as long as there is a loaded QC template. 7-2

103 REVIEWING QC DATA LOADING A QC TEMPLATE 7 Procedure To load an already defined QC template: 1. Applications tt QC. 2. File tt Select. 3. The desired protocol. 4. [Okay]. 5. The QC template appears. 6. Screen tt Levey-Jennings or Data Table to view the QC data in the format you want. 7-3

104 REVIEWING QC DATA RELATIONSHIP BETWEEN THE LEVEY-JENNINGS GRAPHS AND THE QC DATA TABLE 7.3 RELATIONSHIP BETWEEN THE LEVEY-JENNINGS GRAPHS AND THE QC DATA TABLE The Levey-Jennings graphs and the QC Data Table are linked together to better help you review the QC data. Once the QC template is loaded, the QC data files are available for viewing, then you can switch back and forth between the table and graphs to view and modify the data. Data Colors The colors of the data points on the Levey-Jennings graph and the data on the QC data table have specific meanings. When you print the Levey-Jennings graph, different symbols are used to represent the data colors. Data Color Data Description Data Point Print Symbol Black The data is inside the limits. Solid diamond Red The data is outside the limits. Large, diamond outline Blue The data point related to this information is currently N/A under the cursor in the Levey-Jennings graph. Green The mean and its related limits were manually M is after the data point entered in the Levey-Jennings graph. Purple These data points were deleted and [REM] is on in D is after the data point the QC Data Table screen. Yellow These data points were selected as reference points in the QC data table. If [RD REF] is on, the statisitics are recalculated for the table and graph. R is after the data point Lot Numbers r Lot numbers are only present in a QC file if a rectilinear or amorphous lot number region was created in the _A, _C, or _Q protocol that produced the data. The lot number region name must begin with LOT#= followed by up to eight letters and/or numbers. r The _Axyz protocol supplies the lot number of the Flow-Set or Flow-Check Fluorospheres to the nxyz.qcs file and is posted with the voltage and gain values. This same lot number appears next to the nxyz.qcs file name at the top of the screen. r The _C protocol supplies the lot number of the CYTO-COMP compensation reagents into the nxyz.qcs file of the _Axyz protocol run in the Autostandardization panel and is posted with the compensation values. If using multiple _C protocols in the panel, add the lot number of the CYTO-COMP Reagent Kit to the last _C protocol. On the Levey-Jennings graph you can only view this lot number in the Data Info pop-up box for a specific data point. If a nxyz.qcs file entry containing volts, gains, and compensation values appears on the QC Data table, the data appears on two lines: one line with the voltage and gains values and the lot number from the _Axyz protocol that produced them and the second line appears with the compensation values and lot number of the _C protocol that produced them. r The _Q xyzabcd protocol supplies the lot number of the Immuno-Trol Cells, CYTO-TROL Control Cells, or other control used to the nxyzabcd.qcc file and is posted with the QC statistical data. This same lot number appears next to the nxyzabcd.qcc file name at the top of the screen. 7-4

105 REVIEWING QC DATA REVIEWING LEVEY-JENNINGS GRAPHS REVIEWING LEVEY-JENNINGS GRAPHS Use the Levey-Jennings graphs to quickly check the QC data. Load a QC template at the QC Template screen to be able to view its related QC files at the QC Levey-Jennings screen. After you load a QC template or select a QC data file: 1. Screen tt Levey-Jennings. 2. æ and ç to view all the graphs you need to review. 3. [Day/Month/3 Month/6 Month] until the time scale you want appears. 7-5

106 REVIEWING QC DATA REVIEWING LEVEY-JENNINGS GRAPHS 4. Visually check the graphs data points. Note: See the Data Color chart in heading 7.3, Relationship Between the Levey-Jennings Graphs and the QC Data Table, to see the descriptions of the colored data points. 5. The scroll bar to move along the time scale. The corresponding data points appear for viewing. 6. To insert a cursor and display additional information for a data point: [Cursor]to turn it on (green). On or near the needed data point. A cursor spanning the three graphs appears on the data point you selected. The Data Point Info. pop-up box appears for the data point you selected. 7-6

107 REVIEWING QC DATA REVIEWING LEVEY-JENNINGS GRAPHS 7 7. To move the cursor: The cursor to the area of interest. Release the mouse button and the cursor snaps to the closest data point on the selected graph. Click the right mouse button to the left (or right) of the cursor to move the cursor over to the next data point on the left (or right). 8. File tt Print if you need to print a Levey-Jennings graph for your laboratory notebook. Note: See the Data Color chart in heading 7.3, Relationship Between the Levey-Jennings Graphs and the QC Data Table, to see which symbols are printed to correspond to the colored data. 9. [Westg] to view the Westgard Report for the QC files in this template. Rule failures appear for each Westgard rule. 7-7

108 REVIEWING QC DATA REVIEWING THE QC DATA TABLE 7.5 REVIEWING THE QC DATA TABLE Use the QC Data Table to quickly check the QC data. Load a QC template at the QC Template screen to be able to view its related QC files at the QC Data Table screen. The mean, SD, and CV at the bottom of the data table are calculated using unrounded values. If you calculate them manually or in a spreadsheet software application using the rounded values which are displayed, the results may be slightly different. 7-8

109 REVIEWING QC DATA REVIEWING THE QC DATA TABLE 7 After you load a QC template or select a QC data file: 1. Screen tt Data Table. 2. è or é to view all the columns (up to 40) of parameter data. An X appears in a row when there is no available data for a specific Run# and Date. The corresponding statistical data appears in the table below the columns. 3. Visually check the statistical data in the bottom table. It corresponds to the data displayed in the parameter columns above it. 4. The scroll bar to view all the runs for this QC file. 5. To enter and view additional lot number information: [Lot#]. See heading 7.7, Working with the Lot Information Pop-up Box, for more details. 6. To remove data from the Data Table (and Levey-Jennings graphs): [Rem] to turn it on (green). Any run that you want to remove. The entry appears purple and the data is automatically removed from the statistics. When [Rem] is OFF (blue) these data points do not appear. Note: If you want to replace the removed data, the run number again. 7-9

110 REVIEWING QC DATA REVIEWING THE QC DATA TABLE 7. To select reference points and recalculate the statistics based on them: [Rem] to turn it off (blue). Any run number that you want to select as a reference point. The entry appears yellow. [Rd Ref] to turn it on (green). The statistics are then recalculated with only the selected reference points. [Wr Ref] to update the QC file with these selected points. 8. File tt Print if you need to print a copy of the QC data table for your laboratory notebook. 9. [Westg] to view the Westgard Report for the QC files in this template. Rule failures appear for each Westgard rule. 7-10

111 REVIEWING QC DATA COMPARING TWO QC FILES COMPARING TWO QC FILES Use this feature to compare any two QC (*.QCS or *.QCC) data files at the QC Compare screen. For example, compare the *.QCC files from the same protocol for two lot numbers. If a QC template is loaded, its QC file (if it contains data) automatically appears in the bottom tables of the QC Compare screen when you access the screen. You can replace this file by selecting a different QC file. Reference file name and lot number Selected QC file name and lot number Data from the reference file. Data from the QC file. Moves among all the runs in the file B Moves among all the parameters columms. Displays the Lot Information pop-up box. When on, statistics are calculated from only the reference points of the file. 7-11

112 REVIEWING QC DATA COMPARING TWO QC FILES To compare two QC files: 1. Applications tt QC. 2. Screen tt Compare. 3. File tt Select. 4. The desired QC file. 5. [Okay]. The data from the QC file appears in the tables at the bottom of the screen. 6. Reference tt Select. 7. The desired reference QC file. 8. [Okay]. The data from the reference file appears in the tables at the top of the screen. 9. The scroll bars to view all the runs for each file. 10. To see the statistics from both files calculated only with their reference points: [Rd Ref] to turn it on. 7-12

113 REVIEWING QC DATA WORKING WITH THE LOT INFORMATION POP-UP BOX To enter and view additional lot number information: [Lot#]. See heading 7.7, Working with the Lot Information Pop-up Box, for more details. File tt Print if you need to print a copy for your laboratory notebook. 7.7 WORKING WITH THE LOT INFORMATION POP-UP BOX The Lot Information pop-up box can be accessed from the QC Data Table screen or the QC Compare screen. The Lot Information pop-up box displays data for your record keeping only. The system does not use the information in this screen to update files or keep track of lot numbers. The lot number does not automatically update when you change the lot number in a lot number region of a _A, _C, or _Q protocol. At the QC Data Table screen or QC Compare screen: 1. [Lot#] to display the Lot Information screen. 2. The desired row and enter the product name, lot number, expiration date, and the protocol in which it is used. Up to 100 rows can be entered. 3. [Print] if you want to print this screen for your laboratory notebook. 7-13

114 REVIEWING QC DATA WORKING WITH THE LOT INFORMATION POP-UP BOX 7-14

115 6PRINTING AND EXPORTING PATIENT REPORTS INTRODUCTION The Reports application allows you to set up a report template for printing and then allows you to enter specimen information. You can also export selected results from the patient report through the serial port to an ASCII (*.PXP) file. Before you can generate patient reports you must define and verify a report template for each panel you use for patient reporting. In addition to defining the report template, you must also define the export settings in the Patient Export Control pop-up box before exporting any patient reports (*.PXP files). Refer to the Data Management manual for detailed information on defining and verifying report templates and defining export settings in the Patient Export Control pop-up box. 6-1

116 PRINTING AND EXPORTING PATIENT REPORTS PRINTING PATIENT REPORTS 6.2 PRINTING PATIENT REPORTS To print patient reports after acquiring data or after using the MCL: 1. Applications tt Reports. 2. File tt Select. Reports are listed by the run number of the first tube in the panel. 3. The files you want to print. Some panel information is displayed on the right side of the screen to identify the report. 4. [Okay]. 5. [Results_Prn Disabled] to change it to Enabled. 6. [Output Results] and then [Next Report] to print the reports one at a time. OR [Batch] to print all the reports. When checking results, remember: If asterisks (****) replace results, there was either an invalid result or a channel overflow of >65,535 events occurred in one channel. The patient report rounds percentages, cells/µl, and cells/l to one decimal place. Values for counts and equation results that are derived from percentages are calculated after the percentages have been rounded off. 6-2

117 PRINTING AND EXPORTING PATIENT REPORTS EXPORTING PATIENT REPORTS (*.PXP) FILES EXPORTING PATIENT REPORTS (*.PXP) FILES Overview You must first define a report template in the Report application as well as the export settings at the Patient Export Control pop-up box before you can export patient reports. See the Data Management manual for detailed instructions to define a report template and the export settings. You can export patient reports: r At the Acquisition Output Options, or r At the Listmode Output Options, or r At the Reports Data Entry Screen. You can view the exported information while you are in the SYSTEM II software. Exporting from the Acquisition Application 1. Applications tt Acquisition. 2. Panel tt Select. 3. The panel you want. 4. [Okay]. 5. [Output Option]. 6. Each option until the selection you want appears. You must first enable the Patient Report then you can enable the Patient Export feature. Note: Export refers to the Run Reports (EPT and CYT files). 6-3

118 PRINTING AND EXPORTING PATIENT REPORTS EXPORTING PATIENT REPORTS (*.PXP) FILES 7. If you want to edit the Patient Export Control pop-up box but it did not automatically appear: in the Output Options pop-up box change the current selection for the Patient Report, Database, and Patient Export fields. When the Patient Export Control pop-up box appears make any changes that you want. [Pop Up Patient/Bypass Patient] to display [Pop Up Patient] only if you need to enter patient information in the Pop Up Patient box. [Okay]. 8. [Okay] at the Output Options pop-up box. 6-4

119 PRINTING AND EXPORTING PATIENT REPORTS EXPORTING PATIENT REPORTS (*.PXP) FILES 6 Exporting from the Listmode Application 1. Applications tt Listmode. 2. File tt Select. 3. The run numbers you want corresponding to the panel that was run. 4. [Okay]. 5. [Runtime Protocol ]to change it to [New Pnl/Pro]. 6. Panel tt Select. 7. The panel that was used to acquire the selected files. 8. [Okay]. 9. [Output Option]. 10. Each option until the selection you want appears. You must first enable the Patient Report in order to enable the Patient Export feature. Note: Export refers to the Run Reports (EPT and CYT files). 6-5

120 PRINTING AND EXPORTING PATIENT REPORTS EXPORTING PATIENT REPORTS (*.PXP) FILES 11. If you want to edit the Patient Export Control pop-up box but it did not automatically appear: in the Output Options pop-up box change the current selection for the Patient Report, Database, and Patient Export fields. When the Patient Export Control pop-up box appears make any changes that you want. [Pop Up Patient/Bypass Patient] to display [Pop Up Patient] only if you need to enter patient information in the Patient pop-up box. [Okay]. 12. [Okay] at the Output Options box. 13. [Play Next] to replay the files one at a time and export. Only one report is generated for multiple files. OR [Batch Enable] to replay all the files and export. Note: All specimen information except the Specimen ID can be modified during Listmode analysis. Any patient report generated after modifying the data includes the modified information and its patient report file is identified with a different extension (RPL). Any patient report export file generated in the Listmode application after modifying the data includes the modified information, but the patient report export file IS NOT identified with a different extension (PXP). 6-6

121 PRINTING AND EXPORTING PATIENT REPORTS EXPORTING PATIENT REPORTS (*.PXP) FILES 6 Exporting from the Reports Data Entry Screen To export patient reports either after acquiring data (with Patient Report enabled), or after you have replayed the data in Listmode (with Patient Report enabled): 1. Applications tt Reports. 2. The.RPA files or.rpl files you want to export. 3. [Okay]. 4. [Export Control]. Check that the Patient Export Control pop-up box contains the format parameters you want. See the Data Management manual for detailed instructions. 6-7

122 PRINTING AND EXPORTING PATIENT REPORTS EXPORTING PATIENT REPORTS (*.PXP) FILES 5. [Results_Exp Disabled] to change it to Enabled. 6. [Output Results] and then [Next Report] to export the files one at a time. OR [Batch] to export all the files. Note: The patient report export (PXP) file does not identify whether the exported data is from the Acquisition or Listmode application. Viewing a *.PXP File This feature allows you to check that the entries were written to the.pxp file. To view a *.PXP file within the SYSTEM II software: 1. Press É. The Help screen appears. 2. Press É again. The name of the last *.PXP file appears. 3. If this is not the name of the *.PXP file you want, type in another *.PXP file name and press Û. 4. An ASCII version of the file appears. Note: Not all characters may be viewable on the screen. Select File tt Print to view the entire file. 6-8

123 8CLEANING AND SHUTDOWN CLEANING AND SHUTDOWN FREQUENCY When to Clean the Sampling System Routine daily cleaning helps to minimize instrument downtime. r For the XL-MCL flow cytometer, follow both Manual and Auto mode procedures. r For the XL flow cytometer, follow only the Manual mode procedures. The two levels of cleaning for the system are: r r Routine cleaning followed by sample head cleaning. Vacuum line cleaning. 1. Perform the routine and sample head cleaning procedures: r When you change laboratory application procedures, especially if you are using vital fluorescent stains. If vital stains such as propidium iodide, ethidium bromide, acridine orange, thiazole orange, Coriphosphine-O, Fura 3, or fluorescein diacetate are used, perform these cleaning procedures immediately after using the dyes. r Immediately prior to running any immunophenotyping application if vital stains are being used on the same instrument. r When you observe a significant increase in debris or background counts. r Before you perform the vacuum line cleaning procedure. r Before you perform the shutdown procedure. 2. Perform the vacuum line cleaning procedure: r When you change laboratory application procedures, especially if you are using vital fluorescent stains. If vital stains such as propidium iodide, ethidium bromide, acridine orange, thiazole orange, Coriphosphine-O, Fura 3, or fluorescein diacetate are used, perform these cleaning procedures immediately after using the dyes. r Every 8 hours of continuous operation. r Before shutdown. When to Shut Down the Cytometer r Shut down the instrument at least once a day, even if is intended for use 24 hours per day. r Leave the instrument shut down for at least 30 minutes before restarting. 8.2 ROUTINE CLEANING PROCEDURE WARNING The cleaning solution is hazardous and can cause personal injury or damage clothing. Coulter Corporation urges its customers to comply with all national health and safety standards such as the use of barrier protection. This may include, but it is not limited to, protective eyewear, gloves, and suitable laboratory attire when operating or maintaining this or any other automated laboratory analyzer. IMPORTANT A cleaning solution that is not fresh can leave residual stain in the system and misleading results could occur when you change laboratory applications. Be sure to prepare a fresh cleaning solution before performing the cleaning procedure and use it within the same day. 8-1

124 CLEANING AND SHUTDOWN ROUTINE CLEANING PROCEDURE Manual Mode Procedure (XL and XL-MCL Flow Cytometers) 1. Prepare a cleaning solution of 1 part of high-quality, fragrance-free bleach (5% solution of sodium hypochlorite -available chlorine) and 9 parts of distilled water or IsoFlow sheath fluid. Put 2 ml of the bleach solution in a test tube. 2. At the Cytometer: Application tt Acquisition. Panel. Cleaning Panel. [Okay] B Check that the Cytometer status message reads Insert Sample Tube. If it does not call your Beckman Coulter Representative. 4. Put the test tube containing the 2 ml of bleach solution on the Sample Stage A The Sample Stage rises and the bleach solution is aspirated A 8-2

125 CLEANING AND SHUTDOWN ROUTINE CLEANING PROCEDURE 8 When the Sample Stage lowers, remove the tube A 5. Run three tubes of distilled water or IsoFlow sheath fluid. About 2 ml of distilled water or sheath fluid in each B 8-3

126 CLEANING AND SHUTDOWN ROUTINE CLEANING PROCEDURE Auto Mode Procedure (XL-MCL Flow Cytometer Only) 1. Put a test tube containing 2 ml of freshly prepared bleach solution in carousel position 1. Then put three freshly prepared tubes, each containing about 2 ml of distilled water or IsoFlow sheath fluid, in positions 2, 3, and 4 of the carousel. Do not reuse the tubes from the manual mode procedure. Ensure that the cleaning panel is still selected. If not, select it B 2. Put the carousel in the MCL sample loader and close the carousel door. Press the AUTO button or click [Run] on the Workstation screen. Press Y when the system asks Are you starting a new carousel? Press N when the system asks Do you want to enter specimen ID? 3. When the MCL sample loader is done, remove the carousel A A 8-4

127 CLEANING AND SHUTDOWN SAMPLE HEAD AND PROBE CLEANING PROCEDURE 8 Testing for Residual Stain If you use vital stains such as propidium iodide, ethidium bromide, acridine orange, thiazole orange, Coriphosphine-O, Fura 3, or fluorescein diacetate, you may want to test for residual stain after performing the routine cleaning procedure and before proceeding to your next application. To test for residual stain, run unstained Immuno-Trol Cells or CYTO-TROL Control Cells for your application to ensure that the autofluorescent population is where you normally expect it. If it is not, repeat the routine cleaning procedure. 8.3 SAMPLE HEAD AND PROBE CLEANING PROCEDURE Manual Mode Procedure (XL and XL-MCL Flow Cytometers) 1. Prepare a cleaning solution of 1 part of high-quality, fragrance-free bleach (5% solution of sodium hypochlorite -available chlorine) and 9 parts of distilled water or IsoFlow sheath fluid. While wearing latex gloves, apply the 10% bleach solution to a guaze pad B 2. Carefully push the moistened gauze pad up against the inside of the manual sample head and scrub away any debris inside and around the sample probe. Continue scrubbing the sample head and probe by pushing the head up and down 10 times during a 60-second period. Replace gauze as needed A 3. Rinse the sample head and probe with gauze moistened with water. 4. Perform the Vacuum Line Cleaning procedure. 8-5

128 CLEANING AND SHUTDOWN SAMPLE HEAD AND PROBE CLEANING PROCEDURE Auto Mode Procedure (XL-MCL Flow Cytometer Only) 1. Prepare a cleaning solution of 1 part of high-quality, fragrance-free bleach (5% solution of sodium hypochlorite -available chlorine) and 9 parts of distilled water or IsoFlow sheath fluid. While wearing latex gloves, apply the 10% bleach solution to a guaze pad B 2. Open the MCL lid and remove the carousel if installed. Carefully push the moistened gauze pad up against the inside of the MCL sample head and scrub away any debris inside and around the sample probe. Continue scrubbing the sample head and probe by pushing the head up and down 10 times during a 60-second period. Replace gauze as needed A 3. Rinse the MCL sample head and probe with gauze moistened with water. 4. Perform the Vacuum Line Cleaning procedure. 8-6

129 CLEANING AND SHUTDOWN VACUUM LINE CLEANING PROCEDURE VACUUM LINE CLEANING PROCEDURE Auto Mode Procedure (XL-MCL Flow Cytometer Only) 1. Fill two cleaning adaptors with 5 ml of distilled water B 2. Put one adaptor in position 1 of a carousel. Put the second adaptor in position 2. Orient the reservoirs toward the center of the carousel. Adaptor with distilled water in position 1. Adaptor with distilled water in position A 3. Put the carousel into the MCL and close the carousel door. Check that the system is in the Idle mode with the RUN light flashing green. If not, press the RUN button to put the system into Idle. Press the AUTO and CLEANSE buttons at the same time A 4. When Press Run to Initialize appears on the status line, remove the carousel. 8-7

130 CLEANING AND SHUTDOWN VACUUM LINE CLEANING PROCEDURE Manual Mode Procedure (XL and XL-MCL Flow Cytometers) 1. Press the RUN button to take the system out of the Run mode. The RUN light blinks when the unit is not in the Run mode; the RUN light glows steady green when the unit is in the Run mode. 2. Fill a cleaning adaptor reservoir with 5 ml of distilled water B 3. Install the adaptor just as a sample tube is installed on the Sample Stage. Wait for the water to be aspirated before proceeding to step B 4. Remove the adaptor and press the CLEANSE button. When the cleanse cycle ends, the CLEANSE button indicator turns off and the RUN button indicator flashes. The Cytometer status message reads Press Run to Initialize

131 CLEANING AND SHUTDOWN SHUTDOWN PROCEDURE SHUTDOWN PROCEDURE 1. Check that the Routine Cleaning and Vacuum Line Cleaning procedures have been completed. 2. Wipe down all exposed surfaces with 10% bleach solution and then 70% ethanol. Pay special attention to the Sample Stage areas, including the sample probe guides A 3. Put deionized water in a test tube. Put the tube on the manual Sample Stage A 4. Record daily shutdown and cleaning on the QC Maintenance screen: Applications tt QC. Screen tt Maintenance. Each box related to today s daily shutdown and cleaning checks. This enters your operator ID color and symbol. See the Special Procedures and Troubleshooting manual for detailed instructions on the QC Maintenance screen. 8-9

132 CLEANING AND SHUTDOWN SHUTDOWN PROCEDURE 5. Turn off the system at the Start Up box on the Utilities Configuration screen. The box after Power. Respond Y when the message Do you want to turn off the cytometer? appears. Turn off the monitor and Printer separately. 6. Keep the system shut down for 30 minutes. Before running samples, do the daily startup and quality control procedures. Reminder: r Clean the air filters once a week. r Clean the sheath fluid container once a month. r Clean the cleaning agent container every 60 days. Cleaning instructions are in the Special Procedures and Troubleshooting manual. 8-10