Reference. 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 Cytomete COULTER EPICS XL-MCL Flow Cytomete SYSTEM II Softwae Refeence CYTOMETER LASER SAMPLE FLOW LOW MED HIGH (Septembe 2010) Beckman Coulte, Inc. 250 S. Kaeme Blvd. Bea, 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 alet you as follows: WARNING - Can cause injuy. CAUTION - Can cause damage to the instument. IMPORTANT - Can cause misleading esults. 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 opeato injuy if: All doos, coves and panels ae not closed and secued in place pio to and duing instument opeation. The integity of safety intelocks and sensos is compomised. Instument alams and eo messages ae not acknowledged and acted upon. You contact moving pats. You mishandle boken pats. Doos, coves and panels ae not opened, closed, emoved and/o eplaced with cae. Impope tools ae used fo toubleshooting. To avoid injuy: Keep doos, coves and panels closed and secued in place while the instument is in use. Take full advantage of the safety featues of the instument. Do not defeat safety intelocks and sensos. Acknowledge and act upon instument alams and eo messages. Keep away fom moving pats. Repot any boken pats to you Beckman Coulte Repesentative. Open/emove and close/eplace doos, coves and panels with cae. Use the pope tools when toubleshooting. CAUTION System integity might be compomised and opeational failues might occu if: This equipment is used in a manne othe than specified. Opeate the instument as instucted in the Poduct Manuals. You intoduce softwae that is not authoized by Beckman Coulte into you compute. Only opeate you system s compute with softwae authoized by Beckman Coulte. You install softwae that is not an oiginal copyighted vesion. Only use softwae that is an oiginal copyighted vesion to pevent vius contamination. IMPORTANT If you puchased this poduct fom anyone othe than Beckman Coulte o an authoized Beckman Coulte distibuto, and, if it is not pesently unde a Beckman Coulte sevice maintenance ageement, Beckman Coulte cannot guaantee that the poduct is fitted with the most cuent mandatoy engineeing evisions o that you will eceive the most cuent infomation bulletins concening the poduct. If you puchased this poduct fom a thid paty and would like futhe infomation concening this topic, call you Beckman Coulte Repesentative.

3 REVISION STATUS Initial Issue, 1/96 SYSTEM II Softwae Vesion 1.0. Initial issue fo custome distibution. Issue B, 12/98 Complete evision. SYSTEM II Softwae, Vesion 3.0. Includes new quality contol featues, patient epot expot featue, and use-defined un time epots. Issue C, 01/08 Changes wee made to the Intended Use statement on page 1-1 and to the lase desciption on page 4-2. Issue CA, 09/10 Updates wee made to the company copoate addess. Note: Changes that ae pat of the most ecent evision ae indicated in text by a ba in the magin of the amended page. This document applies to the latest softwae listed and highe vesions. When a subsequent softwae vesion changes the infomation in this document, a new issue will be eleased to the Beckman Coulte website. Fo labeling updates, go to and download the most ecent manual o system help fo you instument. iii

4 REVISION STATUS iv

5 CONTENTS WARNINGS AND PRECAUTIONS, ii REVISION STATUS, iii INTRODUCTION, xiii USING YOUR XL AND XL-MCL FLOW CYTOMETER MANUALS, xiii ABOUT THIS MANUAL, xiii CONVENTIONS, xiv Desciption of Repoting Units, xiv 1 USE AND FUNCTION, INTENDED USE, SYSTEM COMPONENTS, 1-2 Cytomete, 1-2 Powe Supply, 1-2 Wokstation, OPTIONS, 1-3 Hadwae Options, 1-3 Multi-tube Caousel Loade (MCL), 1-3 Pinte, 1-4 Ba-Code Pinte, 1-4 Ba-Code Scanne, 1-4 Data Stoage, 1-4 Netwok Kits, 1-4 Wokstation/Netwok Seve,1-4 Fouth FL Senso, 1-5 RAM Upgade Kits, 1-5 Softwae Options, 1-5 tetaone System, 1-5 eticone System, 1-5 MULTICYCLE Softwae, 1-5 Atisoft LANtastic Softwae, 1-5 Sybase SQL Anywhee Softwae, REAGENTS AND QUALITY CONTROL MATERIALS, 1-5 Sheath Fluid, 1-5 Cleaning Agent, 1-6 Quality Contol Mateials, MATERIAL SAFETY DATA SHEETS (MSDS), INSTALLATION, DELIVERY INSPECTION, SPECIAL REQUIREMENTS, 2-1 Space and Accessibility, 2-1 v

6 CONTENTS Electical Input, 2-1 Ambient Tempeatue and Humidity, 2-1 Heat Dissipation, 2-1 Dainage, SYSTEM CONNECTIONS, 2-2 Powe and Signal Cables, 2-2 Waste and Pneumatic Tubing, INSTALLING SOFTWARE VERSION 3.0, OPERATION PRINCIPLES, WHAT THIS CHAPTER EXPLAINS, SAMPLE FLOW, 3-1 Sample Loading, 3-1 Manual, 3-1 Automated, 3-1 Hydodynamic Focusing, LASER BEAM SHAPING, CELL ILLUMINATION, 3-4 Fowad Scatte, 3-4 Side Scatte and Fluoescent Light, LIGHT COLLECTION, SEPARATION AND MEASUREMENT, 3-4 Fowad Scatte Collection, 3-4 Neutal Density Filte, 3-6 Side Scatte and Fluoescent Light Collection, 3-6 Side Scatte, 3-6 Fluoescent Light, SIGNAL PROCESSING, 3-9 Voltage Pulse Signals, 3-9 Peak Signal, 3-9 Integal Signal, 3-9 Amplification, 3-11 Signals Geneated, PARAMETERS, 3-12 AUXiliay Paamete, 3-12 When to Use the AUX Paamete, 3-12 TIME Paamete, 3-12 RATIO Paamete, 3-13 PRISM Paamete, AUTOSTANDARDIZATION PROTOCOLS, 3-15 _A Potocols, 3-15 _C Potocols, 3-15 _Q Potocols, 3-15 vi

7 CONTENTS Examples of Autostandadization Potocols and Panels, QUALITY CONTROL (QC) FEATURES, 3-16 QC Templates, 3-16 Levey-Jennings Gaphs, 3-16 Data Table, 3-17 Westgad Rules,3-17 Compae Sceen, 3-17 Maintenance Sceen, HISTOGRAM DISPLAY, 3-18 Regions, 3-18 Gating, 3-18 Data Stoage, 3-19 Histogam Statistics, 3-19 Linea Region Statistics, 3-19 Log Region Statistics, SPECIFICATIONS/CHARACTERISTICS, SAMPLE REQUIREMENTS, INSTRUMENT SPECIFICATIONS, 4-1 Dimensions, 4-1 Installation Categoy, 4-1 Cytomete, 4-1 Flow Cell, 4-1 Flow Rate, 4-1 Lase, 4-2 Agon Lase Powe, 4-2 Beam-Shaping Optics, 4-2 Lase Beam Spot Size, 4-2 Optical Filtes, 4-2 Sensos, 4-2 Signal Pocessing, 4-2 Wokstation,4-3 Compute, 4-3 Data Stoage, 4-4 Intefaces, 4-4 Input Devices, 4-4 Monito, SOFTWARE SPECIFICATIONS, 4-4 Data Output and Compatibility, 4-4 Repoting Units, 4-4 Two-Digit Date Fomat, 4-5 Fo Non Date-of-Bith Dates, 4-5 Fo Date-of-Bith Dates, 4-5 Acquisition, 4-5 Regions, 4-5 Listmode Analysis, 4-5 vii

8 CONTENTS Multigaph Analysis, PERFORMANCE SPECIFICATIONS, 4-6 Cayove, 4-6 Data Acquisition Thoughput, 4-6 Doublet Discimination, 4-6 Pecision fo Suface Makes, 4-6 Resolution, 4-6 Fowad Scatte, 4-6 Fluoescence, 4-6 Sensitivity, 4-6 Scatte, 4-6 Fluoescence, 4-6 Stability, 4-7 Day-To-Day, 4-7 Within Day, PERFORMANCE CHARACTERISTICS, BAR-CODE SPECIFICATIONS, 4-7 Ba-Code Labels, 4-7 Acceptable Ba Codes, 4-7 Ba-Code Label Optical Chaacteistics at 670 nm ±10%, 4-7 NE Width, 4-8 WE/NE Ratio, 4-8 Pinting Methods, 4-8 MCL Ba-Code Reade, 4-8 Hand-Held Ba-Code Reade, 4-8 Ba-Code Decode, 4-8 Check Sum Algoithm, PRECAUTIONS AND HAZARDS, LASER SAFETY, WARNINGS, WARNING LABELS, 5-1 A B LOG SHEETS,A-1 PREDEFINED PANELS, PROTOCOLS, AND QC TEMPLATES GLOSSARY INDEX BECKMAN COULTER, INC. CUSTOMER END USER LICENSE AGREEMENT TRADEMARKS viii

9 ILLUSTRATIONS 1.1 EPICS XL-MCL Flow Cytomety System, MCL Caousel Ba-Code Labels, Powe and Signal Cable Connections, Waste and Pneumatic Tubing Connections, Installing the Softwae Diskette, Flow Cell (Hydodynamic Focusing), Lase Beam Shaping, Optical System, Filte Configuation, Thee Fluoescence Sensos, Filte Configuation, Fou Fluoescence Sensos, Voltage Pulse Fomation, Peak Signal, Integal and Peak Pulses, Pism Histogam, Lase Labels on the Sensing Compatment Cove, Lase Labels in the Optical Aea, Side View, Lase Labels in the Optical Aea, Font View, Lase Labels on the Lase Head, Lase Label on the Cytomete Back Panel, Lase Labels on the MCL Pobe Housing Cove, Figue 20 Lase Labels on the MCL Ba-Code Reade, 5-6 ix

10 TABLES 1.1 Applications fo the Instument, Code-Related Specifications, Geneal Use Panels and Potocols, B Two-Colo Application Panels and Potocols, B Thee-Colo Application [4 PMT System] Panels and Potocols, B Thee-Colo Application [3PMT System] Panels and Potocols, B Fou-Colo Application Panels and Potocols, B DNA Application Panels and Potocols, B-7 x

11 INTRODUCTION This intoductoy section contains the following topics: Using you COULTER EPICS XL and XL-MCL Flow Cytomete manuals About this Manual Conventions Icons. USING YOUR XL AND XL-MCL FLOW CYTOMETER MANUALS Use the Refeence manual fo in-depth infomation on the pinciples of flow cytomety, infomation about what you instument does, the methods it uses, its specifications, and infomation on installation, safety, and system options. Use the Getting Stated manual to become familia with the contols and indicatos fo you system and to lean about potocols, egions, panels, and the basic skills you need to opeate the system. This manual also has an oveview of the softwae. Use the Opeato s Guide fo the day-to-day unning of you instument. Go though the detailed step-by-step pocedues of statup, quality contol (QC), unning samples, analyzing data, pinting epots, eviewing QC data, and shutdown. Use the Data Management manual fo instuctions on how to expot, save, copy, move, achive, and delete files. It also has infomation about the types of files you system ceates and uses, instuctions fo woking with QC featues, and instuctions fo setting up the epot template that you need to ceate you patient epots. Use the Special Pocedues and Toubleshooting manual to clean, eplace, o adjust a component of the instument. The Toubleshooting tables and eo messages appea at the back of the manual. Use the Opeating Summay as a quick efeence fo basic pocedues. Use the Maste Index to easily locate a topic in any of you manuals. Use the Use's Comment Cad in the Refeence manual to give us you comments about the manual and ways to impove it. ABOUT THIS MANUAL You EPICS XL and XL-MCL Flow Cytomete Refeence manual povides in-depth infomation about what the instument does, the methods it uses, its specifications, and infomation on installation, safety and softwae featues. This infomation is oganized as follows: s s s Chapte 1, Use and Function Contains the intended use of the instument, the eagents used, and a shot desciption of the majo components and options. Chapte 2, Installation Contains instument equiements, and diagams of the inteunit cable connections. Chapte 3, Opeation Pinciples Contains a desciption of flow cytomety, the nomal sample flow though the xi

12 INTRODUCTION CONVENTIONS s s s s s instument, how light collection and signal pocessing ae accomplished and how the paametes ae deived. Chapte 4, Specifications/Chaacteistics Details the instument and pefomance specifications, the pefomance chaacteistics and intefeing substances. Chapte 5, Pecautions and Hazads Descibes lase safety pecautions and the location of the lase-elated labels. Appendices The appendices povide efeence mateial on the following topics: Log Sheets Potocols Glossay The glossay has definitions fo many of the wods and tems used in flow cytomety. Index Use the Index to easily locate specific infomation in this manual. CONVENTIONS This manual uses the following conventions: Thoughout this manual you EPICS XL o XL-MCL flow cytomete is efeed to as the system. Italics indicate sceen messages. Bold indicates a menu item. Couie font indicates text you have to type using the keyboad. ë indicates a key (such as Û). ë+ë indicates that the two keys listed (such as Þ+Ê) ae linked fo a specific function and must be pessed in this sequence: 1. Pess down on the fist key listed and while continuing to pess it, pess down on the second key listed. 2. Release both keys at the same time. ë ë indicates to pess and elease the fist key listed then pess and elease the next key listed. Fo example, Y Û. File tt Save Select the Save item on the File menu. [OKAY] Use the mouse to click on the sceen button labeled [OKAY]. É though Ô Special function keys. Desciption of Repoting Units Unless othewise stated, all paamete units ae shown in the US unit fomat (cells/µl) thoughout the manuals. xii

13 1USE AND FUNCTION INTENDED USE The XL and XL-MCL flow cytometes ae systems intended fo use as in vito diagnostic devices fo the qualitative and quantitative measuement of biological and physical popeties of cells and othe paticles. These popeties ae measued when the cells pass though a lase beam in single-file. The instument can simultaneously measue fowad scatte, side scatte, and thee o, optionally, fou fluoescent dyes using a single lase at 488 nm. Theefoe, the instument can do coelated multipaamete analyses of individual cells. Table 1.1 lists the majo applications fo the instument. In addition to human cells, othe cell types can be analyzed, such as: Plant cells Maine plankton Animal cells Bacteia Table 1.1 Applications fo the Instument Applications Cell suface antigens Nucleic acids Cell Types Whole blood Buffy coats Mononuclea cells Dissociated tissue Platelets Bone Maow Whole blood Dissociated tissue Fozen sections Paaffin sections Body fluids Sample Pepaation (efe to the eagent's package inset) Measuements Paametes ImmunoPep eagents with the Q-PREP Wokstation, the Multi-Q-Pep Wokstation o the TQ-Pep Wokstation Whole blood lysing eagent kit Ficol Fluoescent-labeled antibodies IOTest 3 lysing OptiLyse C lysing VesaLyse Vaious staining methods, including: Ethidium bomide Popidium iodide DNA Pep eagents Cell size and ganulaity FITC, RD1, ECD, and PC5 Nucleic acid content Geen, ed, and oange fluoescence Log/Linea Fowad and side scatte Log/Linea fluoescence one, two, thee, and fou (optional) Pism Ratio Time AUX Fowad and side scatte Log/Linea Fluoescence one, two, and thee (fou is optional). Ratio, AUX 1-1

14 USE AND FUNCTION SYSTEM COMPONENTS Table 1.1 Applications fo the Instument (Continued) Kinetics Cell function Whole blood Buffy coats Mononuclea cells Tissue cultue cells Whole blood Buffy coats Mononuclea cells Tissue cultue cells Vaious methods, including: Fluo 3 fluoescence Vaious methods and dyes, including: DCFH-DA DiOC 5 (3) FDA Reticulocytes Whole blood Thiazole oange coiphosphine O (CPO) 1.2 SYSTEM COMPONENTS The system components ae shown in Figue 1.1. Cell size and ganulaity Geen fluoescence Time Cell size and ganulaity Geen fluoescence Cell size and ganulaity Geen fluoescence Red fluoescence Fowad and side scatte Fluoescence one Ratio Time Fowad and side scatte Fluoescence one Log/Linea Fowad and Side Scatte Log Fluoescence one Log Fluoescence fou Cytomete This unit analyzes the sample. It contains the sheath fluid and cleaning agent bottles. A fouth fluoescence senso and MCL (Multi-tube Caousel Loade) ae optional. Powe Supply This unit povides dc powe, pessue, and vacuum to the Cytomete. A 4-L waste bottle sits in a backet on the ight side of the Powe Supply. Wokstation The Wokstation uns the softwae that enables you to contol the instument. It displays sample esults and othe infomation. It consists of: A monito A compute with data stoage devices A keyboad and a mouse. 1-2

15 CYTOMETER READY LASER ON SAMPLE FLOW LOW MED HIGH FS SS AUX FL1 FL2 FL3 FL4 TM TM USE AND FUNCTION OPTIONS 1 Figue 1.1 EPICS XL-MCL Flow Cytomety System CYTOMETER COMPUTER WORKSTATION MCL OPTION POWER SUPPLY MODULE D 1.3 OPTIONS Hadwae Options Multi-tube Caousel Loade (MCL) The MCL is an automated sample loade fo the instument. It uses a caousel that holds thity-two 12- x 75-mm test tubes. The MCL eads the following ba-code types: Codaba Code 39 ba code Code 128 Inteleave 2-of-5. Fo additional infomation on the ba-code specifications, see Heading?, Ba-Code Specifications. The MCL mixes each sample befoe analysis. You can inteupt the MCL to manually un Stat samples, and then have the MCL etun to whee it left off. A Woklist and MCL caousel epot can be displayed o pinted. Figue 1.2 shows the location of the caousel numbe, tube position, and sample tube ba-code labels on the MCL caousel. 1-3

16 USE AND FUNCTION OPTIONS Figue 1.2 MCL Caousel Ba-Code Labels SAMPLE BAR-CODE LABEL C TUBE POSITION BAR-CODE LABEL CAROUSEL ID BAR-CODE Pinte Povides a pintout of sample esults and othe infomation. The Pintes listed hee ae subject to change, so contact you Beckman Coulte Repesentative fo the coect model selection: HP LaseJet V HP 1200C and 1600C colo pintes HP 560C colo pinte. Ba-Code Pinte Used to pint ba-code labels fom the Woklist fo identification of sample tubes in the MCL caousel. Ba-Code Scanne A hand-held scanne/eade used to ead specimen collection ba-code labels. A fast, accuate method fo inputting specimen and sample tube infomation and tube ID into the softwae database. Data Stoage 2.6-GB optical dive 650-GB extenal, ecodable CD-ROM dive Iomega Zip extenal 100-MB dive (paallel) Netwok Kits Single and multi-node netwok kits used to connect an XL o XL-MCL system to an existing wokstation. Wokstation/Netwok Seve A COULTER FlowCente Multimedia Wokstation can be used as an off-line analysis wokstation and netwok seve fo data and woklist tansfe. The standad wokstation that uns the flow cytomete can be upgaded to a FlowCente Multimedia Wokstation. 1-4

17 USE AND FUNCTION REAGENTS AND QUALITY CONTROL MATERIALS 1 Fouth FL Senso This optional kit contains a photo-multiplie tube with a 200- to 800-nm spectal ange and a 675-BP dichoic filte that ae used to geneate the following signals (paametes): FL4 (can be assigned to the AUX paamete) FL4 LOG FL4 PEAK (must be assigned to the AUX paamete). RAM Upgade Kits Kits to upgade 8 MB RAM systems to 16 MB RAM. Softwae Options tetaone System Softwae and eagent kits fo pefoming automated fou-colo immunofluoescence analysis on the instument. eticone System Softwae and eagent kits fo pefoming automated eticulocyte enumeation on the instument. MULTICYCLE Softwae DNA analysis softwae. Atisoft LANtastic Softwae Netwok opeating system softwae. Sybase SQL Anywhee Softwae Allows access to the System II database fo extended queies. 1.4 REAGENTS AND QUALITY CONTROL MATERIALS Beckman Coulte ecommends these eagents o thei equivalents. All stated pefomance chaacteistics in this manual ae based on the use of the XL and XL-MCL system with the following eagents. Sheath Fluid In the Cytomete, sample is guided into a steam of sheath fluid to make the sample cells flow single file though the lase beam. IsoFlow sheath fluid, a nonfluoescent, balanced electolyte solution, is made fo this pupose. IsoFlow sheath fluid has the following chaacteistics: Filteed to 0.2 µm Tanspaent and nonfluoescent to 488-nm lase light Low backgound Compatible with the chaacteistics of the sample being measued (such as ph, osmolality, conductivity). The sheath containe has a woking capacity of about 2 L. The amount of sheath fluid the containe holds beyond the woking capacity is fo pessuization and level sensing. 1-5

18 USE AND FUNCTION MATERIAL SAFETY DATA SHEETS (MSDS) Cleaning Agent When the Cytomete is in the Cleanse mode, COULTER CLENZ cleaning agent flushes sample tubing and helps to educe potein buildup and paticles in the instument. Each cleanse cycle uses about 15 ml of cleaning agent. The cleaning agent containe has a woking capacity of about 500 ml. That is enough cleaning agent to use the Cleanse mode once a day fo one month. The amount of cleaning agent the containe holds beyond the woking capacity is fo pessuization and level sensing. Read the containe s label fo moe infomation on the cleaning agent. Quality Contol Mateials The quality contol mateials available fom Beckman Coulte ae: Flow-Check Fluoosphees Flow-Set Fluoosphees CYTO-TROL Contol Cells Immuno-Tol Cells CYTO-COMP Cell Kit CYTO-COMP Reagent Kit Fluoosphees used to check the stability of the optical and fluidic systems. Fluoosphees used to standadize light scatte and fluoescence intensity. Lyophilized lymphocytes with assay values fo specific suface antigens. Used to: Assess monoclonal antibody function. Veify pope flow cytomete setup. Stabilized eythocytes and leukocytes with a known quantity of suface antigens. Used to veify monoclonal antibody pefomance as well as veify the pocess of sample staining, lysing, and analysis. Lyophilized lymphocytes used with the CYTO-COMP Reagent Kit to adjust colo compensation settings fo multicolo analysis. Fou two-colo fluoescent eagents used with the CYTO-COMP Cell Kit to adjust colo compensation settings fo multicolo analysis. 1.5 MATERIAL SAFETY DATA SHEETS (MSDS) To obtain an MSDS fo Beckman Coulte eagents used on the XL o XL-MCL flow cytomete: 1. In the USA, eithe call Beckman Coulte Custome Opeations ( ) o wite to: Beckman Coulte, Inc. Attention: MSDS Requests P.O. BOX Miami, FL Outside the USA, contact you Beckman Coulte Repesentative. 1-6

19 2INSTALLATION DELIVERY INSPECTION The instument is tested befoe shipping. Intenational symbols and special handling instuctions ae pinted on the shipping catons to infom the caie of the pecautions and cae applicable to electonic instuments. CAUTION Possible instument damage could occu if you uncate the instument, install it, o set it up. Keep the instument in its packaging until you Beckman Coulte Repesentative uncates it fo installation and setup. When you eceive you instument, caefully inspect all catons. If you see signs of mishandling o damage, file a claim with the caie immediately. If sepaately insued, file the claim with the insuance company. 2.2 SPECIAL REQUIREMENTS Befoe you Beckman Coulte Repesentative aives to install the instument, you must detemine whee you want the system placed and the oveall layout. Conside the factos descibed in the following paagaphs. Space and Accessibility Allow oom to inteconnect the system components. Also, aange fo: Comfotable woking height Space fo ventilation, and access fo maintenance and sevice: Cytomete Powe Supply 30.5 cm (12 in.) fom the back 45.7 cm (18 in.) fom the top 30.5 cm (12 in.) on both sides Powe Supply 12.7 cm (5 in.) fom the back Electical Input CAUTION Possible instument damage could occu if you put the Powe Supply plugs on the same electical cicuit o use an extension cod o a powe stip to connect the Powe Supply. Use two dedicated 115-V, 20-A outlets with isolated gounds fo the Powe Supply plugs. The Powe Supply equies two dedicated 115-V, 20-A outlets with isolated gounds. The compute equies a sepaate 115-V, 20-A outlet, but it does not have to be a dedicated line. Ambient Tempeatue and Humidity Keep the oom tempeatue between 16 C and 32 C (60 F and 90 F), and do not let it change moe than 5 F pe hou. Keep the humidity between 5% and 80%, without condensation. Heat Dissipation Heat dissipation is 2,300 W (7,850 Btu/hou). Povide sufficient ai conditioning (efe to Ambient Tempeatue and Humidity). 2-1

20 LASER POWER MAX 1800 WATTS SYSTEM POWER MAX 1500 WATTS SYSTEM ON/OFF ON OFF TX RX CYT12 INSTALLATION SYSTEM CONNECTIONS Dainage The waste line fom the Cytomete is connected to a 4-L waste bottle, which sits in a backet on the ight side of the Powe Supply. Dispose of the waste in accodance with you local envionmental egulations and acceptable laboatoy pocedues. 2.3 SYSTEM CONNECTIONS Powe and Signal Cables Figue 2.1 shows the inteunit connections of the powe and signal cables. Figue 2.1 Powe and Signal Cable Connections CYTOMETER WORKSTATION MOUSE MCL OPTION TRANS TRANS REC. FLOWCELL PRESSURE VACUUM WASTE WASTE POWER SUPPLY TO OPTIONAL PRINTER AUX POWER ON AC POWER LINE CORDS TO KEYBOARD PRESSURE VACUUM WASTE LEVEL WASTE WASTE VENT AC POWER LINE CORDS B 2-2

21 ON OFF INSTALLATION INSTALLING SOFTWARE VERSION Waste and Pneumatic Tubing Figue 2.2 shows the inteunit connections fo waste and pneumatic tubing. Figue 2.2 Waste and Pneumatic Tubing Connections CYTOMETER FLOW CELL WASTE WASTE TRANS REC. PRESSURE VACUUM WASTE PRESSURE VACUUM POWER SUPPLY AUX POWER ON PRESSURE SYSTEM ON/OFF WASTE WASTE VENT WASTE LEVEL VACUUM WASTE LEVEL WASTE LASER POWER SYSTEM POWER MAX 1800 WATTS MAX 1500 WATTS WASTE VENT D 2.4 INSTALLING SOFTWARE VERSION 3.0 Use this pocedue if you ae installing softwae vesion 3.0 in the MS-DOS opeating envionment. If you have a FlowCente Multimedia Wokstation and want to stat the XL/XL-MCL flow cytomete softwae fom an icon on the Windows 95 desktop, call you local Beckman Coulte Repesentative. To install softwae vesion 3.0 in the MS-DOS opeating envionment: 1. Inset the diskette labeled 1 of 2 into the diskette dive. See Figue 2.3. Figue 2.3 Installing the Softwae Diskette A 2-3

22 INSTALLATION INSTALLING SOFTWARE VERSION At the DOS pompt (C:\>) type: a: (o b:) and pess Û. 3. Type: install and pess Û. 4. Follow the instuctions on the sceen. 5. Be sue to egiste you softwae afte installation is completed. 6. If you need to install the Utilities diskette in an off-line wokstation, o install the Potocol diskette in a Cytomete wokstation: Inset the diskette and follow the instuctions in steps 2 to 4 above. 2-4

23 3OPERATION PRINCIPLES WHAT THIS CHAPTER EXPLAINS This chapte explains how the Cytomete measues scatteed light and fluoescence as cells pass though the lase beam. The illustations in this chapte ae not exact epesentations of the inside of the Cytomete. They ae fo explanatoy puposes only. 3.2 SAMPLE FLOW CAUTION Possible flow cell damage. To avoid clogging the sample pobe, sample tubing o flow cell, ensue that 12 mm x 75 mm test tubes ae fee of debis befoe you use them. Sample Loading Manual When you place a tube containing a pepaed sample on the Cytomete sample stage, the stage ises to position the sample pobe nea the bottom of the tube. The tube is pessuized and sample flow begins. Automated The sample caousel has ba-code labels that identify the caousel and the tube position numbe. Also, you can put ba-code labels on the sample tubes. See Heading 4.6, Ba-Code Specifications. The MCL has a ba-code eade that eads the caousel numbe, the sample tube position, and the sample tube ba-code labels as the caousel otates. The MCL handles a sample tube as follows: It lifts the tube out of the caousel into a centeing cup. It moves the bottom of the tube in a cicula obit fo 3 seconds to mix the sample. It lowes its sample pobe into the tube and the tube is pessuized. Sample flow begins. In both manual and automated sample loading, the appopiate sample pobe is cleaned automatically when sample flow ends. Hydodynamic Focusing If the cells wee to move though the lase beam in diffeent ways duing sample flow, sample analysis could be distoted. The instument uses a pocess called hydodynamic focusing to ensue that the cells move though the lase beam one at a time, along the same path. Hydodynamic focusing occus in the flow cell. The flow cell (Figue 3.1) contains a squae channel. A pessuized steam of sheath fluid entes the channel at the lowe end and flows upwad. The sensing aea of the flow cell is at the cente of the channel. While the sheath steam is flowing though the channel, a steam of sample is injected into the middle of the sheath steam. As shown in Figue 6, the sheath steam suounds, but does not mix with, the sample steam. The pessue of the sheath steam focuses the sample steam so that the cells flow though the lase beam single file that is, one at a time. 3-1

24 OPERATION PRINCIPLES SAMPLE FLOW Figue 3.1 Flow Cell (Hydodynamic Focusing) TO WASTE SENSING AREA HAS SQUARE CHANNEL SAMPLE STREAM SHEATH STREAM DIRECTION OF LASER BEAM CELLS SHEATH STREAM ENTERS HERE TO WASTE C SAMPLE STREAM ENTERS HERE 3-2

25 OPERATION PRINCIPLES LASER BEAM SHAPING LASER BEAM SHAPING Befoe the lase beam eaches the sample steam, coss-cylindical lenses focus the beam (see Figue 3.2). Focusing keeps the beam pependicula to the sample steam flow while making the beam small enough to illuminate only one cell at a time. The fist lens contols the width of the beam; the second, the height. The esulting elliptical beam is focused on the sensing aea of the flow cell. Figue 3.2 Lase Beam Shaping CROSS SECTION AFTER VERTICAL LENS TO WASTE FLOW CELL C 3-3

26 OPERATION PRINCIPLES CELL ILLUMINATION 3.4 CELL ILLUMINATION As cells in the sample steam go though the sensing aea of the flow cell, the elliptical lase beam illuminates them. The cells scatte the lase light and emit fluoescent light fom fluoescent dyes attached to them. Fowad Scatte The amount of lase light scatteed at naow angles to the axis of the lase beam is called fowad scatte (FS). The amount of FS is popotional to the size of the cell that scatteed the lase light. Side Scatte and Fluoescent Light The amount of lase light scatteed at about a 90 angle to the axis of the lase beam is called side scatte (SS). The amount of SS is popotional to the ganulaity of the cell that scatteed the lase light. Fo example, SS is used to diffeentiate between lymphocytes, monocytes, and ganulocytes. In addition to the SS, the cells emit fluoescent light (FL) at all angles to the axis of the lase beam. The amount of FL enables the instument to measue chaacteistics of the cells emitting that light, depending on the eagents used. Fo example, FL is used to identify molecules, such as cell suface antigens. 3.5 LIGHT COLLECTION, SEPARATION AND MEASUREMENT Fowad Scatte Collection The FS senso (see Figue 3.3) collects the fowad scatte the lase light that is scatteed at naow angles to the axis of the lase beam. When light eaches the FS senso, the senso geneates voltage pulse signals. These signals ae popotional to the amount of light the senso eceives. As explained in Heading 3.6, Signal Pocessing, the signals ae pocessed to measue the chaacteistics of the cells that scatteed the light. 3-4

27 OPERATION PRINCIPLES LIGHT COLLECTION, SEPARATION AND MEASUREMENT 3 Figue 3.3 Optical System FL2 SENSOR FL3 SENSOR FL4 SENSOR ( OPTIONAL ) FL1 SENSOR LASER SS SENSOR LASER BEAM BEAM SHAPING LENSES FLOW CELL PICKUP LENS/ SPATIAL FILTER ASSEMBLY OPTICAL FILTER BLOCK NEUTRAL DENSITY FILTER ( IF DESIRED ) FS SENSOR LASER BEAM DIRECTION FS SENSOR FORWARD SCATTERED LIGHT ONTO THE FS SENSOR B 3-5

28 OPERATION PRINCIPLES LIGHT COLLECTION, SEPARATION AND MEASUREMENT Neutal Density Filte When you analyze lage cells (cells geate than 20 µm in diamete, such as plant cells), the lage amount of FS can satuate the FS senso. The instument has a neutal density optical (ND1) filte that solves this poblem by educing the FS signal by a facto of about 10. The ND1 filte is not needed fo cells less than 20 µm. The ND1 filte is built-in and can be positioned fo use as needed. The pocedue to do so is in the Special Pocedues and Toubleshooting manual, in the Replace/Adjust Pocedues chapte. Side Scatte and Fluoescent Light Collection In ode fo the sensos to measue SS and FL, the light must be collected and the SS and fluoescent light must be sepaated. The pickup lens/spatial filte assembly collects SS and FL fom only the sensing aea of the flow cell, and collimates it. This light then goes towad the SS senso. Side Scatte The wavelength of SS is 488 nm. It is much moe intense than FL. SS is the fist light sepaated fom the output of the pickup lens/spatial filte assembly. SS is sepaated using a 488 nm dichoic long-pass (488 DL) filte at a 45-degee angle to the light path (see Figue 3.4 and Figue 3.5). The 488 DL filte eflects the SS to the SS senso but tansmits fluoescent light of longe wavelengths. Fluoescent Light The instument has two fluoescence collection configuations. Thee FL sensos ae standad; a fouth FL senso is optional. Thee FL Sensos. Figue 3.4 shows the filte configuation fo thee FL sensos. The light the 488 DL filte tansmits goes to a 488 nm lase-blocking (488 BK) filte. The 488 BK filte blocks any emaining lase light, tansmitting only fluoescent light. The emaining optical filtes sepaate the light fo the thee FL sensos. A 550 DL filte is at a 45-degee angle to the light path. It eflects light less than 550 nm to a 525 nm band-pass (525 BP) filte that tansmits light between 505 nm and 545 nm to the FL1 senso. The light the 550 DL filte tansmits is between 555 nm and 725 nm. The next dichoic long-pass filte, also positioned at a 45-degee angle to the light path, is a 600 DL. It eflects light between 555 nm and 600 nm to a 575 BP filte in font of the FL2 senso. The 575 BP tansmits light between 560 nm and 590 nm to the FL2 senso. The 600 DL filte tansmits light between 605 nm and 725 nm to a 620 BP filte in font of the FL3 senso. The 620 BP filte tansmits light between 605 nm and 635 nm to the FL3 senso. 3-6

29 OPERATION PRINCIPLES LIGHT COLLECTION, SEPARATION AND MEASUREMENT 3 Figue 3.4 Filte Configuation, Thee Fluoescence Sensos FL1 SENSOR FL2 SENSOR FL3 SENSOR SS SENSOR 525 BP 575 BP 620 BP PICKUP LENS/ SPATIAL FILTER ASSEMBLY 488 BK 550 DL 600 DL LASER BEAM DIRECTION FLOW CELL 488 DL NEUTRAL DENSITY FILTER ( IF DESIRED ) FS SENSOR C 3-7

30 OPERATION PRINCIPLES LIGHT COLLECTION, SEPARATION AND MEASUREMENT Figue 3.5 Filte Configuation, Fou Fluoescence Sensos FL3 SENSOR FL4 SENSOR (OPTIONAL ) FL1 SENSOR FL2 SENSOR 575 BP 620 BP SS SENSOR 525 BP 675 BP 645 DL PICKUP LENS/ SPATIAL FILTER ASSEMBLY 550 DL 600 DL 488 BK LASER BEAM DIRECTION FLOW CELL 488 DL FS SENSOR NEUTRAL DENSITY FILTER ( IF DESIRED ) C Fou FL Sensos. Figue 3.5 shows the filte configuation fo fou FL sensos. It uses the same filtes as the thee FL senso configuation, up to and including the 600 DL filte. A diffeent dichoic long-pass filte, a 645 DL, eflects the light between 605 nm and 645 nm to a 620 BP filte in font of the FL3 senso. The 620 BP filte tansmits light between 605 nm and 635 nm to the FL3 senso. The 645 DL filte tansmits light between 650 nm and 725 nm to a 675 BP filte in font of the FL4 senso. The 675 BP filte tansmits light between 660 nm and 700 nm to the FL4 senso. 3-8

31 OPERATION PRINCIPLES SIGNAL PROCESSING SIGNAL PROCESSING Voltage Pulse Signals The Cytomete has up to six sensos, each geneating a voltage pulse signal as each cell cosses the lase beam. A voltage pulse signal is popotional to the intensity of light the senso eceived. The Cytomete electonics amplifies, conditions, integates, and analyzes these pulses. Peak Signal Figue 3.6 shows how a peak voltage pulse signal foms as a cell cosses the lase beam. The intensity of light scatte o fluoescence detemines the height of the peak pulse (see Figue 3.6). The distibution of the fluoescence detemines the width of the pulse. Theefoe, the total fluoescence (intensity and distibution) detemines the aea unde the pulse. Figue 3.6 shows how thee cells with the same amount of total fluoescence but with diffeent fluoescence intensities, poduce diffeent peak pulses. Integal Signal Because the total fluoescence in all thee cells is the same, but the distibution is diffeent, the pulse can be integated to poduce an integal signal (see Figue 3.7). The height of the integal pulse is popotional to the total fluoescence and is obtained when the cell exits the lase beam. The pulse height, howeve, epesents the most intense amount of fluoescence poduced. The aea unde the pulse is popotional to the total fluoescence. 3-9

32 OPERATION PRINCIPLES SIGNAL PROCESSING Figue 3.6 Voltage Pulse Fomation, Peak Signal VOLTS LASER BEAM CELL TIME Cell entes lase beam; some light is scatteed. VOLTS TIME Cell is in cente of beam. Scatteed light, and theefoe pulse height, eaches a maximum. VOLTS PULSE HEIGHT TIME Cell leaves beam; scatteed light deceases

33 OPERATION PRINCIPLES SIGNAL PROCESSING 3 Figue 3.7 Integal and Peak Pulses CELL IN LASER BEAM DIRECTION OF SAMPLE FLOW PEAK PULSES VOLTS TIME INTEGRAL PULSES VOLTS TIME Amplification Some voltage pulses ae so weak that they must be amplified so that the chaacteistics of the cells that scatteed the light can be measued. The instument softwae lets you: Incease the gain to linealy amplify the integal and peak signals. Logaithmically tansfom the linea data. A logaithmic tansfomation accentuates the diffeences between the smalle pulses and educes the diffeences between the lage pulses. Signals Geneated The instument sensos FS, SS, FL1, FL2, FL3, and FL4 (optional) can geneate these signals (integal unless stated othewise; LOG stands fo logaithmic): FS, FS LOG, FS PEAK SS, SS LOG, SS PEAK FL1, FL1 LOG, FL1 PEAK FL2, FL2 LOG, FL2 PEAK FL3, FL3 LOG, FL3 PEAK FL4, FL4 LOG, FL4 PEAK (optional). Fo any given sample, the instument can geneate up to eight of these signals, plus the AUXiliay (AUX), Pism, TIME, and RATIO paametes. 3-11

34 OPERATION PRINCIPLES PARAMETERS 3.7 PARAMETERS AUXiliay Paamete The only way to acquie a peak signal is to assign it to the AUX paamete. Fo any given sample, only one of the following integal o peak signals can be assigned to AUX: FS, FS PEAK SS, SS PEAK FL1, FL1 PEAK FL2, FL2 PEAK FL3, FL3 PEAK FL4 (optional), FL4 PEAK (optional). When to Use the AUX Paamete When you specify both linea and logaithmic amplification of the same signal, the gain fo the linea amplification is also applied to the logaithmic amplification. Howeve, when you specify only logaithmic amplification of a signal, the instument automatically sets the gain fo that signal to 1.0, and you cannot change it. Use the AUX paamete when you want to: Amplify a signal at two diffeent gains. Obseve a peak signal o integal signal at the same time as the log signal. Obseve a peak signal at the same time as the integal signal. Also, the AUX paamete may be used fo doublet discimination. Assign a peak fluoescence signal to AUX so you can measue peak vs. integal fluoescence. TIME Paamete The TIME paamete is the amount of time, in seconds, the instument acquies data. It is displayed on the histogam axis in 1-second esolution. The axis labels vay, depending on histogam esolution and stop time. The minimum stop time is 10 seconds, the maximum stop time is 1,800 seconds (30 minutes) and the default stop time is 300 seconds (5 minutes). When you assign the TIME paamete to a histogam axis, the divisions on the axis change accodingly. To find the time (in seconds) pe channel in a one-paamete histogam, divide the stop time (in seconds) by 1,024 (0.001 second = 1.0 ms). Fo a two-paamete histogam, divide by 64, 128, o 256 depending upon the histogam esolution you ae using. 3-12

35 OPERATION PRINCIPLES PARAMETERS 3 When stop volume is used, the stop time is as shown in the ight-hand column and cannot be changed because the stop volume is eached long befoe the stop time elapses. Stop Volume 20 µl 40 µl 60 µl 80 µl 100 µl Stop Time 60 seconds 120 seconds 180 seconds 240 seconds 300 seconds RATIO Paamete The RATIO paamete is calculated, not acquied diectly. When you select a paamete, you specify which signal is the numeato and which is the denominato. Numeato RATIO = Denominato A atio of _1 is at channel 1,023. If you assign RATIO to a histogam axis, RATIO events appea at a lowe channel if the intensity of the numeato signal is less than the denominato signal. To calculate the actual atio at a paticula intensity fo a one-paamete histogam, divide the intensity by 1,024. Fo a two-paamete histogam, divide by 64, 128, o 256 depending upon the histogam esolution you ae using. PRISM Paamete Pism is used to analyze multicolo immunofluoescence samples. With multicolo immunofluoescence a cell is eithe positive o negative fo each of two, thee, o fou cell suface makes. A paticula combination is called a phenotype. The Pism paamete allows you to display pecentages on all phenotypic populations in a single histogam. It is a softwae paamete that can be acquied in eithe un time o listmode. Pism is available on up to fou paametes. TIME, RATIO, and Pism itself cannot be used fo the Pism paamete. All othe signals can be used fo the Pism paamete. Geneally, FL1, FL2, FL3, and FL4 ae used. A Pism histogam shows a spike o population fo each antibody combination with a pecent of the total that epesents the pecent of the total events in the Pism histogam. See Figue

36 OPERATION PRINCIPLES PARAMETERS Figue 3.8 Pism Histogam To use Pism, you must fist define egions that sepaate the negative and positive populations and assign these egions to the Pism Equation. See the heading, Pism Equation Regions, in the Getting Stated manual fo instuctions on assigning egions to the Pism Equation. 3-14

37 OPERATION PRINCIPLES AUTOSTANDARDIZATION PROTOCOLS AUTOSTANDARDIZATION PROTOCOLS These potocols ae un in an Autostandadization panel to automatically adjust the high voltage, gain, and compensation settings and to veify that the instument is unning coectly. The Quality Contol chapte in the Opeato s Guide has instuctions fo ceating and unning an Autostandadization panel. _A Potocols The _A potocols automatically adjust high voltage and gain settings. A thee-lette name ("xyz pefix) must follow the _A in the potocol name. The instument settings ae saved to the xyz.qcs file. Use the same thee lette pefix at the beginning of any othe potocol that you want updated with the standadized settings fom the Autostandadization panel. Fo example: _A3QP Flow-Set 3QP FITC/RD1/PC5 _Q3QP FITC/RD1/PC5 the autostandadization potocol fo thee-colo immunofluoescence. It ceates the 3QP.QCS baseline file. the pimay and contol potocol fo thee-colo immunofluoescence the QC potocol fo thee-colo immunofluoescence. It ceates the 3QPFITC.QCC quality contol statistics file. _A potocols use two egions to locate and move the peak of a Flow-Set Fluoosphees population to a specific channel, standadizing the Cytomete s light scatte and fluoescence intensity. One egion (the lage one) captues the population at its cuent position and a second egion (the smalle one) identifies the taget channel that the peak of the population will be moved to. _A potocols must be assigned as Pimay potocols in the Autostandadization panel. _C Potocols The _C potocols ae used to standadize fluoescent (colo) compensation. Compensation eagents and quad-stat egions ae used to automatically adjust colo compensation. _C potocols must be assigned as Seconday potocols in the panel. Up to fou can be used in a panel. _Q Potocols The _Q potocols ae used to geneate quality contol statistics and veify the Cytomete setup. Regions with QC in thei names located in _Q potocols poduce the quality contol statistics which ae stoed in the xyzabcd.qcc file elated to the _QxyzABCD potocol. Use the same fist thee lettes in the _Q potocol name ( xyz pefix) as in the _Axyz potocol. When assigning _Q potocols to a panel: _Q potocols can be assigned as Seconday o Contol potocols in a panel. Multiple _Q potocols can be assigned pe panel. Use a diffeent ABCD descipto afte the _Qxyz (whee xyz epesents the fist thee lettes and designates which baseline 3-15

38 OPERATION PRINCIPLES QUALITY CONTROL (QC) FEATURES file to use) to distinguish each potocol. Spaces and hyphens (-) ae not counted by the system as pat of the xyzabcd that follows the _Q in the name. Example of thee _Q potocols: _Q3QP 45/4/3, _Q3QP 45/8/3, _Q3QP 8/4/3. Use a _Q potocol to teminate the Autostandadization panel. Examples of Autostandadization Potocols and Panels Fist Panel: _Q3CL Flow-Check Second Panel: _A3CL Flow-Set _C1 FITC/RD1 CYTO-COMP _C3 RD1/PC5 CYTO-COMP _Q3CL 45/4/3 _Q3CL 45/8/3 3.9 QUALITY CONTROL (QC) FEATURES QC Templates Use the QC template to define what QC data appeas in the Levey-Jennings gaphs and data table. You can select fom voltages, gains, compensations, and QC egion statistics. QC templates ae only ceated fo _A, _C, o _Q potocols, one QC template pe potocol. The QC template needs to be loaded in the QC Template sceen in ode to view the QC data. Levey-Jennings Gaphs Use the Levey-Jennings gaph featue to automatically gaph the data fom the Cytomete settings baseline files (*.QCS) and QC statistical data files (*.QCC) ove a peiod of time. You define the data you want to appea in the gaph though the QC template. In the Levey-Jennings gaph, you can visually eview the QC data fo tends, shifts, outlies, and violations of you laboatoy s QC guidelines. In addition, you can equest the system to check the data against Westgad ules. Note: See the Data Management manual fo a detailed explanation of the *.QCS and *.QCC files. The Levey-Jennings gaph shows a Mean value, and the inne and oute limits. The oute limits ae 1.5 times the inne limits. Data points ae plotted in elation to the Mean and these limits. You can manually ente the Mean value o have the system calculate it. You detemine whethe to calculate and display the data points on the gaphs in elation to the Mean ±nsd, Mean ±numbe, o Mean ±% of Mean. Each gaph shows the data fo one QC egion statistic o one Cytomete setting signal depending upon the QC template definition. 3-16

39 OPERATION PRINCIPLES QUALITY CONTROL (QC) FEATURES 3 Data Table Use the Data Table to view the data fom the Cytomete settings baseline files (*.QCS) and QC statistical data files (*.QCC) in table fomat and to view the calculated statistics fo the file. You define the data you want to appea in the Data Table in the QC template. Runs can be deleted fom the calculated statistics, o you can select specific efeence uns and calculate the statistics based on them. Westgad Rules Statistical QC data fom the QC egions and instument settings can be checked against Westgad ules. You can view o pint the Westgad epot at the QC Levey-Jennings sceen and the QC Data Table sceen. When you equest the Westgad ules check at these two sceens, any failue of the ules appeas on the Westgad Repot sceen. Hee ae the six Westgad ules used to check QC data by the XL softwae: Rule 1. The point is above the Mean +2 SD o below the Mean -2 SD. Rule 2. The point is above the Mean +3 SD o below the Mean -3 SD. Rule 3. The next two points ae above the Mean +2 SD o below the Mean -2 SD. Rule 4. The point is above the Mean +2 SD, and the next point is below the Mean -2 SD, o vice vesa. Rule 5. The next fou points ae above the Mean +2 SD o below the Mean -2 SD. Rule 6. The next ten points ae above the Mean o below the Mean. Note: Westgad ules used in this system only apply to anges established with the calculated mean and standad deviation (SD). These ules do not apply to manually enteed o system calculated means with Mean ±numbe o Mean ±%. Fo moe detailed infomation on the Westgad ules, visit thei website, Compae Sceen Use this sceen to compae two diffeent QC files (*.QCS o *.QCC). Fo example, you can compae the cuent *.QCC data file fo a _Q potocol to a pevious *.QCC data file fo the same _Q potocol. Note: New *.QCS and *.QCC files ae ceated fo a potocol when you change the lot numbe in the potocol s lot numbe egion. So in the above example you would be compaing simila data fo the same potocol fo two diffeent lot numbes of contol mateial. Maintenance Sceen Use the Maintenance sceen to ecod daily statup and shutdown, and all maintenance pefomed on the XL/XL-MCL flow cytomete. 3-17

40 OPERATION PRINCIPLES HISTOGRAM DISPLAY 3.10 HISTOGRAM DISPLAY The esults of sample analysis appea on the Wokstation sceen as gaphs called histogams. You assign the paametes to the histogam axes. Histogams can be displayed in black and white o colo as: Single-paamete line Dual-paamete dot plot Dual-paamete contou Dual-paamete pojection Dual-paamete isometic Single-o dual-paamete Pism. Dual-paamete histogams can be displayed in 64 x 64, 128 x 128, o 256 x 256 esolution. Regions To analyze data o gate histogams, you must fist ceate and assign egions to these tasks. You can ceate five diffeent types of egions (see Ceating Regions in the Getting Stated manual). The egion types ae: Linea Rectilinea Numeic Quad-stat Amophous. Once a egion is ceated it can be assigned to function in a specific way (see Assigning Regions in the Getting Stated manual). The functions that you can assign a egion to ae: Analysis PRIME CAL (calibation) Cell-Seeke (autogating) Cell-Tacke (event coloing) Gating Listmode gating (LISTGATE) Pism Mapping Positives analysis QC (quality contol statistics) Lot numbe. Gating The softwae lets you use gating to specify that only cetain cells ae to be analyzed. A gate can be defined as the cells that ae inside o out of one o moe egions. 3-18

41 OPERATION PRINCIPLES HISTOGRAM DISPLAY 3 Data Stoage Sample esults can be pinted out, saved to a diskette o othe emoveable media, saved to a netwok dive, o expoted to anothe compute though a seial pot. Specimen infomation and patient epots can be saved to a database. You can stoe sample esults in the fom of a list of the measuements fom each cell, called listmode data. Listmode data can be eplayed into histogams o achived fo analysis late. Histogams can also be saved to a file. Histogam Statistics Note: Wheneve moe than 65,535 events accumulate in any one channel, ****** display instead of the statistics to indicate a channel oveflow. Linea Region Statistics Fo linea signals, statistics fo histogam egions ae calculated as follows: Pecent = Numbe of cells in egion Total numbe of cells in the egion Count o Aea = Numbe of cells in the egion Peak position = Intensity containing the lagest numbe of cells within the egion. Peak count = Numbe of cells in the peak position within the egion Mean = Σ( intensity numbe count in the position) aea SD = Σ[ ( intensity numbe mean) 2 count in the intensity] aea Fo Mean and SD, the summations ae pefomed ove all the channels that lie within the egion. sum( median intensity) aea Median = median intensity count in median intensity Median intensity = the smallest intensity such that sum (intensity) aea Sum (intensity) = sum of events fom the lowe edge of the egion to the intensity CV = SD Mean FPCV = CV width of peak at half the peak height Half Peak CV = peak position 3-19

42 OPERATION PRINCIPLES HISTOGRAM DISPLAY Log Region Statistics Fo log signals, the fomulas ae: Mean intensity = Σlog to lin (channel numbe) count in the channel aea Log to Lin (N) = N L+ D whee L = -1 fo fou-decade logaithmic amplification D = numbe of decades Log SD = Log to Lin SD Mean SD Log to Lin Mean Log CV = Log SD 100 Log Mean Log Mean = Log to Lin (Mean) Log Peak Position = Log to Lin (Peak Position) Log Median = Log to Lin (Median) 3-20

43 4SPECIFICATIONS/CHARACTERISTICS SAMPLE REQUIREMENTS At least 0.5 ml of pepaed sample is needed. It must be in a 12- x 75-mm test tube. Samples analyzed on the instument must be in a single-cell suspension. Typically, cells ae pepaed befoe they ae analyzed. The method used to pepae a specimen depends on the sample type and the assay desied. Fo example, the Q-Pep, Multi-Q Pep, o TQ-Pep wokstations let you pepae antibody-labeled cells fom an anticoagulated whole-blood specimen fo suface make analysis. In geneal, the optimum concentation fo analysis is 5 x 10 6 cells/ml. When this concentation is not possible, efe to the package inset fo the pepaation method you ae using. The instument can measue cells that ae between 0.5 µm and 40 µm in diamete. Fo fluoescent light measuements only, paticles can be macomolecula o up to 40 µm in diamete. 4.2 INSTRUMENT SPECIFICATIONS Dimensions Component Height Width Depth Weight Compute cm cm cm kg (6.2 in.) (21 in.) (16.75 in.) (40 lb) Cytomete with MCL Monito (typical) Powe Supply Installation Categoy Categoy II (pe IEC standad). Cytomete 50.8 cm (20 in.) same as above 42.7 cm (16.81 in.) 48.3 cm (19 in.) 61 cm (24 in.) 86.6 cm (34.1 in.) 40.5 cm (15.94 in.) cm (16 in.) 57.2 cm (22.5 in.) same as above 43.8 cm (17.25 in.) 50.8 cm (20 in.) 63.5 kg (140 lb) 84.8 kg (187 lb) 22 kg (48.4 lb) 54.4 kg (120 lb) Flow Cell Sensing aea: BioSense 250-µm squae channel with an integal lens, mounted with a vetical (upwad) flow path. Flow Rate Continuous pessue is applied to the sample tube. The amount of pessue depends on the flow ate you specify: Low Medium High 4-1

44 SPECIFICATIONS/CHARACTERISTICS INSTRUMENT SPECIFICATIONS Lase Ai-cooled, softwae contolled, 15 mw, agon ion lase opeating at 488 nm. Agon Lase Powe Lase powe is monitoed 5 times pe second within the system softwae and the eading is displayed on the STATUS ba of the cytomete softwae. If the lase powe deviates moe than ±1%, a Lase Powe Eo is displayed on the STATUS ba and the system will not un a sample until the eo message is no longe displayed. Follow the instuctions in the Toubleshooting section fo handling this eo. In ode to extend the life of the agon lase, when the flow cytomete is in idle mode, the lase powe will be less than 8mW. Once the instument is taken out of idle mode, the lase powe will etun to 20mW. Thus, while opeational and out of idle mode, the lase powe specifications ae 20±1%mW. Beam-Shaping Optics Coss cylindical lenses 10 mm by 80 mm. Lase Beam Spot Size An elliptical spot 10-µm high by 80-µm wide. Optical Filtes Neutal density (ND1) filte, if desied. 488-nm, 550-nm and 600-nm dichoic, long-pass (DL) filtes. 488-nm lase-blocking (BK) filte. 525-nm, 575-nm, and 620-nm band-pass (BP) filtes. 675-nm band-pass (optional). Sensos The FS senso and the SS senso ae photodiodes. The thee FL sensos ae photo-multiplie tubes that have a 200- to 800-nm spectal ange (fouth FL senso is optional). Signal Pocessing High voltage amplification, up to 2,000, in incements of 1, fo: FL1 FL2 FL3 FL4 (optional). Venie gain (fine amplification), up to 1,000 (labeled volts), in incements of 1, fo the following. A change of 1 to 1,000 epesents a 1-to-4 change in gain: FS SS AUX 4-2

45 SPECIFICATIONS/CHARACTERISTICS INSTRUMENT SPECIFICATIONS 4 Tue (total) gain is the esult of venie gain (HV) and linea amplification (gain) fo FS, SS and AUX. When HV is 100 and gain is 2.0, tue gain is 2.6. Tue Gain = Gain x [1+ (0.003 x HV)] Linea amplification (gain) by 1.0, 2.0, 5.0, 7.5, 10, 20, 50, 75, 100, 200, o 500 fo: FS SS. Linea amplification (gain) by 1.0, 2.0, 5.0, 7.5, 10, o 20 fo: FL1 FL2 FL3 FL4 (optional). Linea amplification (gain) by 1.0, 2.0, 5.0, 7.5, 10, 20 o 50 fo: AUX. Fou-decade digital logaithmic tansfomation of: FS SS FL1 FL2 FL3 FL4 (optional). Note: A scale of 0.1 to 1,000 is displayed on the histogam axes fo logaithmic paametes, but the statistics ae based on an actual scale of to Fluoescence colo compensation is available in 0.1 incements, fom 0 to 100%, fo: FL1 FL2 FL3 FL4 (optional). A disciminato (maximum value of 1,023) is available fo one of the following signals. Only one disciminato can be specified fo any one sample acquisition. FS SS FL1 FL2 FL3 FL4 (optional) AUX. Wokstation Compute Intel Pentium micopocesso and 16 MB of RAM. 4-3

46 SPECIFICATIONS/CHARACTERISTICS SOFTWARE SPECIFICATIONS Data Stoage 3.5-in., 1.44-MB diskette dive. 2-GB nonemovable had disk. Intefaces Bidiectional asynchonous seial intefaces fo communication with mainfame and pesonal computes. Input Devices Two-button mouse. 101-key IBM PC AT-compatible keyboad. Monito Colo (SVGA) with a 17-in. sceen. 4.3 SOFTWARE SPECIFICATIONS Data Output and Compatibility You can set up the instument fo multiple uses in the Utilities application. The Data Management sceen lets you specify local and off-line diectoies fo saving and etieving files. Othe output and compatibility featues include: FCS 2.0 file fomat fo listmode and histogam files. PCX fomat fo image files. ASCII file fomat fo conveted Expot (*.EPT) files. The ability to copy, move, and delete files. Full-colo pintouts with the appopiate optional Pinte. SQL database fo specimen infomation and patient epots. Automatic data achival. Pintout of sample esults and patient epots. Pintout of Woklist and caousel epot fo the MCL. Compatibility with ALTRA, Pofile, and Elite flow cytomete (without Pism, Gated Amp, o Time of Flight paametes) files. Compatibility with files fom pevious softwae vesions of XL and XL-MCL flow cytometes (1.0, 1.5, SYSTEM II 1.0, SYSTEM II 2.0, and SYSTEM II 2.1). The ability to expot specific line items fom a patient epot though a seial pot o to an ASCII Patient epot expot (*.PXP) file. Repoting Units Repoting units (absolute units) ae selected at the Utilities Configuation sceen. They can be eithe standad US (cells/ul) o SI units (cells/l). 4-4

47 SPECIFICATIONS/CHARACTERISTICS SOFTWARE SPECIFICATIONS 4 Two-Digit Date Fomat Fo Non Date-of-Bith Dates Fo two-digit date enties (othe than date-of-bith enties) if the yea enteed is: 80 to 99, the system assumes the yea is in the ange of 1980 to to 79, the system assumes the yea is in the ange of 2000 to Fo Date-of-Bith Dates Fo D. O. B. (date of bith) enties, if the yea enteed is: Equal to o less than the cuent yea, the system assumes the yea is in the cuent centuy. (Example: If it is cuently 1998 and you ente a DOB of 21JUN91, the system assumes the D.O.B yea is 1991.) Geate than the cuent yea, the system assumes the pevious centuy. (Example: If it is cuently 2001 and you ente a DOB of 13JUL02, the system assumes the D.O.B yea is 1902.) Acquisition Duing data acquisition, the histogams ae updated in eal time. When one histogam is displayed with statistics undeneath, the statistics ae also updated in eal time. Up to eight histogams ae available fo any given sample, and each histogam can have up to two paametes. One-paamete histogams have 1,024-channel esolution. Two-paamete histogams have up to 256- x 256-channel esolution. Regions Twenty-fou egions ae available fo gating, analysis, and autogating. Up to 16 (eight amophous) of those egions can be used at a time in histogam equations as gating egions. The following types of egions ae available fo gating and analysis: Linea Rectilinea Numeic Amophous Quad-stat (quadant statistic) [analysis only] You can select which statistics ae pinted fo each egion in a histogam. Listmode Analysis The instument can stoe up to eight paametes including AUX, TIME, and RATIO as listmode data. The amount of available andom access memoy (RAM) in the compute detemines the maximum size of a listmode file. 4-5

48 SPECIFICATIONS/CHARACTERISTICS PERFORMANCE SPECIFICATIONS Multigaph Analysis Multigaph analysis uses ovelay and galley displays to analyze o display up to eight histogams at a time. 4.4 PERFORMANCE SPECIFICATIONS Cayove Scatte and fluoescence cayove is less than 1% fom one specimen to anothe when the numbe of gated events is between 100 and 10,000. Data Acquisition Thoughput Fo two-colo suface makes, pocesses 60 tubes/hou of nomal whole-blood samples (2,500 lymphocytes/µl, 10,000 leukocytes/µl) when acquiing 10,000 lymphocytes at a ate of 1,000 leukocytes/second. The tubes wee pepaed on a Q-Pep wokstation o a Multi-Q-Pep wokstation. This assumes the use of a black and white lase pinte. Doublet Discimination Detects >90% of cellula doublets in cells _7 µm, using peak vs. integal discimination, depending upon sample pepaation method. Pecision fo Suface Makes See eagent package inset fo pecision specifications of othe suface makes. Resolution Fowad Scatte Within ±25% of the half-peak coefficient of vaiation (HPCV) assay value fo Flow-Check fluoosphees, without the ND1 filte. Geneally a HPCV of <3% is acceptable fo suface make applications and <2% is acceptable fo DNA applications. Fluoescence Within ±25% of the HPCV assay value fo Flow-Check fluoosphees, with optical filtes. Geneally a HPCV of <3% is acceptable fo suface make applications and <2% is acceptable fo DNA applications. Sensitivity Scatte Detects 0.5-µm diamete paticles. Fluoescence Less than 1,000 molecules of equivalent fluoochome when measued with FCSC micobeads and IsoFlow sheath fluid. 4-6

49 SPECIFICATIONS/CHARACTERISTICS PERFORMANCE CHARACTERISTICS 4 Stability Day-To-Day Peak intensity channel values of Flow-Check fluoosphees fo FS, FL1, FL2, FL3, and FL4 (optional) do not vay moe than ±5% fom the peak intensity obtained ove a peiod of 7 days, when the tempeatue does not vay moe than ±5 F fom tempeatue at alignment. Refe to the Flow-Check fluoosphees package inset fo details. Within Day Peak intensity channel values of Flow-Check fluoosphees fo FS, FL1, FL2, FL3, and FL4 (optional) do not vay moe than ±5% fom the peak intensity channel obtained within a peiod of 24 hous, when the tempeatue does not vay moe than ±5 F fom tempeatue at alignment. Refe to the Flow-Check fluoosphees package inset fo details. 4.5 PERFORMANCE CHARACTERISTICS Refe to the package inset fo the pefomance chaacteistics fo the pepaation method you ae using. 4.6 BAR-CODE SPECIFICATIONS Ba-Code Labels A ba code consists of black lines (bas) and white lines (spaces), which ae called elements. Thee ae naow elements (NE) and wide elements (WE). The ba-code symbology detemines thei aangement. IMPORTANT Sample misidentification can occu fom the use of incoect ba-code labels. Follow the specifications in this section to ceate you ba-code labels to pevent incoect sample identification. The instument suppots pepinted labels. Acceptable Ba Codes Within the given specifications, the MCL eade and the optional hand-held ba-code scanne automatically distinguish the following ba codes: Inteleaved 2-of-5 Code 39 ba code Codaba Code 128B Code 128C fixed 14 chaactes (13 data chaactes + 1 check chaacte, must be an even numbe of chaactes, leading zeos can be added) maximum 7 chaactes (6 data chaactes + 1 check chaacte) maximum 10 chaactes (9 data chaactes + 1 check chaacte maximum 8 alphanumeic chaactes maximum 16 numeic chaactes (The use of 15 numeic chaactes is invalid) Ba-Code Label Optical Chaacteistics at 670 nm ±10% Pint Contast Signal (PCS): 80% minimum. 4-7

50 SPECIFICATIONS/CHARACTERISTICS BAR-CODE SPECIFICATIONS Reflectivity of Media (RW): 80% minimum. Reflectivity of Ink (RB): 16% maximum. No spots o voids; no ink smeaing. Edge oughness is included in the ba and space toleances. RW + RB PCS = % RW Table 4.1 Code-Related Specifications Code Inteleaved 2-of-5* Codaba* Code 39* Code 128 * Naow element (NE) width 0.010" ±0.001" 0.010" ±0.001" 0.010" ±0.001" 0.010" ±0.001" Wide element/naow 3:1 N/A 3:1 N/A element atio (WE/NE) Intechaacte gap No 0.010" Minimum. _NE No Data digits 14** 1 to 10** 1 to 7** 2 to 16 * See AIM unifom Symbology specification, Rev fo detailed specification. ** Includes check sum chaacte NE Width 0.01 in. WE/NE Ratio 3:1 Pinting Methods Optional ba-code pinte. See Appendix A in the Special Pocedues and Toubleshooting manual. MCL Ba-Code Reade The MCL uses a visible-lase type eade containing a Class II lase, opeating at 670 nm, with a maximum powe output of 1 mw. Hand-Held Ba-Code Reade The hand-held ba-code eade uses a visible-lase type eade containing a Class II lase, opeating at 670 nm, with a maximum powe output of 1 mw. Ba-Code Decode The MCL sends a GS ASCII chaacte (hexadecimal 1D) to the decode to stat opeation. The decode: Tuns the eade on. Decodes infomation that comes fom the eade. Keeps the eade on fo up to 4 seconds. Tuns the eade off. Sends the decoded infomation (o no-ead message) to the MCL. 4-8

51 SPECIFICATIONS/CHARACTERISTICS BAR-CODE SPECIFICATIONS 4 IMPORTANT To pevent incoect indentification of sample tubes, do not use FNC1, FNC4, and FS (hexadecimal 1C) chaactes in you ba-code infomation. Check Sum Algoithm Beckman Coulte stongly ecommends the use of ba code check sums to povide automatic checks fo ead accuacy. IMPORTANT Use of ba codes is an extemely accuate and effective method of positive patient identification. Cetain featues, such as check sum digits, maximize accuacy in eading Codaba, Code 39 and Inteleaved 2-of-5 labels. In one study, the use of check sum digits detected 97% of misead eos. Use checksums to povide potection against occasional misead eos caused by poblems such as damaged o misapplied labels. If you must use ba codes without check sums, Beckman Coulte ecommends that you veify each ba-code eading to assue coect patient identification. 4-9

52 SPECIFICATIONS/CHARACTERISTICS BAR-CODE SPECIFICATIONS 4-10

53 5PRECAUTIONS AND HAZARDS LASER SAFETY The Cytomete and the optional MCL each contain a lase. Beckman Coulte's design and manufactue of the instument complies with the equiements govening the use and application of a lase as specified in egulatoy documents issued by the: U.S. Depatment of Health and Human Sevices and Cente fo Devices and Radiological Health (CDRH). In compliance with these egulatoy documents, evey measue has been taken to ensue the health and safety of uses and laboatoy pesonnel fom the possible danges of lase use. Use the instument accoding to the infomation in the manuals. 5.2 WARNINGS Use of contols o adjustments o pefomance of pocedues othe than those specified heein might esult in hazadous adiation exposue. To ensue you safety, the Cytomete lase is coveed with potective shields. Do not emove these shields. No use-seviceable assemblies ae accessible. Do not attempt to emove the lase o open it. The instument has components that ae dangeous to the opeato. If any attempt has been made to defeat a safety featue, o if the instument fails to pefom as descibed in its manuals, disconnect the powe and call you Beckman Coulte Repesentative. 5.3 WARNING LABELS CDRH-equied waning labels ae placed nea o on coves that, if emoved, might expose lase adiation. They ae also placed nea openings that, if looked into, might expose you to lase adiation. See Figue 5.1 fo the Sensing Compatment cove waning label. See Figues 5.2 and 5.3 fo the Optical aea waning labels. See Figue 5.4 fo the Lase head waning labels. See Figue 5.5 fo the Cytomete back panel lase label. See Figue 5.6 fo the MCL pobe housing waning labels. See Figue 5.7 fo the MCL ba-code eade waning labels. 5-1

54 FS SS AUX FL4 TM PRECAUTIONS AND HAZARDS WARNING LABELS Figue 5.1 Lase Labels on the Sensing Compatment Cove CYTOMETER READY LASER ON FL1 SAMPLE FLOW FL2 LOW MED HIGH FL C Figue 5.2 Lase Labels in the Optical Aea, Side View C 5-2

55 READY ON FS SS AUX FL1 FL2 FL3 FL4 TM PRECAUTIONS AND HAZARDS WARNING LABELS 5 Figue 5.3 Lase Labels in the Optical Aea, Font View CYTOMETER LASER SAMPLE FLOW LOW MEDHIGH AVOID EXPOSURE AVOID EXPOSURE VISIBLE AND/OR INVISIBLE LASER RADIATION IS EMITTED FROM THIS APERTURE. VISIBLE AND/OR INVISIBLE LASER RADIATION WHEN OPEN AVOID EYE OR SKIN EXPOSURE TO DIRECT OR SCATTERED RADIATION C Figue 5.4 Lase Labels on the Lase Head C 5-3

56 PRECAUTIONS AND HAZARDS WARNING LABELS Figue 5.5 Lase Label on the Cytomete Back Panel 5-4

57 PRECAUTIONS AND HAZARDS WARNING LABELS 5 Figue 5.6 Lase Labels on the MCL Pobe Housing Cove AVOID EXPOSURE VISIBLE AND/OR INVISIBLE LASER RADIATION IS EMITTED FROM THIS APERTURE. CLASS 1 LASER PRODUCT CAUTION LASER LIGHT DO NOT STARE INTO BEAM 670 nm DIODE LASER 1.0 MILLIWATT MAXUMCLASS II LASER PRODUCT CAUTION Lase adiation when open and intelock defeated. DO NOT STARE INTO BEAM C 5-5

58 PRECAUTIONS AND HAZARDS WARNING LABELS Figue 5.7 Figue 20 Lase Labels on the MCL Ba-Code Reade DANGER LASER LIGHT WHEN OPEN. AVOID DIRECT EYE EXPOSURE. 939 INDUSTRY DR. TUKWILLA WA CLASS PRODUCT CONFORMS TO DHHS 21 CFR SUBCHAPTER J MADE IN THE USA MS 510 MODEL I I SERIAL NUMBER MANUFACTURED FIS PART NO. CAUTION LASER LIGHT. DO NOT STARE INTO BEAM 670 nm DIODE LASER 1.0 MILLIWATT MAXIMUM CLASS II LASER PRODUCT C 5-6

59 ALOG SHEETS A This appendix contains the following log sheets: Reagent Log Action Log Flow-Set Fluoosphees Chats t Daily Log fo Instument Standadization t Establishing HV/Total Gain Ranges t Establishing Fluoescence and/o Light Scatte Taget Ranges Flow-Check Fluoosphees Chats t Daily Log fo Instument Veification of Alignment and Fluidics t Establishing Peak Position and HPCV Taget Ranges Make photocopies as needed. A-1

60 LOG SHEETS A-2

61 LOG SHEETS A REAGENT LOG Reagent Name Date Opened Lot Numbe Expiation Date Tech Seial No. Lab. COULTER EPICS XL Flow Cytomete COULTER EPICS XL-MCL Flow Cytomete C A-3

62 LOG SHEETS A-4

63 LOG SHEETS A ACTION LOG Date Condition Noted Tech Date Action Taken Tech Seial No. Lab. COULTER EPICS XL Flow Cytomete COULTER EPICS XL-MCL Flow Cytomete C A-5

64 LOG SHEETS A-6

65 LOG SHEETS A DAILY LOG FOR INSTRUMENT STANDARDIZATION Flow-Set Fluoosphees Application Lot Numbe Expiation Date Instument HV/Total Gain Taget Ranges: FS SS/LOG SS LOG FL1 LOG FL2 LOG FL3 LOG FL4 FS SS o LOG SS Run Peak HV Gain/ Total Gain* Peak HV Gain/ Total Gain* *Recod total gain if using autostandadization. LOG FL1 Peak HV LOG FL2 Peak HV LOG FL3 Peak HV LOG FL4 Peak HV Tech/ Date Seial No. Lab. COULTER EPICS XL Flow Cytomete COULTER EPICS XL-MCL Flow Cytomete C A-7

66 LOG SHEETS A-8

67 LOG SHEETS A ESTABLISHING HV/TOTAL GAIN RANGES Flow-Set Fluoosphees Application Lot Numbe Expiation Date Peak Intensity Taget Ranges: FS SS/LOG SS LOG FL1 LOG FL2 LOG FL3 LOG FL4 FS Run Peak HV Aveage HV/Gain Gain/ Total Gain* SS o LOG SS Gain/ Total Peak HV Gain* LOG FL1 Peak HV LOG FL2 Peak HV LOG FL3 Peak HV LOG FL4 Peak HV Tech/ Date Aveage +2SD o +1% Aveage -2SD o -1% *Recod total gain if using autostandadization. Seial No. Lab. COULTER EPICS XL Flow Cytomete COULTER EPICS XL-MCL Flow Cytomete C A-9

68 LOG SHEETS A-10

69 LOG SHEETS A ESTABLISHING FLUORESCENCE AND/OR LIGHT SCATTER TARGET RANGES Flow-Set Fluoosphees Application Lot Numbe Expiation Date FS Run Peak HV Gain/ Total Gain* SS o LOG SS Gain/ Total Peak HV Gain* LOG FL1 Peak HV LOG FL2 Peak HV LOG FL3 Peak HV LOG FL4 Peak HV Tech/ Date n Aveage Channel *Recod total gain if using autostandadization. See package inset fo diections on how to detemine the taget ange fo each paamete. Seial No. Lab. COULTER EPICS XL Flow Cytomete COULTER EPICS XL-MCL Flow Cytomete C A-11

70 LOG SHEETS A-12

71 LOG SHEETS A DAILY LOG FOR INSTRUMENT VERIFICATION OF ALIGNMENT AND FLUIDICS Flow-Check Fluoosphees Lot Numbe Expiation Date Taget Range FS FL1 FL2 FL3 FL4 Run Peak HPCV Peak HPCV Peak HPCV Peak HPCV Peak HPCV Tech/Date Seial No. Lab. COULTER EPICS XL Flow Cytomete COULTER EPICS XL-MCL Flow Cytomete C A-13

72 LOG SHEETS A-14

73 LOG SHEETS A ESTABLISHING PEAK POSITION AND HPCV TARGET RANGES Flow-Check Fluoosphees Lot Numbe Expiation Date FS FL1 FL2 FL3 FL4 Run Peak HPCV Peak HPCV Mean SD +2SD -2SD Peak HPCV Peak HPCV Peak HPCV Tech/Date Seial No. Lab. COULTER EPICS XL Flow Cytomete COULTER EPICS XL-MCL Flow Cytomete C A-15

74 LOG SHEETS A-16

75 BPREDEFINED PANELS, PROTOCOLS, AND QC TEMPLATES B This Appendix contains Tables 2.1 though 2.6 which descibe the panels, potocols, and QC templates included with vesion 3.0 of the SYSTEM II softwae. These ae geneic templates that you can use as is, o modify fo you specific application needs. The potocols listed in theses panels can also be un sepaately o combined to fom othe panels. Table 2.1 Geneal Use Panels and Potocols Panel Name Cleaning Potocol Name (and QC Template Name if applicable) BLEACH Rinse Rinse Rinse Desciption Use this panel as the cleaning pocedue at the end of a shift o application as pe you laboatoy pocedue. N/A CLENZ Use this potocol as a single potocol cleaning pocedue. N/A _Axyz Flow-Check* Use this potocol to set up the taget Cytomete settings fo the Flow-Check Fluoosphees. N/A _Qxyz Flow-Check* Use this potocol afte the taget Cytomete settings have been set fo a specific lot of Flow-Check Fluoosphees. The nxyz.qcs baseline file updates the Cytomete settings when unning this potocol. * Each _A and _Q potocol listed in this table has a coesponding QC template pedefined in the softwae. The QC template has the same name as the potocol and esides in the same diectoy. Table 2.2 Two-Colo Application Panels and Potocols Panel Name 2 Colo Auto Setup Potocol Name (and QC Template Name if applicable) _A2CL Flow-Set* _C1 FITC/RD1 CYTO-COMP* _Q2CL CD45-FITC/CD14-RD1* _Q2CL MsIgG1-RD1/MsIgG1-FITC* _Q2CL 1 CD3-FITC/CD4-RD1* _Q2CL 2 CD3-FITC/CD8-RD1* _Q2CL 3 CD3-FITC/CD19-RD1* _Q2CL 4 CD3-FITC/CD56-RD1* Desciption _ Adjusts and monitos the Cytomete settings. _ Adjusts the compensation pecentages. _ The _Q templates in this panel monito the % positive, mean channel, and absolute counts (if using Flow-Count fluoosphees) fo the petinent egions in each potocol. Use the potocols to analyze contol cells. Modify the panel and potocols as needed fo you application. N/A _Q2CL FITC/RD1* Use this potocol to ceate additional 2 Colo Auto Setup potocols fo diffeent antibody combinations fo the contol cells. Subsequently, additional _Q potocols can be added to the oiginal 2 Colo Auto Setup Panel, as you applications equie. B-1

76 PREDEFINED PANELS, PROTOCOLS, AND QC TEMPLATES Table 2.2 Two-Colo Application Panels and Potocols (Continued) N/A _Q2CL Kappa-FITC/CD19-RD1* Add this potocol to the 2 Colo Auto Setup Panel if you application includes Kappa analysis. N/A _Q2CL Lambda-FITC/CD19-RD1* Add this potocol to the 2 Colo Auto Setup Panel if you application includes Lambda analysis. 2 Colo Analysis 2CL MsIgG1-RD1/MsIgG1-FITC 2CL CD45-FITC/CD14-RD1 2CL CD3-FITC/CD4-RD1 2CL CD3-FITC/CD8-RD1 2CL CD3-FITC/CD19-RD1 2CL CD3-FITC/CD56-RD1 Use the 2 Colo Analysis panel to analyze you two-colo applications. Modify the panel and potocols as needed fo you application. The 2CL.qcs baseline file updates the Cytomete settings when unning this panel. N/A 2CL FITC/RD1 Use this potocol to ceate additional 2 Colo Analysis potocols fo diffeent antibody combinations. Subsequently, these additional potocols can be added to the oiginal 2 Colo Analysis Panel as you applications equie. These potocols may be un alone o combined to ceate panels specific to you applications. The 2CL.qcs baseline file updates the Cytomete settings when unning this potocol. N/A 2CL Kappa-FITC/CD19-RD1 Add this potocol to the 2 Colo Analysis panel if you application includes Kappa analysis.the 2CL.qcs baseline file updates the Cytomete settings when unning this potocol. N/A 2CL Lambda-FITC/CD19-RD1 Add this potocol to the 2 Colo Analysis panel if you application includes Lambda analysis. The 2CL.qcs baseline file updates the Cytomete settings when unning this potocol. * Each _A, _C, and _Q potocol listed in this table has a coesponding QC template pedefined in the softwae. The QC template has the same name as the potocol and esides in the same diectoy. B-2

77 PREDEFINED PANELS, PROTOCOLS, AND QC TEMPLATES B Table 2.3 Thee-Colo Application [4 PMT System] Panels and Potocols Panel Name Potocol Name (and QC Template Name if applicable) Desciption 3 Colo Auto Setup [4 PMT System] _A3CL Flow-Set [4 PMT]* _C1 FITC/RD1 CYTO-COMP* _C3 RD1/PC5 CYTO-COMP [3CL-4PMT]* _Q3CL 1 45-FITC/G1RD1/G1PC5 [4]* _Q3CL 3 45-FITC/4-RD1/3-PC5 [4]* _Q3CL 5 45-FITC/8-RD1/3-PC5 [4]* _Q3CL 7 45-FITC/19-RD1/3PC5 [4]* _Q3CL 9 45-FITC/56-RD1/3PC5 [4]* _ Adjusts and monitos the Cytomete settings. _ Adjusts the compensation pecentages. _ The _Q templates in this panel monito the % positive, mean channel, and absolute counts (if using Flow-Count fluoosphees) fo the petinent egions in each potocol. Use the potocols to analyze contol cells. Modify the panel and potocols as needed. The 3CL.qcs baseline file updates the Cytomete settings when unning this panel. N/A _A3ECD Flow-Set Adjusts and monitos Cytomete settings fo thee-colo immunophenotyping using FITC, RD1, and ECD on eithe a 3 PMT o 4 PMT system. N/A _Q3ECD 2 FITC/RD1/ECD [4]* Use this potocol to ceate additional 3 Colo Auto Setup potocols fo diffeent antibody combinations of FITC, RD1, and ECD fo contol cells. Subsequently, additional _Q potocols can be added to the oiginal 3 Colo Auto Setup Panel, as you applications equie. The 3CL.qcs baseline file updates the Cytomete settings when unning this potocol. N/A _Q3CL A4 FITC/RD1/PC5 [4]* Use this potocol to ceate additional 3 Colo Auto Setup potocols fo diffeent antibody combinations of FITC, RD1, and PC5 fo contol cells. Subsequently, additional _Q potocols can be added to the oiginal 3 Colo Auto Setup Panel, as you applications equie. The 3CL.qcs baseline file updates the Cytomete settings when unning this potocol. N/A _Q3CL B2 G1-FITC/G1-RD1/G1-PC5 [4]* Add this potocol to the 3 Colo Auto Setup Panel if you application includes this antibody. Modify the potocol as needed fo you application. The 3CL.qcs baseline file updates the Cytomete settings when unning this potocol. N/A _Q3CL B4 8-FITC/4-RD1/3-PC5 [4]* Add this potocol to the 3 Colo Auto Setup Panel if you application includes this antibody. Modify the potocol as equied fo you application. The 3CL.qcs baseline file updates the Cytomete settings when unning this potocol. B-3

78 PREDEFINED PANELS, PROTOCOLS, AND QC TEMPLATES Table 2.3 Thee-Colo Application [4 PMT System] Panels and Potocols (Continued) 3 Colo Analysis [4 PMT System] 3CL 45-FITC/G1-RD1/G1-PC5 [4] 3CL 45-FITC/4-RD1/3-PC5 [4] 3CL 45-FITC/8-RD1/3-PC5 [4] 3CL 45-FITC/19-RD1/3-PC5 [4] 3CL 45-FITC/56-RD1/3-PC5 [4] Use the 3 Colo Analysis panel to analyze you thee-colo applications. Modify the panel and potocols as needed fo you application.the 3CL.qcs baseline file updates the Cytomete settings when unning this panel. N/A 3ECD FITC/RD1/ECD [4] Use this potocol to ceate additional 3 Colo Analysis potocols fo diffeent antibody combinations of FITC, RD1, and ECD. Subsequently, these additional potocols can be added to the oiginal 3 Colo Analysis Panel as you applications equie. These potocols can be un alone o combined to ceate panels specific to you applications. The 3CL.qcs baseline file updates the Cytomete settings when unning this potocol. N/A 3CL FITC/RD1/PC5 [4] Use this potocol to ceate additional 3 Colo Analysis potocols fo diffeent antibody combinations of FITC, RD1, and PC5. Subsequently, these additional potocols can be added to the oiginal 3 Colo Analysis Panel as you applications equie. These potocols can be un alone o combined to ceate panels specific to you applications. The 3CL.qcs baseline file updates the Cytomete settings when unning this potocol. N/A 3CL 8-FITC/4-RD1/3-PC5 [4] Add this potocol to the 3 Colo Analysis panel o use as a single potocol if you application includes analysis of this antibody combination. Modify the potocol as needed fo you application. The 3CL.qcs baseline file updates the Cytomete settings when unning this potocol. N/A 3CL G1-FITC/G1-RD1/G1-PC5 [4] Add this potocol to the 3 Colo Analysis panel o use as a single potocol if you application includes analysis of this antibody combination. Modify the potocol as needed fo you application. The 3CL.qcs baseline file updates the Cytomete settings when unning this potocol. * Each _A, _C, and _Q potocol listed in this table has a coesponding QC template pedefined in the softwae. The QC template has the same name as the potocol and esides in the same diectoy. B-4

79 PREDEFINED PANELS, PROTOCOLS, AND QC TEMPLATES B Table 2.4 Thee-Colo Application [3PMT System] Panels and Potocols Panel Name Potocol Name (and QC Template Name if applicable) Desciption 3 Colo Auto Setup [3 PMT System] _A3CL Flow-Set [3 PMT]* _C1 FITC/RD1 CYTO-COMP* _C3 RD1/PC5 CYTO-COMP [3CL-3PMT]* _Q3CL 0 45-FITC/G1RD1/G1PC5 [3]* _Q3CL 2 45-FITC/4-RD1/3-PC5 [3]* _Q3CL 4 45-FITC/8-RD1/3-PC5 [3]* _Q3CL 6 45-FITC/19-RD1/3PC5 [3]* _Q3CL 8 45-FITC/56-RD1/3PC5 [3]* _ Adjusts and monitos the Cytomete settings. _ Adjusts the compensation pecentages. _ The _Q Templates in this panel monito the % positive, mean channel, and absolute counts (if using Flow-Count fluoosphees) fo the petinent egions in each potocol. Use these potocols to analyze contol cells. Modify the panel and potocols as equied fo you application. The 3CL.qcs baseline file updates the Cytomete settings when unning this panel. N/A _Q3ECD 1 FITC/RD1/ECD [3] 620 BP* Use this potocol to ceate additional 3 Colo Auto Setup potocols fo diffeent antibody combinations of FITC, RD1, and ECD. Subsequently, additional _Q potocols can be added to the oiginal 3 Colo Auto Setup Panel, as you applications equie. The 3CL.qcs baseline file updates the Cytomete settings when unning this potocol. N/A _Q3CL A3 FITC/RD1/PC5 [3] 675 BP* Use this potocol to ceate additional 3 Colo Auto Setup potocols fo diffeent antibody combinations of FITC, RD1, and PC5. Subsequently, additional _Q potocols can be added to the oiginal 3 Colo Auto Setup Panel as you applications equie. The 3CL.qcs baseline file updates the Cytomete settings when unning this potocol. N/A _Q3CL B1 G1-FITC/G1-RD1/G1-PC5 [3]* Add this potocol to the 3 Colo Auto Setup Panel if you application includes this antibody. Modify the potocol as needed fo you application. The 3CL.qcs baseline file updates the Cytomete settings when unning this potocol. N/A _Q3CL B3 8-FITC/CD4-RD1/CD3-PC5[3]* Add this potocol to the 3 Colo Auto Setup Panel if you application includes this antibody. Modify the potocol as needed fo you application. The 3CL.qcs baseline file updates the Cytomete settings when unning this potocol. 3 Colo Analysis [3 PMT System] 3CL 45-FITC/G1-RD1/G1-PC5 [3] 3CL 45-FITC/4-RD1/3-PC5 [3] 3CL 45-FITC/8-RD1/3-PC5 [3] 3CL 45-FITC/19-RD1/3-PC5 [3] 3CL 45-FITC/56-RD1/3-PC5 [3] Use the 3 Colo Analysis panel to analyze you thee-colo applications. Modify the panel and potocols as needed fo you application. The 3CL.qcs baseline file updates the Cytomete settings when unning this panel. B-5

80 PREDEFINED PANELS, PROTOCOLS, AND QC TEMPLATES Table 2.4 Thee-Colo Application [3PMT System] Panels and Potocols (Continued) N/A 3ECD FITC/RD1/ECD [3] 620 BP Use this potocol to ceate additional 3 Colo Analysis potocols fo diffeent antibody combinations of FITC, RD1, and ECD. Subsequently, these additional potocols can be added to the oiginal 3 Colo Analysis Panel, as you applications equie. These potocols can be un alone o combined to ceate panels specific to you applications. The 3CL.qcs baseline file updates the Cytomete settings when unning this potocol. N/A 3CL FITC/RD1/PC5 [3] 675 BP Use this potocol to ceate additional 3 Colo Analysis potocols fo diffeent antibody combinations of FITC, RD1, and PC5. Subsequently, these additional potocols can be added to the oiginal 3 Colo Analysis Panel, as you applications equie. These potocols can be un alone o combined to ceate panels specific to you applications. The 3CL.qcs baseline file updates the Cytomete settings when unning this potocol. N/A 3CL 8-FITC/4-RD1/3-PC5 [3] Add this potocol to the 3 Colo Analysis panel o use as a single potocol if you application includes analysis of this antibody combination. Modify the potocol as equied fo you application. The 3CL.qcs baseline file updates the Cytomete settings when unning this potocol. N/A 3CL G1-FITC/G1-RD1/G1-PC5 [3] Add this potocol to the 3 Colo Analysis panel o use as a single potocol if you application includes analysis of this antibody combination. Modify the potocol as needed fo you application. The 3CL.qcs baseline file updates the Cytomete settings when unning this potocol. * Each _A, _C, and _Q potocol listed in this table has a coesponding QC template pedefined in the softwae. The QC template has the same name as the potocol and esides in the same diectoy. B-6

81 PREDEFINED PANELS, PROTOCOLS, AND QC TEMPLATES B Table 2.5 Fou-Colo Application Panels and Potocols Panel Name 4 Colo Auto Setup 4 Colo Analysis Potocol Name (and QC Template Name if applicable) _A4CL Flow-Set* _C1 FITC/RD1 CYTO-COMP* _C2 RD1/ECD CYTO-COMP* _C3 RD1/PC5 CYTO-COMP [4CL]* _C4 ECD/PC5 CYTO-COMP* _Q4CL FITC/RD1/ECD/PC5* 4CL FITC/RD1/ECD/PC5 Desciption _ Adjusts and monitos the Cytomete settings. _ Adjusts the compensation pecentages. _ The _Q Templates in this panel monito the % positive, mean channel, and absolute counts (if using Flow-Count fluoosphees) fo the petinent egions in each potocol. Modify the panel and potocols as needed fo you application. Use these potocols to analyze contol cells. The 4CL.qcs baseline file updates the Cytomete settings when unning this panel. Use the 4 Colo Analysis panel to analyze you fou-colo applications. Modify the panel as equied fo you application. The 4CL.qcs baseline file updates the Cytomete settings when unning this panel. * Each _A, _C, and _Q potocol listed in this table has a coesponding QC template pedefined in the softwae. The QC template has the same name as the potocol (but a diffeent extension) and esides in the same diectoy. Table 2.6 DNA Application Panels and Potocols Panel Name Potocol Name (and QC Template Name if applicable) Desciption N/A _ADNA INDEX Contol* Use this Autostandadization potocol to adjust the Cytomete settings fo DNA analysis. Use the QC template to monito the Cytomete settings. N/A _QDNA INDEX Contol* Use this potocol to collect and monito DNA INDEX Contol pecentages and mean channels. The dna.qcs baseline file updates the Cytomete settings when unning this potocol. N/A DNA Analysis Use this potocol to analyze you DNA specimens. Modify as needed fo you application. The dna.qcs file updates the Cytomete settings when unning this potocol. * Each _A and _Q potocol listed in this table has a coesponding QC template pedefined in the softwae. The QC template has the same name as the potocol and esides in the same diectoy. B-7

82 PREDEFINED PANELS, PROTOCOLS, AND QC TEMPLATES B-8

83 GLOSSARY _A potocols - These potocols ae used to automatically adjust high voltage and gain settings. One is always assigned as a pimay potocol in an Autostandadization panel so that the settings ae passed to othe potocols in the panel. Accuacy - The ability of an instument to agee with a pedetemined efeence value at any point within the opeating ange. Contast with pecision. ASCII - Abbeviation fo Ameican Standad Code fo Infomation Intechange. An ASCII file is a type of text file. Assay values - Values fo a contol established by extensive epeat testing of that contol. AUX Signal - Auxillay acquisition pathway that allows eithe contol of simultaneous Lin and Log signals o acquisition of a Peak signal. Backgound count - Measue of the amount of electical o paticle intefeence. BK filte - A lase-blocking optical filte that passes the fluoescence wavelengths but does not pass the lase wavelength. BP filte - A band-pass optical filte that passes a band of wavelengths and blocks othes. Button - The Wokstation sceens have named aeas (fo example, a ectangle labeled Abot) that you select with the mouse to tell the instument what to do. The Cytomete has buttons that you pess to specify the opeating mode (fo example, the RUN button). _C potocols - These potocols ae used to standadize fluoescent compensation. They ae always assigned as seconday potocols in an autostandadization panel, following the _Axyz potocol. CAL Facto - A numbe used in conjunction with a known numbe of paticles identified by a CAL egion, that adjusts the egion counts obtained. Cell Seeke - 3 Levels of AutoGating. Cell Stat - Algoithm that pefoms automatic gating and analysis used in tetaone System. Cell Tacke - Colo event analysis with geen, blue, ed, and magenta colos assigned to egions and gates. Channel - In an analog-to-digital convete, the numbe of equally spaced divisions of the amplified input signal voltage. All XL and XL-MCL flow cytomete signals ae esolved into 1024 channels. Fo dual-paamete histogams, the numbe of channels is educed to 64, 128, o 256. Cleaning agent - A detegent used to flush sample fom tubing and eliminate potein buildup. Click - To pess and elease a mouse button. Coefficient of vaiation (CV%) - A measue of the vaiability in signal intensity that is geneated as paticles pass epeatedly though the lase beam. This vaiability is expessed as a pecentage of the aveage signal intensity. Collimate - To make paallel (fo example, collimate ays of light). GLOSSARY-1

84 GLOSSARY Colo compensation - The subtaction of: a pecentage of the signal fom one fluoescence light senso fom the signal fom anothe fluoescence light senso to coect fo the ovelap of one dye's emission into anothe dye's emission measuement. Contol - A substance used to outinely monito the pefomance of an analytical pocess that does not have the chaacteistic being measued (fo example, Immuno-Tol cells o CYTO-TROL contol cells). Contols and indicatos - Instument contols ae the mechanisms you use to communicate with the instument. Indicatos ae the mechanisms the instument uses to communicate with you. Contols and Indicatos is the fist chapte in the Getting Stated manual. Coss-cylindical lenses - Used in the Cytomete to focus the lase beam and fom an elliptical beam spot. Cytomete - The system component that analyzes the sample and contains the sheath fluid and cleaning agent bottles. Database file - SQL fomat file that contains ecods of the patient epot. Database ecod - Individual patient panel epot esiding in the database file. dc - Abbeviation fo diect cuent. Defaults - Oiginal settings fo the instument. You can change them to customize the settings fo you laboatoy. Disciminato - A channel setting fo a paamete that lets you ignoe events below the setting. This lets you eliminate signals caused by debis. Display Options - Acquisition button that changes cytomete adjustment options in the Cytomete Contol Window. DL filte - A dichoic, long-pass optical filte that diects light in diffeent spectal egions to diffeent detectos. DiOC5(3) - Abbeviation fo oxacabocyanine dye. EPT2ASC2 - Pogam that convets Expot (*. EPT) files to ASCII (*. EPA) files and (*. CYT) files to ASCII (*. CYA) files. Expot file (*.EPT) - Binay file geneated by unning a panel that contains the egion statistics. Event - A paticle passing though the lase beam. Fast Set - Diect histogam manipulation to adjust cytomete voltages and gain. Fast Comp - Diect histogam manipulation to adjust cytomete colo compensation. GLOSSARY-2

85 GLOSSARY FDA - Abbeviation fo fluoescein diacetate dye. File Timming - Pocess that deletes data fom the database file and fom the QC histoy file by a date olde than the time peiod you ente when to tim Auto Setup Recods in the Data Management sceen. Used to minimize the memoy equiements of these files. FITC - Abbeviation fo fluoescein isothiocyanate dye. Flow cell - A device though which paticles pass, in a steam of fluid, one at a time, though a lase beam. Flow cytomety - A pocess fo measuing the chaacteistics of cells o othe biological paticles as they pass though a measuing appaatus in a fluid steam. Fluoescent light - The emission of electomagnetic adiation that occus when the emitting body absobs adiation fom some othe souce. Fo example, when a fluoescent dye is excited (absobs adiation), it emits fluoescent light at a wavelength that is diffeent fom the wavelength of the light that excited it. Fluoescent light (FL1, FL2, FL3, FL4 optional) sensos - Collect the fluoescent light and geneate voltage pulse signals. The "1" efes to the fist fluoescence senso; "2" the second; and so foth. Fowad scatte (FS) - The lase light scatteed at naow angles to the axis of the lase beam. The amount of fowad scatte is popotional to the size of the cell that scatteed the lase light. Fowad scatte (FS) senso - Collects the fowad scatte and geneates voltage pulse signals. Gain - The amount of amplification applied to a signal. In linea amplification, all of a senso's signals ae inceased by the same amount. Contast with logaithmic amplification. Gating - The use of citeia that must be met befoe an event is included in a histogam. GB - The abbeviation fo gigabyte. High voltage - Can be adjusted to change the sensitivity of a fluoescent light senso. Histogam - A gaph showing the elative numbe and distibution of events. Hist Mode - Multigaph analysis of individual histogams fom diffeent un numbes. Hot keys - A keyboad shotcut fo changing sceens. Instead of using the menu ba to change sceens, you can pess and hold down Þ while pessing a cetain lette key. Fo example, pessing Þ and C simultaneously displays the Cytosettings sceen. Hydodynamic focusing - A pocess that focuses the sample steam though the flow cell. It ensues that cells move though the lase beam one at a time, along the same path. Instance Numbe - A Specimen can have multiple sets of esults geneated fom acquisition and listmode. Each set of esults saved to the Database is given an incemental instance numbe that intenally links the esults table to the cyto un table and the specimen table. GLOSSARY-3

86 GLOSSARY Integal signal - A voltage pulse with height and aea popotional to the total amount of fluoescent mateial in a cell. IQAP - Abbeviation fo Beckman Coulte's Intelaboatoy Quality Assuance Pogam. A sevice fo all woldwide uses of Immuno-Tol cells and CYTO-TROL contol cells, the IQAP statistically compaes you contol data with that of othe laboatoies. Lase - Abbeviation fo light amplification by stimulated emission of adiation. Two lases ae in the instument: one in the MCL fo eading ba codes and one in the flow cell fo analyzing cells. Levey-Jennings - Gaphs that display the contol data fo easy eview fo tends, shifts, outlies, and violations of you laboatoy s guidelines. The gaph consists of a middle line that shows the mean, and a set of inne limit lines and oute limit lines that appea above and below the mean. The contol data appeas as connected dots within the gaph. Linea amplification - See gain. Listmode data - A list of measuements fom each cell. LISTGATE - Listmode Gate used as a live gate in acquisition and fo listmode achival. Loadlist - A goup of panels. Logaithmic amplification - A method of inceasing the gain and dynamic ange of a signal. A lage gain is applied to a senso's smalle signals than to the senso's lage signals. See also gain. MB - Abbeviation fo megabyte. Mean - Aithmetic aveage of a goup of data. See also standad deviation and coefficient of vaiation. Menu - On a Wokstation sceen, a list of items fom which you can choose. Minimum Event Counte - The count that must minimally be achieved in ode to stop acquisition. Used to ensue collection of ae events. Mouse - A pointing device. The cuso on the Wokstation sceen moves as you slide the mouse on you desk o othe flat suface. Multi-tube Caousel Loade (MCL) - An optional automated sample loade fo the instument. New Pnl/Po - Used when listmode eplay is equied to be pefomed with a Panel o with a diffeent potocol fom the untime potocol. Next File - Listmode button that incements though the queue of listmode files without eplaying the data. Next Set - Button in Multigaph Application that is active in Histogam mode. Incements the display to the next eight histogams in the Hist queue. GLOSSARY-4

87 GLOSSARY Next Test - Button in Multigaph Application that is active in Test mode. Incements the display to the next un numbe in the Hist queue. Neutal density (ND1) filte - An optical filte that can be used with the fowad scatte senso to educe the intensity of the fowad scatte, thus enabling the instument to analyze lage paticles without satuating the senso. Nomalization - Applied to linea statistics fom diffeent histogam esolutions to ensue esults ae compaable. Scales to 1,024. Optical filtes - Mediums, such as glass, that sepaate fluoescent light by wavelength, which is measued in nanometes (nm). See also BK, BP, and DL filtes. Output Options - Acquisition and Listmode global options fo pinting hadcopy and caousel summay. Also contols saving expot files, database ecods, and patient epot files. Panel - A goup of potocols fo analyzing a seies of tubes coesponding to one patient specimen. The Cytomete settings ae passed on though the panel with identification of the pimay tubes. Pass/Load Regions - Button in Multigaph Application that is active in Test mode. Load egions display egions fom the histogam file. Pass egions allow the cuently displayed egions to pass on to the next test. Patient Repot - Repot geneated at the end of a Panel that contains esults manipulated by the Repot Template. Patient Repot Expot File (*.PXP) - The data that is tansmitted though a seial pot o stoed in a file accoding to the Patient Repot template. Patient Repot Template - Template fo a patient panel epot that allows calculations to be pefomed on egion statistics. PCX File - A gaphics file ceated using the mouse to identify the captue aea. Peak signal - A voltage pulse with height popotional to the amount of light the cell scattes o fluoesces. Photo-multiplie tube (PMT) - A light-sensitive senso that convets light enegy into electical cuent and geneates a voltage pulse signal. Pickup lens/spatial filte assembly - Collects side scatte and fluoescent light fom only the sensing aea of the flow cell, and collimates it. Play List - Listmode button that eplays the cuent listmode file in memoy. Play Next - Listmode button that loads and eplays the next listmode file in the queue. Pop-up window - A ectangula aea that appeas on top of the cuent sceen displayed on the Wokstation. You must close the window befoe you can use the cuent sceen again. Positives analysis - Analysis pefomed on the negative contol to set egions automatically to exclude the negative population fom the positives statistics. GLOSSARY-5

88 GLOSSARY Powe Supply - The system component that povides diect cuent powe, pessue, and vacuum to the Cytomete, and collects waste fom the Cytomete. Pecision - Ability of an instument to epoduce simila esults when a sample is un epeatedly. Pecision shows the closeness of test esults when epeated analyses of the same mateial ae pefomed. Also known as epoducibility. Contast with accuacy. PRIME egion - When a histogam Peak is not within a PRIME Region the system pefoms an AutoPime. Pinte - An optional system component that povides a pintout of sample esults and othe infomation. Pism - Phenotype paamete fo multicolo analysis. Pism histogam - Histogam that displays the phenotype of an identified population. Potocol - A set of instuctions that tells the Cytomete what and how to acquie data and elay listmode data. _Q potocols - These potocols ae used to geneate quality contol statistics fom thei QC egions. One o moe can be assigned as seconday o contol potocols at the end of an autostandadization panel. QC Template - A customized design fomat instucting the system to display opeato specified data fom a specific test potocol. QCC file - A QC data file geneated by a _Q potocol that contains the statistics fom all the defined QC egions in that potocol. The data can be viewed in Levey-Jennings and data table fomat. QCS file - A QC Cytomete setting baseline file geneated by a _A potocol that contains the voltage and gain settings fom the _A potocol and the compensation values fom the last _C potocol in the Autostandadization panel. The data can be viewed in Levey-Jennings and data table fomat. Quality contol (QC) - A compehensive set of pocedues a laboatoy sets up to ensue that an instument is woking accuately and pecisely. RCVEXP3 - Pogam that esides in a Wokstation that eceives the seial tansmission of an Expot file fom an XL o XL-MCL flow cytomete. RD1 - Red dye 1. Rebuild - Button used to update a diectoy listing wheneve files ae copied, moved, o deleted. Region Mapping - Regions whose coodinates map to an identified egion. Results DB - That potion of the Database ecod that contains infomation petaining to the esults fom the patient epot. Retic Stat - Algoithm that pefoms automatic gating and analysis used in eticone System. GLOSSARY-6

89 GLOSSARY Runtime Potocol - The Potocol stoed with the Listmode file at acquisition. Listmode eplays identical to the acquisition potocol. Runtime SQL - The database engine accessed fom within SYSTEM II softwae. Scoll ba - The aea on the left of a pop-up window. The ba's aows let you move (scoll) the window's content up o down so that you can see othe pats of it. Fo example, the scoll ba in the Potocol Select window lets you scoll though the entie list of potocol names. Select - To position the mouse cuso on an item, and then pess and elease a mouse button to choose that item. Select Di - A button in the file selection window that allows you to change to a diffeent diectoy o dive to select files. Sensitivity - The ability of the instument to distinguish vey low levels of light scatte and fluoescence fom backgound light o electonic noise. Sheath fluid - A balanced electolyte solution. Side scatte - The amount of lase light scatteed at about a 90 angle to the axis of the lase beam. The amount of side scatte is popotional to the ganulaity of the cell that scatteed the lase light. Side scatte (SS) senso - Collects the side scatte and geneates voltage pulse signals. Specimen DB - That potion of the Database ecod that contains infomation petaining to the specimen and patient demogaphics. Specimen ID - ID assigned to a Specimen daw as opposed to a tube. SQL - Standad quey language. Standad deviation (SD) - A measue of diffeence fom the mean. A measue of pecision. Test Mode - Multigaph analysis of histogams fom the same un numbe eithe tube by tube o batched. Tube ID - The Ba code ID on individual eaction sample tubes. Voltage pulse signals - The signals that the fowad scatte, side scatte, and fluoescence sensos geneate. They ae popotional to the intensity of light the senso eceived. Volume Stop - Stops acquisition based on a pedefined volume of 20, 40, 60, 80, o 100 µl. Westgad ules - A set of ules in the softwae that checks the quality contol data against standad deviation elated citeia. Window - See pop-up window. Wokstation - The system component that uns the softwae that lets you contol the instument. It displays sample esults and othe infomation. GLOSSARY-7

90 GLOSSARY GLOSSARY-8

91 INDEX Symbols _A potocols definition, GLOSSARY-1 desciption, 3-15 _C potocols definition, GLOSSARY-1 desciption, 3-15 _Q potocols definition, GLOSSARY-6 desciption, 3-15 A about this manual, xiii accessibility, instument installation equiements, 2-1 accuacy definition, GLOSSARY-1 action log, A-1 ai conditioning special equiements, 2-1 ambient tempeatue instument, 2-1 amplification desciption, 3-11 application list, fo the instument, 1-1 Atisoft LANtastic softwae option, desciption, 1-5 ASCII definition, GLOSSARY-1 assay values definition, GLOSSARY-1 autostandadization potocols, desciption, 3-15 potocols, examples, 3-16 potocols, pedefined, B-1 AUX paamete desciption, 3-12 signal, definition, GLOSSARY-1 when to use, 3-12 B backgound count definition, GLOSSARY-1 ba-code acceptable, 4-7 Codaba, 4-7 Code 128, 4-7 Code 39 ba code, 4-7 Inteleaved 2-of-5, 4-7 specifications, 4-7 use of checksum, 4-9 ba-code decode desciption, 4-8 ba-code labels optical chaacteistics, 4-7 specifications, 4-7 used on sample caousel, 3-1 ba-code pinte desciption, 1-4 ba-code scanne/eade acceptable ba codes, 4-7 desciption, 1-4 hand-held, specifications, 4-8 MCL, specifications, 4-8 BK filtes definition, GLOSSARY-1 bottle, waste See waste containe, 2-2 BP filtes definition, GLOSSARY-1 Btus equied fo opeating system, 2-1 buttons definition, GLOSSARY-1 C cable connections location, 2-2 powe, 2-2 signal, 2-2 CAL facto definition, GLOSSARY-1 caousel, sample ba-code labels, 3-1 desciption, 3-1 See also MCL, 3-1 cayove pefomance specifications, 4-6 CDRH equied labels, 5-1 cell illumination desciption, 3-4 Cell Seeke definition, GLOSSARY-1 Cell Stat definition, GLOSSARY-1 INDEX-1

92 INDEX Cell Tacke definition, GLOSSARY-1 channel definition, GLOSSARY-1 chaacteistics, pefomance, 4-7 checksum algoithm Codaba, 4-9 Code 39, 4-9 Inteleaved 2-of-5, 4-9 cleaning agent definition, GLOSSARY-1 See eagents, 1-6 Cleanse Mode use of cleaning agent, 1-6 click definition, GLOSSARY-1 Codaba ba code checksum algoithm, 4-9 specifications, 4-7 Code 128 specifications, 4-7 Code 39 ba code checksum algoithm, 4-9 specifications, 4-7 coefficient of vaiation (CV) definition, GLOSSARY-1 collimate definition, GLOSSARY-1 colo compensation definition, GLOSSARY-2 components See system components, 1-2 compute electical input equiements, 2-1 specifications, 4-3 connections, cables location, 2-2 powe, 2-2 signal, 2-2 connections, tubing location, 2-3 pneumatic, 2-3 pessue, 2-3 vacuum, 2-3 waste containe, 2-3 containe, waste capacity, 2-2 connections, 2-3 contols and indicatos, definition, GLOSSARY-2 CYTO-TROL contol cells, 1-6 definition, GLOSSARY-2 Flow-Check fluoosphees, 1-6 Flow-Set fluoosphees, 1-6 fluoosphees, 1-6 Immuno-Tol cells, 1-6 conventions used in the manual, xiv COULTER CLENZ cleaning agent See eagents, 1-6 coss-cylindical lenses beam shaping, 3-3 definition, GLOSSARY-2 CV definition, GLOSSARY-1 Cytomete accessibility, 2-1 desciption, 1-2 sensos, 3-9 space needed, 2-1 specifications, 4-1 cytomete definition, GLOSSARY-2 cytomety See flow cytomety, GLOSSARY-3 CYTO-TROL contol cells See contols, 1-6 D daily log fo instument standadization, A-1 daily log fo instument veification of alignment and fluidics, A-1 data acquisition thoughput pefomance specifications, 4-6 data sheets, mateial safety how to ode, 1-6 data stoage dives, 1-4 options, 1-4 database file, definition, GLOSSARY-2 ecod, definition, GLOSSARY-2 dc definition, GLOSSARY-2 defaults definition, GLOSSARY-2 definitions, GLOSSARY-1 delivey inspection INDEX-2

93 INDEX instument, 2-1 dimensions compute, 4-1 Cytomete, 4-1 instument, 4-1 Powe Supply, 4-1 Wokstation, 4-1 DiOC5(3) definition, GLOSSARY-2 disciminato definition, GLOSSARY-2 Display Options definition, GLOSSARY-2 dissipation, heat special equiements, 2-1 DL filtes definition, GLOSSARY-2 documentation fo you instument about you efeence manual, xiii conventions used, xiv intoduction to you manuals, xiii using the manuals, xiii doublet discimination pefomance specifications, 4-6 dainage equiements, 2-2 E electical input special equiements, 2-1 EPT2ASC2 definition, GLOSSARY-2 equations linea egion statistics, 3-19 log egion statistics, 3-20 establishing fluoescence and/o light scatte taget anges log, A-1 establishing HV/total gain anges log, A-1 establishing peak position and HPCV taget anges log, A-1 event definition, GLOSSARY-2 expot file definition, GLOSSARY-2 extension cod Caution, 2-1 F Fast Comp definition, GLOSSARY-2 Fast Set definition, GLOSSARY-2 FDA definition, GLOSSARY-3 file database, definition, GLOSSARY-2 expot, definition, GLOSSARY-2 PCX, definition, GLOSSARY-5 timming, definition, GLOSSARY-3 filtes BK, definition, GLOSSARY-1 BP, definition, GLOSSARY-1 configuation, fou FL sensos, 3-8 configuation, thee FL sensos, 3-7 DL, definition, GLOSSARY-2 ND1 filte, desciption, 3-6 ND1, definition, GLOSSARY-5 neutal density (ND1) filte, desciption, 3-6 optical, definition, GLOSSARY-5 optical, specifications, 4-2 spatial, function, 3-6 FITC definition, GLOSSARY-3 FL See fluoescent light (FL), 3-6 flow cell definition, GLOSSARY-3 specifications, 4-1 flow cytomety definition, GLOSSARY-3 flow ate specifications, 4-1 Flow-Check fluoosphees log sheets, A-1 See contols, 1-6 Flow-Set fluoosphees log sheets, A-1 See contols, 1-6 fluoescence esolution, pefomance specifications, 4-6 sensitivity, pefomance specifications, 4-6 fluoescent light (FL) cell illumination, 3-4 collection, 3-6 definition, GLOSSARY-3 senso, definition, GLOSSARY-3 INDEX-3

94 INDEX sensos, fou, 3-8 sensos, thee, 3-6 focusing, hydodynamic desciption, 3-1 fomulas linea egion statistics, 3-19 log egion statistics, 3-20 fowad scatte (FS) cell illumination, 3-4 definition, GLOSSARY-3 light collection, 3-4 esolution, pefomance specifications, 4-6 senso, 3-4 senso, definition, GLOSSARY-3 G gain definition, GLOSSARY-3 gating definition, GLOSSARY-3 GB definition, GLOSSARY-3 geneated signals desciption, 3-11 gound path equiement, 2-1 H handheld scanne See ba-code scanne/eade, 1-4 hadwae options, 1-3 hazads adiation exposue, 5-1 heat dissipation, 2-1 high voltage definition, GLOSSARY-3 Hist Mode definition, GLOSSARY-3 histogam definition, GLOSSARY-3 Multigaph, specifications, 4-6 pism, definition, GLOSSARY-6 hot keys definition, GLOSSARY-3 humidity, allowance, 2-1 hydodynamic focusing definition, GLOSSARY-3 desciption, 3-1 I Immuno-Tol cells See contols, 1-6 input, electical special equiements, 2-1 installation categoy, 4-1 equiements, 2-1 softwae vesion 3.0, 2-3 instance numbe definition, GLOSSARY-3 instument accessibility, 2-1 delivey inspection, 2-1 dimensions, 4-1 electical input equiements, 2-1 installation special equiements, 2-1 layout, 2-1 location, 2-1 optical system, 3-5 pefomance specifications, 4-6 softwae specifications, 4-4 space needed, 2-1 unpacking, 2-1 view, 1-3 integal signal definition, GLOSSARY-4 intended use of the instument, 1-1 Inteleaved 2-of-5 ba code checksum algoithm, 4-9 specifications, 4-7 intoduction to you manuals, xiii IQAP definition, GLOSSARY-4 IsoFLow sheath fluid See eagents, 1-5 K keyboad cable connection, 2-2 L labels ba-code, specifications, 4-7 CDRH-equied, 5-1 INDEX-4

95 INDEX lase, waning, 5-1 labels, ba-code specifications, 4-7 used on MCL caousel, 3-1 LANtastic softwae desciption, 1-5 lase ba-code eade, hand held, 4-8 ba-code eade, MCL, 4-8 beam shaping, 3-3 definition, GLOSSARY-4 safety, 5-1 specifications, 4-2 waning labels, 5-1 layout of the instument, 1-3 lenses beam shaping, 3-3 coss-cylindical, 3-3 Levey-Jennings gaphs definition, GLOSSARY-4 desciption, 3-16 light collection, sepaation and measuement desciption, 3-4 linea amplification See gain, GLOSSARY-4 LISTGATE definition, GLOSSARY-4 Listmode specifications, 4-5 Listmode data definition, GLOSSARY-4 loadlist definition, GLOSSARY-4 log sheets action log, A-1 daily log fo instument standadization, A-1 daily log fo instument veification of alignment and fluidics, A-1 establishing fluoescence and/o light scatte taget anges, A-1 establishing HV/total gain anges, A-1 eagent log, A-1 logithmic amplification definition, GLOSSARY-4 M manuals fo you instument about you efeence manual, xiii conventions used, xiv how to use, xiii intoduction to you manuals, xiii using the manuals, xiii mateial safety data sheets (MSDS) how to ode, 1-6 MB definition, GLOSSARY-4 MCL (Multi-Tube Caousel Loade) definition, GLOSSARY-4 desciption, 1-3 loading the sample, 3-1 mean definition, GLOSSARY-4 menu definition, GLOSSARY-4 minimum event counte definition, GLOSSARY-4 monito specifications, 4-4 mouse definition, GLOSSARY-4 MSDS (mateial safety data sheets) how to ode, 1-6 MULTICYCLE softwae option, desciption, 1-5 MultiGaph analysis, specifications, 4-6 Multi-tube Caousel Loade (MCL) See MCL, GLOSSARY-4 N ND1 filte See filtes, 3-6 netwok LANtastic softwae, desciption, 1-5 opeating system softwae option, 1-5 netwok kit desciption, 1-4 netwok seve desciption, 1-4 neutal density (ND1) filte See filtes, GLOSSARY-5 New Pnl/Po definition, GLOSSARY-4 Next File definition, GLOSSARY-4 Next Set definition, GLOSSARY-4 INDEX-5

96 INDEX Next Test definition, GLOSSARY-5 nomalization definition, GLOSSARY-5 O opeation pinciples of, 3-1 optical system, 3-5 optical chaacteistics fo ba-code labels, 4-7 optical filtes See filtes, 3-6 options Atisoft LANtastic softwae, 1-5 ba-code pinte, 1-4 ba-code scanne/eade, 1-4 data stoage, 1-4 fouth FL senso, 1-5 hadwae, 1-3 MULTICYCLE softwae, 1-5 Multi-tube Caousel Loade (MCL), 1-3 netwok kit, 1-4 Pinte, 1-4 RAM upgade kit, 1-5 eticone system, 1-5 softwae, 1-5 tetaone system, 1-5 Wokstation/netwok seve, 1-4 Output Options definition, GLOSSARY-5 P panels definition, GLOSSARY-5 pedefined by Coulte, B-1 paametes AUX, 3-12 desciption, 3-12 PRISM, 3-13 RATIO, 3-13 TIME, 3-12 pass/load egions definition, GLOSSARY-5 patient epot definition, GLOSSARY-5 template, definition, GLOSSARY-5 patient epot expot file definition, GLOSSARY-5 PCX file See file, GLOSSARY-5 peak signal desciption, 3-9 See signal, GLOSSARY-5 pefomance chaacteistics, 4-7 pefomance specifications cayove, 4-6 doublet discimination, 4-6 pecision fo suface makes, 4-6 esolution, 4-6 sensitivity, 4-6 stability, 4-7 thoughput, 4-6 photo-multiplie tube See PMT, GLOSSARY-5 physical specifications, instument, 4-1 pickup lens/spacial filte assembly definition, GLOSSARY-5 function, 3-6 Play List definition, GLOSSARY-5 Play Next definition, GLOSSARY-5 PMT definition, GLOSSARY-5 optional fouth FL senso, desciption, 1-5 pneumatic powe supply desciption, 1-2 location, 2-3 pneumatic tubing connections location, 2-3 pop-up window definition, GLOSSARY-5 positives analysis definition, GLOSSARY-5 powe cable connections, 2-2 electical input, 2-1 powe cables connection, 2-2 Powe Supply accessibility, 2-1 definition, GLOSSARY-6 desciption, 1-2 dimensions, 4-1 INDEX-6

97 INDEX electical input equiements, 2-1 space needed, 2-1 pecautions against adiation, 5-1 pecision (epoducibility) definition, GLOSSARY-6 suface makes, pefomance specifications, 4-6 Pime egion definition, GLOSSARY-6 pinciples of opeation, 3-1 Pinte definition, GLOSSARY-6 pinting methods of ba-code labels, 4-8 pism histogam See histogam, GLOSSARY-6 PRISM paamete desciption, 3-13 pism paamete definition, GLOSSARY-6 potocols autostandadization, 3-15 definition, GLOSSARY-6 pedefined by Coulte, B-1 untime, definition, GLOSSARY-7 Q QC template definition, GLOSSARY-6 pedefined by Coulte, B-1 QCC file definition, GLOSSARY-6 QCS file definition, GLOSSARY-6 Quality Contol (QC) featues, 3-16 quality contol (QC) definition, GLOSSARY-6 mateials, use and function, 1-6 R adiation hazads, 5-1 RAM upgade kit desciption, 1-5 RATIO paamete desciption, 3-13 RCVEXP definition, GLOSSARY-6 RD1 definition, GLOSSARY-6 eagent log, A-1 eagents cleaning agent, 1-6 connections, 2-3 desciption, 1-5 sheath fluid, 1-5 sheath fluid, definition, GLOSSARY-7 types used, 1-5 Rebuild definition, GLOSSARY-6 efeence manual conventions used, xiv intoduction to you manuals, xiv using the manuals, xiv egion mapping See egions, GLOSSARY-6 egions definition, GLOSSARY-6 specifications, 4-5 epoducibility (pecision) definition, GLOSSARY-6 suface makes, pefomance specifications, 4-6 equiements sample, 4-1 equiements, installation instument accessibility, 2-1 space needed, 2-1 esolution pefomance specifications, 4-6 Results DB definition, GLOSSARY-6 Retic Stat definition, GLOSSARY-6 eticone system option, desciption, 1-5 oom tempeatue ambient opeating, 2-1 special equiements, 2-1 specifications, 2-1 untime potocol See potocols, GLOSSARY-7 untime SQL See SQL, GLOSSARY-7 S safety pecautions INDEX-7

98 INDEX lase, 5-1 sample flow, desciption, 3-1 loading, automated, 3-1 loading, manual, 3-1 measuable cell sizes, 4-1 optimum concentation, 4-1 pepaation, 4-1 equiements, 4-1 sample caousel ba-code labels, 3-1 desciption, 3-1 See also MCL, 3-1 sample flow desciption, 3-1 sample stage loading the sample, 3-1 sample tube size equiements, 4-1 scanne, hand-held See ba-code scanne/eade, 4-8 scoll ba definition, GLOSSARY-7 SD (standad deviation) definition, GLOSSARY-7 select definition, GLOSSARY-7 Select Di definition, GLOSSARY-7 sensitivity, 4-6 definition, GLOSSARY-7 pefomance specifications, 4-6 sensos Cytomete, 3-9 fluoescent light, definition, GLOSSARY-3 fowad scatte (FS), definition, GLOSSARY-3 specifications, 4-2 sheath fluid See eagents, GLOSSARY-7 side scatte (SS) cell illumination, 3-4 collection, 3-6 senso, 3-6 senso, definition, GLOSSARY-7 signal cables connections, 2-2 signals geneated, 3-11 peak, 3-9 pocessing of, opeation pinciples, 3-9 pocessing of, specifications, 4-2 voltage pulse, 3-9 softwae options, 1-5 specifications, 4-4 space needed, instument, 2-1 spatial filte See filtes, 3-6 special equiements installation, 2-1 specifications ba-code labels, 4-7 instument, 4-1 Listmode, 4-5 pefomance, 4-6 physical, 4-1 egions, 4-5 softwae, 4-4 specimen See sample, 3-1 specimen DB definition, GLOSSARY-7 specimen ID definition, GLOSSARY-7 SQL definition, GLOSSARY-7 untime, definition, GLOSSARY-7 stability day-to-day, 4-7 pefomance specifications, 4-7 within day, 4-7 standad deviation (SD) definition, GLOSSARY-7 statistics histogam, 3-19 log egion, 3-20 suface makes pecision, 4-6 thoughput, 4-6 system components, 1-2 connections, 2-2 optical, 3-5 system components desciption Cytomete, 1-2 Powe Supply, 1-2 Wokstation, 1-2 INDEX-8

99 INDEX T tempeatue, ambient opeating, 2-1 test mode definition, GLOSSARY-7 test tube size, 4-1 tetaone system option, desciption, 1-5 thoughput pefomance specifications, 4-6 TIME paamete desciption, 3-12 Tube ID definition, GLOSSARY-7 tube, sample See test tube, 4-1 tubing Powe Supply pessue connection, 2-3 Powe Supply vacuum connection, 2-3 waste containe connection, 2-3 Wokstation definition, GLOSSARY-7 seve option, 1-4 specifications, 4-3 wokstation desciption, 1-2 X XL/XL-MCL system components desciption, 1-2 installation, 2-1 intended use, 1-1 manuals, had-copy, xiii pupose, 1-1 specifications, 4-1 view of instument, 1-3 U unpacking of instument, 2-1 use and function, instument, 1-1 V ventilation equiements, 2-1 voltage pulse signals definition, GLOSSARY-7 desciption, 3-9 volume stop definition, GLOSSARY-7 W waste disposal, 2-2 equiements, 2-2 waste containe capacity, 2-2 connections, 2-3 waste line connection, 2-3 Westgad ules definition, GLOSSARY-7 desciption, 3-17 window See pop-up window, GLOSSARY-5 INDEX-9

100 INDEX INDEX-10

101 BECKMAN COULTER, INC. CUSTOMER END USER LICENSE AGREEMENT This Poduct contains softwae that is owned by Beckman Coulte, Inc. o its supplies and is potected by United States and intenational copyight laws and intenational tade povisions. You must teat the softwae contained in this Poduct like any othe copyighted mateial. This license and you ight to use the Poduct teminate automatically if you violate any pat of this ageement. This is a license ageement and not an ageement fo sale. Beckman Coulte heeby licenses this Softwae to you unde the following tems and conditions: You May: 1. Use this softwae in the compute supplied to you by Beckman Coulte; 2. Maintain one copy of this softwae fo backup puposes (the backup copy shall be supplied by Beckman Coulte); 3. Afte witten notification to Beckman Coulte, tansfe the entie Poduct to anothe peson o entity, povided you etain no copies of the Poduct softwae and the tansfeee agees to the tems of this license ageement. You May Not: 1. Use, copy o tansfe copies of this Softwae except as povided in this license ageement; 2. Alte, mege, modify o adapt this Softwae in any way including disassembling o decompiling; 3. Loan, ent, lease, o sublicense this Softwae o any copy. Limited Waanty Beckman Coulte waants that the softwae will substantially confom to the published specifications fo the Poduct in which it is contained, povided that it is used on the compute hadwae and in the opeating system envionment fo which it was designed. Should the media on which you softwae aives pove defective, Beckman Coulte will eplace said media fee of chage within 90 days of delivey of the Poduct. This is you sole emedy fo any beech of waanty fo this softwae. Except as specifically noted above, Beckman Coulte makes no waanty o epesentation, eithe expessed o implied, with espect to this softwae o its documentation including quality, pefomance, mechantability, o fitness fo a paticula pupose. No Liability fo Consequential Damages In no event shall Beckman Coulte o its supplies be liable fo any damages whatsoeve (including, without limitation, damages fo loss of pofits, business inteuption, loss of infomation, o othe pecuniay loss) aising out of the use of o inability to use the Beckman Coulte Poduct softwae. Because some states do not allow the exclusion o limitation of liability fo consequential damages, the above limitation might not apply to you. Geneal This ageement constitutes the entie ageement between you and Beckman Coulte and supesedes any pio ageement concening this Poduct softwae. It shall not be modified except by witten ageement dated subsequent to the date of this ageement signed by an authoized Beckman Coulte epesentative. Beckman Coulte is not bound by any povision of any puchase ode, eceipt, acceptance, confimation, coespondence, o othewise, unless Beckman Coulte specifically agees to the povision in witing. This ageement is govened by the laws of the State of Califonia.

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103 TRADEMARKS ALTRA, the Beckman Coulte logo, Cell-Seeke, Cell-Tacke, COULTER, COULTER CLENZ, CYTO-COMP, CYTO-TROL, Elite, EPICS, FlowCente, Flow-Check, Flow-Set, ImmunoPep, Immuno-Tol, IOTest 3, IsoFlow, Multi-Q-Pep, OptiLyse, Pofile, Q-Pep, eticone, SYSTEM II, tetaone, TQ-Pep, VesaLyse, XL, and XL-MCL ae tademaks of Beckman Coulte Inc. All othe tademaks, sevice maks, poducts, o sevices ae tademaks o egisteed tademaks of thei espective holdes.

104 COULTER EPICS XL and XL-MCL Flow Cytomete Documentation s s s s s s s Refeence PN (Black binding) Special Pocedues and Toubleshooting PN (Blue binding) Opeato's Guide PN (Red binding) Data Management PN (White binding) Getting Stated PN (Clea binding) Opeating Summay PN Maste Index PN Use and Function Installation Opeation Pinciples Specifications/Chaacteistics Pecautions and Hazads Log Sheets Pedefined Panels, Potocols, and QC Templates Glossay Index Cleaning Pocedues Replace/Adjust Pocedues Toubleshooting Ba-Code Options QC Maintenance Sceen Index Statup Quality Contol Ceating a Woklist Running Samples Data Analysis Pinting and Expoting Patient Repots Reviewing QC Data Cleaning and Shutdown Index Data Management System Configuation Woking with QC Featues Ceating Repots Expoting Data File Fomats Index Contols and Indicatos Leaning the Basics Softwae Oveview Ceating Potocols Using Regions Ceating Panels The Opeating System Index Quick efeence vesions of daily pocedues. Combined index fo the Opeato's Guide, Special Pocedues and Toubleshooting, Refeence, Getting Stated, and Data Management manuals. Come visit us at Pinted on Recycled Pape Copyight Beckman Coulte, Inc All Rights Reseved