Beckman Coulter CytoFLEX Series Instructions For Use Manual
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Beckman Coulter CytoFLEX Series Instructions For Use Manual

Flow cytometers
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Instructions for Use
CytoFLEX Series
CytoFLEX, CytoFLEX S, and CytoFLEX LX Flow Cytometers
For Research Use Only. Not for use in diagnostic procedures.
B49006AH
March 2017
Beckman Coulter, Inc.
250 S. Kraemer Blvd.
Brea, CA 92821 U.S.A.

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Summary of Contents for Beckman Coulter CytoFLEX Series

  • Page 1 Instructions for Use CytoFLEX Series CytoFLEX, CytoFLEX S, and CytoFLEX LX Flow Cytometers For Research Use Only. Not for use in diagnostic procedures. B49006AH March 2017 Beckman Coulter, Inc. 250 S. Kraemer Blvd. Brea, CA 92821 U.S.A.
  • Page 2 PN B49006AH (March2017) © 2017 Beckman Coulter, Inc. All rights reserved. Beckman Coulter, the stylized logo, and the Beckman Coulter product and service marks mentioned herein are trademarks or registered trademarks of Beckman Coulter, Inc. in the United States and other countries.

  • Page 3: Revision History

    This document applies to the latest software listed and higher versions. When a subsequent software version affects the information in this document, a new issue will be released to the Beckman Coulter Web site. For labeling updates, go to www.beckmancoulter.com and download the latest version of the manual or system help for your instrument.
  • Page 4 This document applies to the latest software listed and higher versions. When a subsequent software version affects the information in this document, a new issue will be released to the Beckman Coulter Web site. For labeling updates, go to www.beckmancoulter.com and download the latest version of the manual or system help for your instrument.
  • Page 5 This document applies to the latest software listed and higher versions. When a subsequent software version affects the information in this document, a new issue will be released to the Beckman Coulter Web site. For labeling updates, go to www.beckmancoulter.com and download the latest version of the manual or system help for your instrument.
  • Page 6 This document applies to the latest software listed and higher versions. When a subsequent software version affects the information in this document, a new issue will be released to the Beckman Coulter Web site. For labeling updates, go to www.beckmancoulter.com and download the latest version of the manual or system help for your instrument.
  • Page 7 This document applies to the latest software listed and higher versions. When a subsequent software version affects the information in this document, a new issue will be released to the Beckman Coulter Web site. For labeling updates, go to www.beckmancoulter.com and download the latest version of the manual or system help for your instrument.
  • Page 8 Revision History viii B49006AH...

  • Page 9: Safety Notices

    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.

  • Page 10: Safety Precautions

    • This equipment is used in a manner other than specified. Operate the instrument as instructed in the product manuals. • You introduce software that is not authorized by Beckman Coulter into your computer. Only operate your system’s software with software authorized by Beckman Coulter.

  • Page 11: Symbol Explanations

    Safety Notices Symbol Explanations WARNING California Proposition 65: This product may contain chemicals known to the State of California to cause cancer and birth defects or other reproductive harm. Symbol Explanations Symbol meanings Caution / Warning Electrical shock hazard. Operate with caution! Possible laser radiation inside.
  • Page 12 Safety Notices Symbol Explanations B49006AH...

  • Page 13: Table Of Contents

    Contents Revision History, iii Safety Notices, ix Alerts for Danger, Warning, and Caution, ix Safety Precautions, x Symbol Explanations, xi Introduction, xxvii Overview, xxvii How to Use Your Manual, xxvii About this Manual, xxvii Conventions Used, xxix Graphics, xxix CHAPTER 1: System Overview, 1-1 Overview, 1-1...
  • Page 14 Contents Instrument Specifications, 1-25 Dimensions [CytoFLEX], 1-25 Dimensions [CytoFLEX LX], 1-25 Installation Category, 1-25 Pollution Degree, 1-25 Acoustic Noise Level, 1-25 Electrical Ratings, 1-25 Cytometer, 1-26 Performance Characteristics, 1-28 Performance Characteristics [CytoFLEX], 1-28 Performance Characteristics [CytoFLEX LX], 1-29 Reagent Limitations, 1-29 CHAPTER 2: Using the CytExpert Software, 2-1...
  • Page 15 Contents Account Policies, 2-30 User Management Operation Log, 2-31 Viewing and Exporting User Logs, 2-31 Graphic and Gating Styles, 2-32 Plots, 2-32 Gates, 2-33 Software Settings, 2-34 Language Settings, 2-37 Setting Up CytExpert Application Programming Interface (API) Test Client, 2-37 CHAPTER 3: Daily Startup, 3-1...
  • Page 16 Contents Collecting QC Data, 4-10 Collecting QC Data [With Plate Loader], 4-12 Confirming Results, 4-16 Creating Levey-Jennings Charts, 4-19 QC Result Manager, 4-21 Standardization, 4-22 Adding, Editing, Importing, and Deleting Standardization Laser Target Values in the QC Experiment, 4-23 Adding a New Standardization Item, 4-24 Editing Standardization Item Parameters, 4-25...
  • Page 17 Contents Setting Collection Conditions, 5-58 Setting Plot Display Conditions, 5-60 Setting Customized Parameters, 5-61 Setting Custom Statistics, 5-62 Analyzing and Exporting Data, 5-64 Exporting FCS Files, 5-70 Exporting Single Tube Files, 5-70 Exporting Multiple FCS Files, 5-72 Exporting Plots or the Statistics Table of Multiple Tubes as Picture Files, 5-73 Importing and Exporting Instrument Settings, 5-73...
  • Page 18 Contents Copying a Previously Acquired Experiment, 7-1 Importing Previously Acquired Data, 7-1 Setting the Plots and Statistics, 7-3 Opening the Analysis Screen, 7-3 Creating Histogram and Dot Plot Overlays, 7-5 Calculating Sample Volume and Concentration, 7-7 Adjusting Compensation Settings, 7-8 Exporting Results, 7-9 CHAPTER 8:...
  • Page 19 Contents Cleaning the Sample Station, 10-7 Cleaning the Sample Probe, 10-8 Deep Clean Procedure, 10-8 Cleaning the 4 L Sheath Fluid Container, 10-10 Cleaning the 4 L Waste Container, 10-11 Nonscheduled Cleaning, 10-13 Surface Cleaning and Disinfection, 10-13 Preparing the Instrument for Transport or Storage, 10-14 Lifting and Carrying Instructions, 10-18...
  • Page 20 Contents Changing Sample Mixing and Backflush Settings, 11-71 Calibrating the Plate Position [With Plate Loader], 11-73 APPENDIX A: Instrument Installation, A-1 Overview, A-1 Instrument Transportation and Storage, A-1 Installation Environment Validation, A-2 Worktable, A-2 Ventilation and Cleaning, A-2 Power Source, A-3 Temperature and Humidity, A-4 Waste...
  • Page 21 Switching the Sample Probe from the Plate Loader to the Single Tube Sample Station [With Sample Injection Mode Control Knob] , C-4 APPENDIX D: Table of Hazardous Substances, D-1 Table of Hazardous Substances, D-1 Abbreviations Index Beckman Coulter, Inc. Customer End User License Agreement Related Documents...
  • Page 22 Illustrations Illustrations Main Components [CytoFLEX Without Plate Loader], 1-2 Main Components [CytoFLEX LX], 1-3 Optical Filter Mounts, 1-5 Optical Filter Mount with Optical Filter, 1-5 Optical Filter Mount Labeled with the Band-Pass Information, 1-6 Optical Filter Mount (Top), 1-6 Fluid Containers [CytoFLEX], 1-10 Fluid Cubitainers [CytoFLEX LX], 1-11...
  • Page 23 Illustrations All Gates - Example Experiment, 5-46 Circular Gating Logic - Example Experiment, 5-46 Movement - Default Setting, 5-51 Movement - Max Setting, 5-52 Extent - Default Setting, 5-52 Extent - Maximum Setting, 5-53 Laser Setting Window [CytoFLEX LX shown], 5-53 Before Compensation, 6-1 After...
  • Page 24 Illustrations Experiment Operation Log Window, B-12 Select Experiment Profile Window, B-13 Print and Export Preview Window, B-14 System Operation Log Window, B-15 User Management Operation Log Window, B-16 Sample Injection Mode Control Knob, C-2 xxiv...
  • Page 25 Tables Tables WDM Optical Filter Mount Color Codes [CytoFLEX], 1-7 WDM Optical Filter Mount Color Codes [CytoFLEX LX], 1-7 Troubleshooting, 9-10 Troubleshooting [With Plate Loader], 9-22 Table of Hazardous Substances Name and Concentration [CytoFLEX], D-2 Table of Hazardous Substances Name and Concentration [CytoFLEX LX], D-3...
  • Page 26 Tables xxvi...

  • Page 27: Introduction

    Introduction Overview This introduction contains the following information: • How to Use Your Manual • About this Manual • Conventions Used • Graphics How to Use Your Manual Your CytoFLEX flow cytometer system includes the manuals listed below: • Use this Instructions for Use manual for information on the day-to-day operation of your CytoFLEX flow cytometer.
  • Page 28 Introduction About this Manual CHAPTER 3, Daily Startup Provides instructions for starting your CytoFLEX flow cytometer and navigating to the sample testing standby state. CHAPTER 4, Instrument Quality Control and Standardization Provides instructions for performing daily quality control (QC) on your CytoFLEX flow cytometer to confirm the instrument is working correctly and to ensure accurate experimental data measurement.

  • Page 29: Conventions Used

    Introduction Conventions Used Conventions Used This document uses the following conventions: • Bold face font indicates buttons or selections that appear on the workstation screen. • The term “select” is used to indicate the following action: — To click with a mouse. NOTE The verb “press”...
  • Page 30 Introduction Graphics B49006AH...

  • Page 31: System Overview

    CHAPTER 1 System Overview Overview This chapter describes the individual components of the CytoFLEX flow cytometer and the corresponding functions of these components. This chapter contains information on: • Product Description • Main Components • Optical Components • Fluidics System •...

  • Page 32: Main Components

    System Overview Main Components Main Components CAUTION Risk of instrument damage and/or instrument stability. Do not place any objects on top of the instrument, as this could cause warping of the top cover or affect the stability of the optical path. The instrument consists of three main components: Fluid Containers/Cubitainers, Cytometer, and the Workstation.

  • Page 33: Main Components [Cytoflex Lx]

    Cyto FLEX Sheath Fluid A nonionic, non-flourescent, and azide free sheath fluid for use on Beckman Coulter CytoFLEX flow cytometers. REACTIVE INGREDIENTS: Anti-microbial compound 1. Fluid Cubitainers. Accommodates sheath fluid and waste liquids as required for operation of the instrument.

  • Page 34: Optical Components

    System Overview Optical Components Optical Components CAUTION Risk of operator injury. When operating the instrument, keep the top cover in the closed position to prevent the top cover from falling. When opening the top cover, be cautious to avoid any possible pinch points. The optical components are located in the upper portion of the Cytometer and are visible when the top cover is open.

  • Page 35: Wavelength Division Multiplexer (Wdm)

    System Overview Optical Components Wavelength Division Multiplexer (WDM) Each WDM corresponds to a different laser, or in some cases two lasers. The color of the ring on each cap corresponds to the color of the respective laser. Pressing the two release buttons on opposite edges of the cap allows you to open the unit and replace the filters inside.

  • Page 36: Optical Filter Mount Labeled With The Band-Pass

    System Overview Optical Components Each optical filter mount is labeled with the corresponding laser and band-pass information. See Figure 1.5. Figure 1.5 Optical Filter Mount Labeled with the Band-Pass Information The top of each optical filter mount has two marks. The color of the dot indicates the color of the laser.

  • Page 37: Wdm Optical Filter Mount Color Codes [Cytoflex]

    System Overview Optical Components Table 1.1 WDM Optical Filter Mount Color Codes [CytoFLEX] CytoFLEX Channel Commonly used Laser Fluorescent Channel Names Fluorescent Dyes FITC FITC, Alexa Fluor™ 488, 488 nm 525/40 BP CFSE, Fluo-3 PE, PI 585/42 BP ECD, PE-Texas Red®, 610/20 BP PE-CF594, PI PC5.5...
  • Page 38 System Overview Optical Components Table 1.2 WDM Optical Filter Mount Color Codes [CytoFLEX LX] (Continued) CytoFLEX Channel Commonly used Laser Fluorescent Channel Names Fluorescent Dyes V450-PB450 Pacific Blue™dye, V450, 405 nm 450/45 BP eFluor™ 450, BV421 V525-KrO Krome Orange, AmCyan, 525/40 BP V500, BV510 V610...

  • Page 39: Optical Fiber

    System Overview Optical Components Optical Fiber CAUTION Risk of data integrity damage. • During use, verify that the optical fibers are securely connected to the WDM. A loose connection can alter the optical path and affect fluorescence detection. • Do not disconnect the fiber as this could contaminate the tip and weaken the signal.

  • Page 40: Fluidics System

    NOTE The 10 L sheath fluid and waste cubitainers are available from Beckman Coulter as an alternative to the 4 L Fluid Containers (refer to Figure 1.7) provided with your CytoFLEX and CytoFLEX S Flow...

  • Page 41: Fluid Containers/Cubitainers

    2. Sheath fluid cubitainer. 10 L capacity, for holding sheath fluid. Beckman Coulter recommends using CytoFLEX Sheath Fluid or a similar nonionic sheath fluid to ensure system performance.

  • Page 42: Fluidics Module

    System Overview Fluidics System Fluidics Module The Fluidics module is on the right side of the Cytometer. To access it, you must first open the right-side cover. Refer to Right-Side Cover Removal and Reinstallation CHAPTER 11, Replacement/Adjustment Procedures. Inside the module, in addition to working pumps, valves, and tubing, there is a sheath filter and a Deep Clean solution bottle.

  • Page 43: Fluidic Connections

    System Overview Fluidics System Figure 1.10 Fluidic Connections 1. Waste level sensor connector.Connects to the waste liquid sensor cable. 2. Flow cell waste out. Connects to the waste tubing of the wash station. 3. Sheath fluid level sensor connector. Connects to the sheath fluid sensor cable. 4.

  • Page 44: Sample Station

    System Overview Sample Station Sample Station WARNING Risk of biohazardous contamination and/or instrument damage. When running samples, it is important to insert the sample tube all the way down into the sample tube holder, until the bottom of the sample tube touches the base of the holder. Failing to do this could cause the sample probe to bend or break on entry.

  • Page 45: Sample Tube Holder Positions

    System Overview Sample Station Sample Tube Holder Positions Three of the sample tube holder positions are shown in Figure 1.12: sample loading position standby position , and sample acquisition position . You can only distinguish the mixing position from the sample acquisition position if you are looking directly at the sample tube holder while the Cytometer is processing a sample.

  • Page 46: Plate Loader Components

    System Overview Plate Loader Components Plate Loader Components Figure 1.13 Plate Loader [CytoFLEX Shown] 1. Plate loader door 2. Plate holder stage 3. Plate holder (removable) 1-16 B49006AH...
  • Page 47 System Overview Plate Loader Components Figure 1.14 Plate Loader (Front Cover Removed) 1. Plate loader PEEK tubing 2. Plate loader sample probe assembly 3. Plate loader wash station 4. Plate holder 1-17 B49006AH...

  • Page 48: Plate Holder Components

    System Overview Plate Loader Components Plate Holder Components Figure 1.15 Plate Holder 1. Spring leaves to hold plate 2. Plate holder notches NOTE The Plate holder is removable and replaceable. Refer to Replacing the Plate Holder [With Plate Loader] CHAPTER 11, Replacement/Adjustment Procedures.

  • Page 49: Plate Holder Stage

    System Overview Plate Loader Components Figure 1.16 Plate Holder Stage 1. Pins 2. Position A1 1-19 B49006AH...

  • Page 50: System Configuration

    System Overview System Configuration System Configuration CAUTION Risk of data loss and/or instrument damage. Never shut off the power or disconnect a data cable while the Cytometer is in the process of performing a task. This could cause data loss or damage to the system. System Configuration [CytoFLEX] Figure 1.17 System Connections CLASS 1 LASER PRODUCT...

  • Page 51: Back Cover Connections

    System Overview System Configuration Figure 1.18 Back Cover Connections CLASS 1 LASER PRODUCT COMPLIES WITH 21 CFR 1040.10 AND 1040.11 EXCEPT FOR DEVIATIONS PURSUANT TO LASER NOTICE NO. 50 DATED JUNE 24, 2007 MANUFACTURED 1. Power switch. Turns Cytometer on and off. An indicator light glows when the power is on. 2.

  • Page 52: System Configuration [Cytoflex Lx]

    LASER NOTICE NO. 50 DATED JUNE 24, 2007 MANUFACTURED 10 L Cyto FLEX Sheath Fluid A nonionic, non-flourescent, and azide free sheath fluid for use on Beckman Coulter CytoFLEX flow cytometers. REACTIVE INGREDIENTS: Anti-microbial compound 1. Monitor 4. Computer 2. Keyboard 5. Cytometer 3.

  • Page 53: Back Cover Connections

    System Overview System Configuration Figure 1.21 Back Cover Connections CLASS 1 LASER PRODUCT COMPLIES WITH 21 CFR 1040.10 AND 1040.11 EXCEPT FOR DEVIATIONS PURSUANT TO LASER NOTICE NO. 50 DATED JUNE 24, 2007 MANUFACTURED 1. Power switch. Turns Cytometer on and off. An indicator light glows when the power is on. 2.

  • Page 54: Consumables And Supplies

    CytoFLEX flow cytometer’s Infra Red optical alignment. Material Safety Data Sheets (SDS/MSDS) To obtain an SDS or MSDS for CytoFLEX Series reagents used on the CytoFLEX Series systems: • On the Internet, go to www.beckmancoulter.com: 1.

  • Page 55: Instrument Specifications

    System Overview Instrument Specifications Instrument Specifications Dimensions [CytoFLEX] Dimensions Instrument Cytometer [With or Without 42.5 cm x 42.5 cm x 34 cm dimensions (Length x Plate Loader] Width x Height) Fluid Containers and Fluid 14 cm x 35.6 cm x 35.6 cm Container holder Weight Cytometer [Without Plate...

  • Page 56: Cytometer

    System Overview Instrument Specifications Cytometer Optics Excitation Optics The CytoFLEX system can be configured with up to three spatially-separated lasers. The optical system is alignment free. The laser delays are automatically adjusted by the daily QC system, if required. No user intervention is required to ensure optimum system performance.
  • Page 57 System Overview Instrument Specifications Fluidics System Sample loading speed Defaults Slow 10 μL/min Medium 30 μL/min Fast 60 μL/min Fluid capacity CytoFLEX: Standard 4-L sheath fluid and waste containers; Optional 10 L sheath fluid and waste cubitainers CytoFLEX LX: Standard 10-L sheath fluid and waste cubitainers Automated Startup (initialize), system startup program, sample mix, backflush, prime, maintenance cycles...

  • Page 58: Performance Characteristics

    System Overview Performance Characteristics Data Management Minimum Operating system Windows® 7, 8, 10 Professional 64-bit Workstation/ Processor 6th Generation Intel Core i7 (8MB cache 4.0 ghz) computer Memory 8 GB RAM requirements [CytoFLEX LX] Storage 256 GB Port 1 GB Ethernet port 5 USB 2.0 and above ports Compensation Full matrix compensation, manual and automatic.

  • Page 59: Performance Characteristics [Cytoflex Lx]

    System Overview Reagent Limitations Performance Characteristics [CytoFLEX LX] Performance Sensitivity MESF FITC: <30 molecules of equivalent soluble fluorochrome (MESF-FITC) PE: <10 molecules of equivalent soluble fluorochrome (MESF-PE) Fluorescence rCV <5% resolution The CytoFLEX LX Flow Cytometer is capable of achieving <5% rCV only with calibration reagents that have the capability to achieve this specification.
  • Page 60 System Overview Reagent Limitations 1-30 B49006AH...

  • Page 61: Chapter 2: Using The Cytexpert Software

    CHAPTER 2 Using the CytExpert Software Overview The CytExpert software is a full-feature software package that controls the instrument's operation, collection of experiment data, and analysis of the results. This chapter will explain the software’s functions and features. This chapter contains information on: •...

  • Page 62: Start Page

    Using the CytExpert Software Main Software Screen Start Page The start page automatically opens after the software has been launched. The following operations can be selected from the start page: • New Experiment . For creating a new experiment. The process creates a file with the .xit extension and a folder with the same file name where the raw data (.fcs files) are kept.

  • Page 63: Acquisition Screen

    Using the CytExpert Software Main Software Screen The Experiment, Template, and Compensation tabs below give you the option of opening one of the 10 most recently opened experiments. Acquisition Screen New Experiment New Experiment From Template Open Experiment Selecting , or automatically opens the Acquisition screen.

  • Page 64: Acquisition Screen Navigation

    Using the CytExpert Software Main Software Screen Acquisition Screen Navigation The Acquisition screens have two navigation icons, one for the Acquisition screen and the other for the Analysis screen. 1. Acquisition screen icon. Accesses the Acquisition screen. 2. Analysis screen icon. Accesses the Analysis screen. B49006AH...

  • Page 65: Collection [With Plate Loader]

    Using the CytExpert Software Main Software Screen Collection Standby state Initialized state 1. Acquisition control. Controls sample loading/unloading and data acquisition and recording. 2. Acquisition status. Displays such information as the acquisition rate (Events/Sec), event count, duration, and abort (%). 3.
  • Page 66 Using the CytExpert Software Main Software Screen Collection [With Plate Loader] Standby state Initialized state 1. Acquisition control. Controls sample loading/unloading and data acquisition and recording. 2. Acquisition status. Displays such information as the acquisition rate (Events/Sec), event count, duration, and abort (%). 3.

  • Page 67: Test Tubes

    Using the CytExpert Software Main Software Screen Test Tubes Manual/Semi Automatic Sample Injection Plate Loader Sample Injection Mode Test Tube Status Mode 1. Tube management controls. Manages sample tubes. Used to add, copy, or delete attributes, open the tube property, and open the compensation matrix. 2.

  • Page 68: Plot Area

    Using the CytExpert Software Main Software Screen Plot area 1. Plot controls. For creating single or multiple plots, such as dot plots, histograms, density plots, pseudo color plots, and contour plots. 2. Statistics and hierarchy controls. For creating statistical and hierarchical charts. 3.

  • Page 69: Status Bar

    Using the CytExpert Software Main Software Screen Status Bar 1. Communication connection status. Displays whether the Cytometer and the Workstation are connected. 2. Instrument status information. Displays the status of the Cytometer. 3. Laser status. Displays the status of each laser. NOTE The laser status only displays when a required laser is disabled.

  • Page 70: Compensation Experiment Screen

    Using the CytExpert Software Main Software Screen The Tube management module cannot add new sample tubes. Return to the Acquisition screen to add new sample tubes. Semi Automatic Sample Injection Mode Shown Drawing controls (see Figure 2.1) include the multi-data histograms and graphical display data controls.
  • Page 71 Using the CytExpert Software Main Software Screen Semi-Automatic Sample Injection Mode Shown 1. Tube management. Displays sample tubes required for the compensation experiment. 2. Plot area. Displays compensation plots and gating. The Tube management section of the screen can import saved data (.fcs) files for computational purposes.

  • Page 72: Qc Experiment Screen

    Using the CytExpert Software Main Software Screen QC Experiment Screen The Quality Control (QC) Experiment screen appears when you access a QC experiment. QC Report Screen Before starting the QC routine, a Settings screen appears. Figure 2.2 QC Report Screen [CytoFLEX LX Semi-Automatic Sample Injection Mode Shown] 1.

  • Page 73: Qc Experiment Screen

    Using the CytExpert Software Main Software Screen QC Experiment Screen When acquiring QC samples, the software opens the QC screen. 1. QC experiment progress indicator. Displays the QC stage. 2. Plot area. Displays the QC plots. QC Screen Navigation The Analysis screens have two navigation icons, one for the QC screen and the other for the Levey-Jennings (LJ) charts.

  • Page 74: Software Menu

    Using the CytExpert Software Main Software Screen Software Menu IMPORTANT All menu items apply to the CytExpert Default software option unless otherwise specified. The CytExpert software contains the following selectable menu items: Figure 2.3 Software Menu Tree* Cytometer Settings Advanced Account* * * Log* * * Signature††...

  • Page 75: Acquisition And Analysis Screen Menu

    Using the CytExpert Software Main Software Screen Figure 2.4 QC Software Menu Tree Cytometer Settings Advanced Account‡ ‡ Log‡ ‡ Signature** Help File QC Setting View Help Delay User Experiment Detector Experiment Target Library File Setting Manager‡ ‡ Operation Configuration Standardization About Laser...
  • Page 76 Using the CytExpert Software Main Software Screen File Menu For creating new experiments, opening existing experiments, saving new experiments and data, and importing/exporting FCS data files. CytExpert Default Software Option CytExpert User Management Software Option 2-16 B49006AH...
  • Page 77 Using the CytExpert Software Main Software Screen CytExpert Electronic Record Management Software Option Cytometer Menu For configuring Cytometer settings and controlling Cytometer functions. Depending on the Cytometer state, certain functions may not be available. CytExpert Default Software Option - CytExpert Default Software Option - Standby state Initialized state 2-17...
  • Page 78 Using the CytExpert Software Main Software Screen CytExpert User Management Software Option - CytExpert User Management Software Option - Standby state Initialized state 2-18 B49006AH...
  • Page 79 Using the CytExpert Software Main Software Screen CytExpert Electronic Record Management CytExpert Electronic Record Management Software Option - Standby state Software Option - Initialized state NOTE The Turn On and Turn Off selections are only available on the CytoFLEX LX. Settings Menu Used to select and/or change software options and settings.
  • Page 80 Using the CytExpert Software Main Software Screen CytExpert User Management Software Option CytExpert Electronic Record Management Software Option QC/Standardization Menu Select Start QC/Standardization from the QC/Standardization menu to start the QC routine. NOTE The QC/Standardization menu is the same for the CytExpert Default, CytExpert User Management, and the CytExpert Electronic Record Management software options.
  • Page 81 Using the CytExpert Software Main Software Screen CytExpert User Management Option - Semi- CytExpert User Management Software Option - Automatic/Manual Sample Injection Mode Plate Loader Sample Injection Mode CytExpert Electronic Record Management CytExpert Electronic Record Management Software Option - Semi-Automatic/Manual Software Option - Plate Loader Sample Injection Sample Injection Mode Mode...

  • Page 82: User Management

    Using the CytExpert Software User Management CytExpert Electronic Record Management Software Option NOTE The Log menu is not available in the CytExpert Default software option. Signature Menu Used to sign experiment and view signature details. CytExpert Electronic Record Management Software Option NOTE The Signature menu is only available in the CytExpert Electronic Record Management software option.

  • Page 83: User Manager (Card View)

    Using the CytExpert Software User Management Figure 2.5 User Manager (Card View) Figure 2.6 User Manager (Grid View) 1. Search text box: Filters users by username and 6. Unlock: Used to unlock an existing account that display name. has been locked. 2.

  • Page 84: Creating, Deleting, And Modifying Users In User Manager

    Using the CytExpert Software User Management Creating, Deleting, and Modifying Users in User Manager Creating a New User in User Manager Select in the User Manager window. The New window appears. Fill in the new user information. a. Enter the Username. b.

  • Page 85: Deleting Users In User Manager

    Using the CytExpert Software User Management Deleting Users in User Manager IMPORTANT If an account has been used and log information has been generated related to it, the account cannot be deleted, but it can be disabled. Select the user to be deleted in the User Manager window then select NOTE The user 'Admin' is a system default user and cannot be deleted.

  • Page 86: Unlocking A User Account

    Using the CytExpert Software Role Management Unlocking a User Account Select a Locked user in the User Manager window and select Unlock NOTE You cannot unlock an active user. Resetting a User Passwords Select a user in the User Manager window then select .

  • Page 87: Role Manager

    Using the CytExpert Software Role Management Account Role Manager Select . The Role Manager window appears. Refer to Figure 2.7. > Figure 2.7 Role Manager 1. New: Used to create a new role profile. 2. Modify: Used to modify an existing role profile. 3.

  • Page 88: Creating, Deleting, And Modifying User Roles In Role Manager

    Using the CytExpert Software Role Management Creating, Deleting, and Modifying User Roles in Role Manager Creating New User Roles in Role Manager Select . The New window appears. Fill in the new role information. a. Enter the role name. b. Enter the role description. c.

  • Page 89: Deleting User Roles In Role Manager

    Using the CytExpert Software Role Management Deleting User Roles in Role Manager IMPORTANT If a role has already been assigned to a user, that role cannot be deleted. IMPORTANT The Administrator and Operator Roles are system defaults and may not be deleted. Select the Role to be deleted in Role Manager then select Select Modifying User Roles in the Role Window...

  • Page 90: Account Policies

    Using the CytExpert Software Account Policies Select Account Policies IMPORTANT Only an Administrator can manage users. You must have either the CytExpert User Management or CytExpert Electronic Record Management software option installed to use this feature. Refer to CytExpert Software Installation Options APPENDIX A, Instrument Installation.

  • Page 91: User Management Operation Log

    Using the CytExpert Software User Management Operation Log User Management Operation Log IMPORTANT You must have either the CytExpert User Management or CytExpert Electronic Record Management software option installed to use this feature. Refer to CytExpert Software Installation Options APPENDIX A, Instrument Installation.

  • Page 92: Graphic And Gating Styles

    Using the CytExpert Software Graphic and Gating Styles Graphic and Gating Styles Plots The CytExpert software offers a variety of plot formats including: • Single-parameter plots and histogram overlays • Dual-parameter plots: dot plots, density plots, pseudo color plots, contour plots, and dot plot overlays NOTE Histogram Overlays and Dot Plot Overlays can only be created from multiple samples in the...

  • Page 93: Gates

    Using the CytExpert Software Graphic and Gating Styles Pseudo color plot Contour plot Dot plot overlay Gates Various gating choices are available. The software includes the following gate types: • For dual-parameter plots: lasso, polygon, rectangle, four-quadrant, hinged gates, and auto polygon •...

  • Page 94: Software Settings

    Using the CytExpert Software Software Settings Vertical gate a. This gate can be created using the autogate functionality. Refer to Creating and Adjusting Auto Gates CHAPTER 5, Data Acquisition and Sample Analysis Software Settings Options Select in the Settings menu to configure the software settings. In the experiment settings, you can set the experiment’s default save path.
  • Page 95 Using the CytExpert Software Software Settings In the tube settings, you can select the columns that display in the tube section of the screen. In the plot settings, you can define the background of the graphics display area, configure the histograms, and set the default signal parameters to either the channel’s area or the channel’s height.
  • Page 96 Using the CytExpert Software Software Settings In the Gate settings, you can choose to display population percentage on all plots except overlay. In the Page Setup settings, you can change the page size, orientation, margin size, and display options. 2-36 B49006AH...

  • Page 97: Language Settings

    The two options currently offered are English and Simplified Chinese. Setting Up CytExpert Application Programming Interface (API) Test Client The CytExpert API is available for external software to control CytoFLEX series instruments. It allows external software to perform operations such as running methods and allows for basic control of the plate loader.
  • Page 98 Using the CytExpert Software Software Settings 2-38 B49006AH...

  • Page 99: Daily Startup

    CHAPTER 3 Daily Startup Overview IMPORTANT Verify that the correct USB configuration key is securely connected to a computer USB port. If the USB configuration key is not connected, the following error message appears: CytExpert cannot find the license. Please check whether the correct USB configuration key has been plugged in. This chapter describes the instrument startup procedure.

  • Page 100: Check Waste And Reagent Levels [4 L Fluid Containers]

    Risk of instrument damage. Do not use a saline-based sheath fluid on the CytoFLEX instrument. Saline-based sheath fluid could damage instrument components. Beckman Coulter recommends using CytoFLEX Sheath Fluid or a similar nonionic sheath fluid to ensure system performance. Examine the sheath fluid and waste containers. Verify that there is sufficient sheath fluid in the sheath fluid container and that the waste container is empty.
  • Page 101 Daily Startup Pre-Startup Inspection WARNING Risk of chemical injury from bleach. To avoid contact with the bleach, use barrier protection, including protective eyeware, gloves, and suitable laboratory attire. Refer to the Safety Data Sheet for details about chemical exposure before using the chemical.

  • Page 102: Check Waste And Reagent Levels [10 L Fluid Cubitainers]

    CAUTION Risk of instrument damage. Do not use a saline-based sheath fluid on the CytoFLEX Series instrument. Saline-based sheath fluid could damage instrument components. Beckman Coulter recommends using CytoFLEX Sheath Fluid or a similar nonionic sheath fluid to ensure system performance.

  • Page 103: Power Source Inspection

    Daily Startup Pre-Startup Inspection Verify that all sheath fluid tubing, waste tubing, and sensor cables are properly connected, as shown in the figure: 1. Waste level sensor connector. Connects to the waste liquid sensor cable. 2. Flow cell waste out. Connects to the flow cell waste tubing. 3.

  • Page 104: Turning On The Instrument

    Daily Startup Turning On the Instrument Turning On the Instrument CAUTION 1. If the Cytometer or Workstation fails to start properly, check first to see whether the power cable and connection cables are properly connected. 2. Never shut off the power or disconnect a data cable while the Cytometer is performing a task.
  • Page 105 Daily Startup Logging Into the Software Enter your username and password. Select NOTE The display name of the user that is currently logged in displays in the top, right corner of the software screen. Confirm that the software and the Cytometer are properly connected. a.
  • Page 106 • Select the status information in the lower left to open the system log. Send a copy of the system log to your Beckman Coulter Representative for support if a service call is requested. B49006AH...

  • Page 107: Logging Out Of The Software

    Daily Startup Logging Into the Software Logging Out of the Software If you have the CytExpert User Management or the CytExpert Electronic Record Management software option installed, select the username displayed in the top-right corner of the software screen and select File Exit If you have the CytExpert Default software option installed, log out is not required.

  • Page 108: Using Semi-Automatic Injection Mode

    Daily Startup Logging Into the Software Using Semi-Automatic Injection Mode Sample Injection Mode > Semi Automatic Select in the Cytometer menu to change the Sample Injection mode selection. The sampler status icon located in the bottom right side of the screen changes to display Semi-automatic Sampler.

  • Page 109: Using The Manual Injection Mode

    Daily Startup Logging Into the Software WARNING Risk of biohazardous contamination. When using 1.5-mL and/or 2-mL sample tubes, always cut the cap off and do not exceed 300-μL sample volume. Running samples with a cap attached to the sample tube or with volumes exceeding 300 μL can result in sample splashing.

  • Page 110: Selecting The Plate Loader Sample Injection Mode [With Plate Loader]

    Daily Startup Logging Into the Software Backflush Select to clean the probe. Selecting the Plate Loader Sample Injection Mode [With Plate Loader] Sample injection Mode Select in the Cytometer Menu to change between the Semi-Automatic Injection mode, the Manual Injection mode, and the Plate Loader Injection mode. The Plate Loader Injection mode can be used for running small volumes using the following plates: 96-well flat-bottom, 96-well V-bottom, and 96-well U-bottom.
  • Page 111 Cytometer Turn Off. > IMPORTANT If you have the Sample Injection Mode Control Kit installed on your CytoFLEX Series instrument, refer to APPENDIX C, Sample Injection Mode Control Kit for detailed instructions on switching from the single tube sample probe to the Plate Loader.

  • Page 112: Running The System Startup Program With The Single Tube Loader

    Daily Startup Logging Into the Software Load Select to load the plate. Select . The plate loader automatically loads the plate holder stage and begins to acquire data. Record When you are satisfied with the data, select to record the data. Stop Wait for the data acquisition to finish or select .
  • Page 113 Daily Startup Logging Into the Software System Startup Program Select in the Cytometer menu. [CytoFLEX LX Shown] The System Startup Program window appears. Select Initialize. System Startup Program Window in Semi-Automatic Injection Mode 3-15 B49006AH...
  • Page 114 Daily Startup Logging Into the Software System Startup Program Window in Manual Injection Mode Wait for the system to initialize. Follow the on screen software prompts, then select Start. The instrument begins to prime. This process takes about 1 minute. After priming, the system initializes again.

  • Page 115: Running The System Startup Program [With Plate Loader]

    Daily Startup Logging Into the Software The sample tube is unloaded after sample acquisition has finished. The system uses the remaining time to warm up. Close When warm up is finished, select to quit the startup program. The system is now initialized.
  • Page 116 Daily Startup Logging Into the Software System Startup Program Select from the Cytometer menu to open the System Startup Program wizard. [CytoFLEX LX Shown] 3-18 B49006AH...
  • Page 117 Daily Startup Logging Into the Software The plate loader automatically ejects the plate holder stage and the System Startup Program window appears. 3-19 B49006AH...
  • Page 118 Daily Startup Logging Into the Software Set As Deionized Follow the on screen software prompts and select the desired wells and select Water Well NOTE To deselect water wells, select the desired well and select Set As Empty Well. NOTE Prepare two to six sample wells with deionized water.
  • Page 119 Daily Startup Logging Into the Software Wait for the system to initialize. The instrument begins to prime. This process takes about 1 minute. 3-21 B49006AH...
  • Page 120 Daily Startup Logging Into the Software After priming, the system initializes again. The sample is loaded automatically. This process takes about 1 minute per well. 3-22 B49006AH...
  • Page 121 Daily Startup Logging Into the Software When the system finishes acquiring the selected sample well, it uses the remaining time to warm up. 3-23 B49006AH...

  • Page 122: Selecting Experiments From The Start Page

    Daily Startup Logging Into the Software Close When warm up is complete the plate loader ejects the plate holder stage. Select to quit the startup program. The system is now initialized. Selecting Experiments from the Start Page Refer to Start Page CHAPTER 2, Using the CytExpert Software.

  • Page 123: Initializing The Instrument

    Daily Startup Initializing the Instrument Initializing the Instrument Select Initialize in the Data Acquisition Control screen or select Initialize in the Cytometer Menu to initialize the instrument. NOTE If the instrument is in Semi-Automatic Injection mode during the initialization process, the sample tube holder automatically shifts into the sample loading position (see Figure 1.12).
  • Page 124 Daily Startup Initializing the Instrument 3-26 B49006AH...

  • Page 125: Instrument Quality Control And Standardization

    CHAPTER 4 Instrument Quality Control and Standardization Overview This chapter provides information on performing daily quality control (QC) on the CytoFLEX and CytoFLEX LX flow cytometer and how to confirm that the instrument is working properly within the specified parameters. Quality control allows you to determine whether your instrument can provide adequate signal strength and precision.
  • Page 126 If the system is unable to adjust these parameters to fall within the operating limits, the system notifies you. NOTE Beckman Coulter recommends performing QC on a daily basis. NOTE QC can only be run in channels with the standard detector sets including, laser and band-pass. Refer...

  • Page 127: Preparing The Qc Sample

    Instrument Quality Control and Standardization Preparing the QC Sample Preparing the QC Sample Required Materials The following materials are required to complete the QC process: • CytoFLEX Daily QC Fluorospheres • CytoFLEX Sheath Fluid or another nonionic sheath fluid • Sample tubes (12 x 75 mm). •...

  • Page 128: Preparing The Qc Sample [With Plate Loader]

    Instrument Quality Control and Standardization Preparing the QC Sample [With Plate Loader] Use the vortexer or shake vigorously to thoroughly mix the bottle of CytoFLEX IR Daily QC Fluorospheres. Pipette 100 μL of CytoFLEX IR Daily QC Fluorospheres into the sample tube. Vortex the sample tube until a uniform mixture is achieved.

  • Page 129: Preparation Process Cytoflex Ir Daily Qc Fluorospheres

    Instrument Quality Control and Standardization Preparing the QC Sample [With Plate Loader] Add one drop of CytoFLEX Daily QC Fluorospheres to the sample well. Add approximately 200 L of deionized water to the sample well. Place the well plate in a dark location at 2-8 C until ready to load the well plate into the instrument for QC.

  • Page 130: Importing Lot-Specific Target Values

    Instrument Quality Control and Standardization Importing Lot-Specific Target Values Importing Lot-Specific Target Values Import lot-specific target values for each new lot of CytoFLEX QC Fluorospheres and CytoFLEX IR Daily QC Fluorospheres. CAUTION Risk of erroneous QC results. Different target value information correspond to different lot numbers.
  • Page 131 Instrument Quality Control and Standardization Importing Lot-Specific Target Values IMPORTANT The Beckman Coulter website may prompt you to select your Region and Country prior to the Beckman Coulter Technical Documents and Software page. Download Target File Select . The Beckman Coulter Technical Documents and Software Downloads page appears.
  • Page 132 Instrument Quality Control and Standardization Importing Lot-Specific Target Values g. Select English from the Language drop-down menu. Search Select The search results appear below the Search By Lot Number tab. B49006AH...
  • Page 133 Instrument Quality Control and Standardization Importing Lot-Specific Target Values CytoFLEX QC Fluorospheres Target Values Select under the Software Name column. The CytoFLEX QC Fluorospheres Target Values page appears. Download Select under the correct lot number from the CytoFLEX QC Fluorospheres Target Values page.

  • Page 134: Collecting Qc Data

    Ensure that the instrument configuration is properly configured for the QC experiment. The QC experiment may not be completed or may end in erroneous results if incorrect settings are chosen. Beckman Coulter recommends using the factory configuration and ensuring that the proper optical filters are in place.
  • Page 135 Instrument Quality Control and Standardization Collecting QC Data Start QC/Standardization Select in the QC/Standardization menu to access the QC experiment. [CytoFLEX LX Shown] Ensure that the QC bead lot number is selectable in the Lot No. drop down menu. If the lot number is not selectable, refer to Importing Lot-Specific Target Values, then select the proper...
  • Page 136 Instrument Quality Control and Standardization Collecting QC Data [With Plate Loader] During QC, the software automatically seeks the CytoFLEX Daily QC Fluorospheres and computes the results. The software returns to the QC Report screen after the QC run is complete. If the sampling rate is too low, the Cytometer stops the QC run and displays a prompt alerting you that the QC run failed to reach the required event flow rate.
  • Page 137 Ensure that the instrument configuration is properly configured for the QC experiment. The QC experiment may not be completed or may end in erroneous QC results if incorrect settings are chosen. Beckman Coulter recommends using the factory configuration and ensuring that the proper optical filters are in place.
  • Page 138 Instrument Quality Control and Standardization Collecting QC Data [With Plate Loader] Initialize Select Select Eject Insert the prepared QC well plate (see Preparation Process CytoFLEX Daily QC Fluorospheres) into the plate holder. Select . The Plate Settings window appears. IMPORTANT Ensure the well position on the plate matches the well position selected in the software.
  • Page 139 Instrument Quality Control and Standardization Collecting QC Data [With Plate Loader] Select the desired plate type from the Plate Type dropdown menu. Select the Mix and Backflush settings in the top of the Plate Settings window. Select Start Select to load the sample and begin to run the QC procedure. The message Please confirm that the correct plate is placed properly and press OK appears.

  • Page 140: Confirming Results

    Instrument Quality Control and Standardization Confirming Results During QC, the software automatically seeks the CytoFLEX Daily QC Fluorospheres and computes the results. The software returns to the QC screen after the QC run is complete. If the sampling rate is too low, the Cytometer stops the QC run and displays a prompt that the QC run fails to reach the required event flow rate.
  • Page 141 Instrument Quality Control and Standardization Confirming Results Select a QC run from the QC Process list on the left and a QC report appears on the right. NOTE The results column indicates a passing QC result with a and a failed QC result with QC results must meet the following criteria to pass: •...
  • Page 142 Instrument Quality Control and Standardization Confirming Results The report area on the right displays detailed experiment results, including laser power, delay, testing conditions, and signal results. The same symbols are used to indicate each result. For items that fail, values falling outside the prescribed range are displayed in red font. In the Comment area, an explanation appears for each failed item.

  • Page 143: Creating Levey-Jennings

    Instrument Quality Control and Standardization Confirming Results d. Run Daily Clean CHAPTER 10, Cleaning Procedures, and retest. e. Run Deep Clean Procedure CHAPTER 10, Cleaning Procedures, and retest. Repeat Steps c-d. NOTE If QC fails two times in a row on the same day after repeating Steps a-f, contact If necessary, you can select (for CSV format) or...
  • Page 144 Instrument Quality Control and Standardization Confirming Results Laser Select the tab, and select the power and/or delay checkboxes for each laser as needed. Select the Channels tab, and select each channel checkbox as needed. 4-20 B49006AH...

  • Page 145: Qc Result Manager

    Instrument Quality Control and Standardization Confirming Results Apply Select Select Select the Levey-Jennings plot and select the start and end date from the drop down boxes at the top of the LJ Chart screen to specify the desired date range. NOTE Select the desired configuration and date range from the drop-down menus located at the top of the LJ Chart screen to sort by the configuration used during the specified date range.

  • Page 146: Standardization

    QC screen. The QC Result Manager window appears. Standardization Use Beckman Coulter CytoFLEX Daily QC fluorospheres or any other reference material that is relevant for your application. Ensure that either the Beckman Coulter CytoFLEX Daily QC fluorospheres or other reference material has optimized settings appropriate for your experiment.

  • Page 147: Adding, Editing, Importing, And Deleting Standardization Laser

    Instrument Quality Control and Standardization Standardization Adding, Editing, Importing, and Deleting Standardization Laser Target Values in the QC Experiment Standardization Target Library... Select from the Settings menu. The standardization Target Library window appears. NOTE The Item name displays in the Acquisition Setting Catalog window as the saved acquisition setting name.

  • Page 148: Adding A New Standardization

    Instrument Quality Control and Standardization Standardization Adding a New Standardization Item Select . The Add Standardization Target Value window appears. NOTE Select to create a duplicate copy of an existing standardization item. NOTE Unchecking the Use the same threshold setting for acquisition setting checkbox allows you to specify custom threshold settings and when to save the test item into the Acquisition Setting Catalog.

  • Page 149: Editing Standardization Item

    Instrument Quality Control and Standardization Standardization Choose either Manual or Automatic threshold from the Standardization Test Setting or Acquisition Setting section of the screen. NOTE If you select Manual threshold, enter a value greater than 0, but less than 8,388,600. Set the channels, median, and median tolerance values.

  • Page 150: Importing A Standardization

    Instrument Quality Control and Standardization Standardization Edit the parameters for that item and select NOTE Task Item, Lot No., and Expire date cannot be edited. Ensure the item parameters are correct then select and save the file. Select to exit the Standardization Target Library window. Importing a Standardization Item Select on the Standardization Target Library window.
  • Page 151 Instrument Quality Control and Standardization Standardization Standardization Select the radio button. Select the Lot No. and the Items to be applied. 4-27 B49006AH...
  • Page 152 Instrument Quality Control and Standardization Standardization Select The Process section of the screen displays the process details. Once the process is complete, the Custom Setting Calibration Report displays. 4-28 B49006AH...
  • Page 153 Instrument Quality Control and Standardization Standardization Verify the gain settings. a. Select Acq. Setting Catalog from the Cytometer menu. The Acq. Setting Catalog window appears. NOTE designates test items from Standardization. b. Select the desired Test Item, from the list in the Gain, Threshold, and Width tabs. 4-29 B49006AH...
  • Page 154 Instrument Quality Control and Standardization Standardization 4-30 B49006AH...

  • Page 155: Chapter 5: Data Acquisition And Sample

    CHAPTER 5 Data Acquisition and Sample Analysis Overview This chapter contains information on how to use your CytoFLEX and CytoFLEX LX flow cytometer, including data acquisition, analyzing and exporting results, and compensation procedures that will be executed manually during the process. Workflow: Adjust instrument Create...
  • Page 156 Data Acquisition and Sample Analysis Creating an Experiment Verify the laser settings. Refer to Laser Settings CHAPTER 5, Data Acquisition and Sample Analysis. Create or open an experiment using one of the following methods: • Create a new experiment: — Select New Experiment on the Start page, specify the file path, and save the experiment.

  • Page 157: Creating An Experiment

    Data Acquisition and Sample Analysis Creating an Experiment [With Plate Loader] NOTE If you are using either the CytExpert Default Software Option or the CytExpert User Management Software Option: If you need to change the default save path, select Options in the Settings menu and modify the Default Path displayed to the right of the Experiment tab.
  • Page 158 Data Acquisition and Sample Analysis Creating an Experiment [With Plate Loader] Select . The Plate window appears. [CytoFLEX LX Shown] B49006AH...
  • Page 159 Data Acquisition and Sample Analysis Creating an Experiment [With Plate Loader] Add Plate Select the Add Plate dropdown and select [CytoFLEX LX Shown] The Add Plate window appears. NOTE Select Add Plate from Template to add a plate template with preset plate settings. NOTE Select Duplicate Current Plate without Data to create a copy of the selected plate without data.
  • Page 160 Data Acquisition and Sample Analysis Creating an Experiment [With Plate Loader] Select the direction of the workflow from the sampling sequence from the plate section of the window. The Plate window appears. [CytoFLEX LX Shown] If you need to automatically turn off the cytometer after auto acquisition is complete [CytoFLEX LX Only]: a.

  • Page 161: Setting Sample Wells

    Data Acquisition and Sample Analysis Creating an Experiment [With Plate Loader] Setting Sample Wells Once the plate protocol is created, the plate window appears. Refer to Figure 5.1. Figure 5.1 Plate Window [CytoFLEX LX Shown] Empty well The well with color is set as sample well, but is not set for Auto Record The well is set as sample well, and is ready for Auto Record.
  • Page 162 Data Acquisition and Sample Analysis Creating an Experiment [With Plate Loader] Left-click and drag your mouse to highlight the desired wells or hold the Control key and select each desired well. Set As Sample Wells Select or right-click the selected wells and select .
  • Page 163 Data Acquisition and Sample Analysis Creating an Experiment [With Plate Loader] Select the desired acquisition settings under the Acq. Setting tab. NOTE Select Import from File to import the settings from a FCS file. NOTE If desired, import saved settings/standardization settings from the catalog. Select the channels and create label names under the Channel tab.
  • Page 164 Select Events to record or time to record under the Stopping Rules tab. NOTE Beckman Coulter recommends setting an acquisition time limit, so the acquisition will not continue if the event limit cannot be reached. 5-10...

  • Page 165: Modifying Well

    Data Acquisition and Sample Analysis Creating an Experiment [With Plate Loader] Select Modifying Well Settings If any well settings require modification, right-click the sample well and select the setting to change. [CytoFLEX LX Shown] 5-11 B49006AH...

  • Page 166: Applying Existing Well Settings To Additional

    Data Acquisition and Sample Analysis Creating an Experiment [With Plate Loader] Applying Existing Well Settings to Additional Wells Apply Well Settings to Right-click the well with the desired settings and select . The Apply Well Settings of window appears. NOTE The Convert Compensation Matrix by the Gain of Tube checkbox is only available when Acq.

  • Page 167: Moving The Location Of A

    Data Acquisition and Sample Analysis Creating an Experiment [With Plate Loader] Copy Right-click and select NOTE When Copy or Cut is selected, the well displays as follows: Right-click an empty well and select Paste NOTE The same number of wells will paste in the same orientation the wells were selected. Moving the Location of a Well Right-click the desired well to move and select Move location...
  • Page 168 Data Acquisition and Sample Analysis Creating an Experiment [With Plate Loader] b. Select or right-click and select Auto Record to set the selected wells for auto record. Number labels appear in the bottom right corner of each well set for auto record. Sample acquisition occurs in the order indicated by the numbers.

  • Page 169: Creating A Heat

    Data Acquisition and Sample Analysis Creating an Experiment [With Plate Loader] Creating a Heat Map Select from the Tube management controls (refer to Test Tubes CHAPTER 2, Using the CytExpert SoftwareCHAPTER 5, Data Acquisition and Sample Analysis). The Heat Map window appears.
  • Page 170 Data Acquisition and Sample Analysis Creating an Experiment [With Plate Loader] Select from the Heat Map window. The New Heat Map window appears. NOTE You must have at least one plate that contains data in at least two wells to create a new heat map. NOTE Select Display Value to display the value within the heat map well on the Heat Map window.
  • Page 171 Data Acquisition and Sample Analysis Creating an Experiment [With Plate Loader] Select to add a parameter item to the Parameter section of the New Heat Map window. NOTE A maximum of 6 parameters can be added to a single heat map. NOTE to delete a parameter.
  • Page 172 Data Acquisition and Sample Analysis Creating an Experiment [With Plate Loader] d. Select the Use Custom Ranges checkbox if needed and enter the Min and Max ranges. NOTE The Actual Range displays when the statistics parameter can be calculated. Select any wells that should be excluded from the heat map in the Wells section of the New Heat Map window then select NOTE designates that a well is included.
  • Page 173 Data Acquisition and Sample Analysis Creating an Experiment [With Plate Loader] Edit the color elements in the Color section of the New Heat Map window. a. Select a color from the base color drop down menu. b. Select the number of color bands desired from the Bands drop down menu. The window refreshes to display the appropriate number of bands.
  • Page 174 Data Acquisition and Sample Analysis Creating an Experiment [With Plate Loader] Select Fixed Range to assign colors based on a fixed range specified by the user. The heat map is created directly based on the result of the expression. The color of the heat map displays according to the legend range.

  • Page 175: Refreshing A Heat

    Data Acquisition and Sample Analysis Creating an Experiment [With Plate Loader] Refreshing a Heat Map When data displayed in a heat map is no longer current, the symbol appears next to the heat map name and the message Analysis changed. Please refresh. appears on the Heat Map window. Select from the heat map toolbar to refresh the analysis.

  • Page 176: Modifying Existing Heat Map

    Data Acquisition and Sample Analysis Creating an Experiment [With Plate Loader] Modifying Existing Heat Map Settings Select from the heat map toolbar to modify existing heat map settings. The Modify Heat Map Settings window appears. 5-22 B49006AH...

  • Page 177: Deleting An Existing Heat

    Data Acquisition and Sample Analysis Creating an Experiment [With Plate Loader] Deleting an Existing Heat Map To delete a single heat map, select the heat map to be deleted from the list of heat maps in the Heat Map window then select from the heat map toolbar.

  • Page 178: Load Sample And Record Data

    Data Acquisition and Sample Analysis Load Sample and Record Data Load Sample and Record Data Before Running Samples CAUTION Risk of erroneous results if the Cytometer has been idle for an extended period of time. Perform a prime if the system has been idle for an extended period of time (see Priming the Flow Cell CHAPTER 11, Replacement/Adjustment...
  • Page 179 Data Acquisition and Sample Analysis Load Sample and Record Data Verifying, Selecting, Editing, and Creating Detector Configuration CAUTION Risk of erroneous results. The system will read the selected Detector Configuration even if the optical filters do not match the selected Detector Configuration.
  • Page 180 Data Acquisition and Sample Analysis Load Sample and Record Data If a saved configuration requires changes, edit that configuration. NOTE The factory configuration is in bold and cannot be edited. a. Select the configuration, then select Edit to access the Edit Detector Configuration screen. b.
  • Page 181 Data Acquisition and Sample Analysis Load Sample and Record Data d. Ensure the new configuration is highlighted, then select Edit . The Edit Detector Configuration window appears. e. Customize the new configuration. Channels with a white background can be edited. Drag the names of the appropriate fluorescence channels and optical filters on the left to the correct channels.
  • Page 182 Data Acquisition and Sample Analysis Load Sample and Record Data When finished, select Select the appropriate configuration. Verify that the correct optical filters are installed in the Cytometer and match the newly created configuration. Set As Current Select Select Delete To delete a configuration created in error, select .
  • Page 183 Setting Up Violet Side Scatter (VSSC) Channel For microparticles, a VSSC option can be added to better separate side scatter signals from noise. Beckman Coulter recommends using this channel to detect particles smaller than 500 nm. NOTE Since the total available channel numbers remain the same when the VSSC channel is used, the number of fluorescent channels in the violet WDM is reduced by 1 channel.
  • Page 184 Place the third filter in position 1, the 405-nm filter in position 2, and the 450-nm filter in position 3. NOTE For the Violet WDM, Beckman Coulter recommends placing the filters in sequential order from the shortest wavelength to the longest wavelength in positions 2 to 6. Position 1 will always contain the unused filter.
  • Page 185 Data Acquisition and Sample Analysis Load Sample and Record Data Save As Select the Default Configuration and select . The Configuration Save As window appears. Name the new configuration VSSC and select Select the VSSC configuration and select Edit . The Edit Detector Configuration window appears. Change the filters and channel names according to the filter order in the violet WDM.

  • Page 186: Sampling And Collecting

    Data Acquisition and Sample Analysis Load Sample and Record Data Set SSC Right-click the VSSC channel, and select to set it as the Violet SSC channel. Select to save the changes and close the Edit Detector Configuration window. Set as Current Select Select to save the changes and close the Detector Configuration window.
  • Page 187 Data Acquisition and Sample Analysis Load Sample and Record Data Change the tube name if necessary. Refer to Changing the Tube Name. Mix the sample tube intended for testing. Ensure that the sample tube holder is in the sample loading position (see Figure 1.12).
  • Page 188 Data Acquisition and Sample Analysis Load Sample and Record Data Select to load the sample. NOTE When you select a tube that only contains acquired data, as indicated by the blue tube in the test tube section of the screen, the following message appears: •...
  • Page 189 Data Acquisition and Sample Analysis Load Sample and Record Data • Append data to existing file. Adds new data to the existing data. NOTE When you select a tube that only contains acquired data, as indicated by the blue tube in the test tube section of the screen, the following message appears: •...

  • Page 190: Configuring Acquisition

    Data Acquisition and Sample Analysis Configuring Acquisition Settings Configuring Acquisition Settings Changing the Tube Name To change the name of a new sample tube or the sample ID, right-click the tube name or the sample Edit Name ID name in the Tube section of the screen and select or simply double-click the sample tube or sample ID name.
  • Page 191 Data Acquisition and Sample Analysis Configuring Acquisition Settings In the Set Channel window, modify which channels are used and how they are displayed. a. Select the channel signal checkbox, then you can add the reagent name in the Label column. The information you add appears in the corresponding axis of the relevant plot in the plot area.
  • Page 192 Data Acquisition and Sample Analysis Configuring Acquisition Settings d. If you only need to modify the label name, select Set Label in the Settings menu to make the required changes. The Set Label screen appears. The Set Label screen does not allow you to select which channels to use, but it does allow you to apply the modified label to all the sample tubes.
  • Page 193 Data Acquisition and Sample Analysis Configuring Acquisition Settings Creating Plots and Gates Use the plotting controls (see Figure 2.1) in the plot area to create plots and gates and to generate graphs as shown. Use the icons to generate histograms, dot plots, density plots, pseudo color plots, and contour plot.
  • Page 194 Data Acquisition and Sample Analysis Configuring Acquisition Settings b. Select an axis name to change which channel is displayed. An “A” after the channel name indicates signal pulse area, while an “H” indicates height. The default setting is "A". NOTE To modify the default settings, select Options in the Settings menu.
  • Page 195 Data Acquisition and Sample Analysis Configuring Acquisition Settings c. Signal width can be used as a tool for doublet discrimination and to differentiate somatic cell adhesion. If necessary, select to open the Acq. Setting window. 5-41 B49006AH...
  • Page 196 Data Acquisition and Sample Analysis Configuring Acquisition Settings d. Select the Width tab, and select a channel with the required signal width. e. Plot properties can be configured to display axes in Log, Log-Linear, or Linear format. 1) Double-click the plot or right-click the plot and select Property from the drop-down menu.
  • Page 197 Data Acquisition and Sample Analysis Configuring Acquisition Settings 2) Select whether to display the axes in logarithmic or linear format for both the X-axis and Y-axis. Enter a value for log-linear coefficient if the log-linear view is desired. 3) Select Close Select the logarithmic axis on the plot.
  • Page 198 Data Acquisition and Sample Analysis Configuring Acquisition Settings • Select to shift the axes. The mouse pointer appears as a hand. It allows you to drag the graph to reveal the axis segment you need. — Pan: Modifies the axis display range dimensions when panning both axes. When the pan control is selected, you can right-click the graph and select which axis you need to adjust when dragging.
  • Page 199 Data Acquisition and Sample Analysis Configuring Acquisition Settings To create gates, use the control buttons or right-click the plot and select the gate type required. Gates can be set according to different requirements to differentiate cell populations. NOTE To add a vertex to a polygon gate: 1.

  • Page 200: All Gates - Example

    Data Acquisition and Sample Analysis Configuring Acquisition Settings Select the gates to display. a. Select the heading area of the plot, select the parent population/gate to display in the plot from the drop-down menu. The selected parent gate appears in the tab area of the plot. NOTE The CytExpert software will not list gates which would create circular gating logic.
  • Page 201 Data Acquisition and Sample Analysis Configuring Acquisition Settings b. If necessary, you can select the Combo Population option from the drop-down menu to create a combination gate, using the boolean relationships “and”, “or”, and “not” to produce a new gate. You can also select the population color or change the gate name. •...

  • Page 202: Creating And Adjusting Auto

    Data Acquisition and Sample Analysis Configuring Acquisition Settings The Population Hierarchy function allows you to view how gates rank in relation to one another. To change the display color, double-click the default color and select the desired color from the drop-down color palette.
  • Page 203 Data Acquisition and Sample Analysis Configuring Acquisition Settings Select the population you want to gate in the histogram to automatically gate that population. To create an auto polygon gate, select from the toolbar or right-click on the 2D plot and select Auto Polygon from the drop down menu.

  • Page 204: Turning Auto Recalculate

    Data Acquisition and Sample Analysis Configuring Acquisition Settings Select the population you want to gate in the 2-D plot. The gate will automatically be drawn to fit the population. NOTE To add a vertex to an auto polygon gate: 1. Select the gate. 2.

  • Page 205: Adjusting Autogate Movement And Laser Settings

    Data Acquisition and Sample Analysis Configuring Acquisition Settings Adjusting Autogate Movement and Extent Movement — The distance an autogate can move to find the target population. To adjust movement, right-click an autogate and drag the Movement handle in the auto gate menu left and right.

  • Page 206: Movement - Max

    Data Acquisition and Sample Analysis Configuring Acquisition Settings Figure 5.5 Movement - Max Setting Extent — Shrinks or expands the gate around the population. To adjust extent, right-click an autogate and drag the Extent handle in the auto gate menu left and right.

  • Page 207: Extent - Maximum

    Data Acquisition and Sample Analysis Configuring Acquisition Settings Figure 5.7 Extent - Maximum Setting Laser Settings To access the Laser Setting window, select Advanced Laser Setting . The Laser Setting window > appears. Refer to Figure 5.8. NOTE The instrument must be in Standby mode to access the Laser Setting window. Figure 5.8 Laser Setting Window [CytoFLEX LX shown] 1.
  • Page 208 Data Acquisition and Sample Analysis Configuring Acquisition Settings Enable Disable Select the radio button next to each laser on the Laser Setting window to enable or disable lasers. The laser status for each laser displays in the software status bar. Hover your mouse over to display details for each laser.

  • Page 209: Adjusting The

    Data Acquisition and Sample Analysis Configuring Acquisition Settings Standardize your laser target values in the QC Experiment. Refer to Standardization CHAPTER 4, Instrument Quality Control and Standardization. NOTE Disabled lasers are marked Laser XXX is disabled in the QC screen and do not provide laser power values.
  • Page 210 Data Acquisition and Sample Analysis Configuring Acquisition Settings Adjust the gain setting of each channel under the Gain tab in the Acq. Setting window. Raising the gain increases the signal. Lowering the gain reduces the signal. Gain Control Another option is to use the button on the toolbar in the graphic control area to adjust the gain values for cell population data to their desired levels, directly on the plots...
  • Page 211 Data Acquisition and Sample Analysis Configuring Acquisition Settings Adjusting the Threshold By adjusting the threshold, the user can remove unnecessary signal noise to ensure that most of the data collected consists of desired signal data. After the threshold settings have been configured for a given channel, the acquisition of data from this channel will only be triggered by signals that exceed the established threshold.
  • Page 212 Data Acquisition and Sample Analysis Configuring Acquisition Settings Set the desired threshold using one of the following methods: • Choose the channel that is used for setting the threshold. Manually enter the threshold value in the Threshold tab. NOTE For dual-parameter plots, you can right-click the plot and select both parameters if desired. Then, select the desired threshold boundary for the second parameter.
  • Page 213 Data Acquisition and Sample Analysis Configuring Acquisition Settings For example, if the event to record is set to record 1,000 P1 events, the software automatically stops recording when P1 events reach 1,000 events. However, the software saves all data acquired, including events outside of P1, when 1,000 P1 events is reached. You can also specify the time to store if necessary.
  • Page 214 Data Acquisition and Sample Analysis Configuring Acquisition Settings Setting Plot Display Conditions Select Events Display Setting in the Settings menu. The Events Display Setting window appears. Three display options are available: • Display all events. Used to view all events on the plot. •...
  • Page 215 Data Acquisition and Sample Analysis Configuring Acquisition Settings Setting Customized Parameters Set custom parameter to create fluorescence calculations. Select Set Customized Parameter from the Settings menu. Or, right-click a test tube from the test tube menu and select Set Customized Parameter .
  • Page 216 Data Acquisition and Sample Analysis Configuring Acquisition Settings Setting Custom Statistics Set custom statistics to create calculations based on populations of interest. Right-click the statistics table and select Statistics Setting . The Statistics Setting window appears. Select Expression 5-62 B49006AH...
  • Page 217 Data Acquisition and Sample Analysis Configuring Acquisition Settings Edit Select . The Expression window appears. Enter the expression name in the Name section and enter the expression using the equation buttons. 5-63 B49006AH...
  • Page 218 Data Acquisition and Sample Analysis Analyzing and Exporting Data Select NOTE The equation populates in the Statistics Setting window under the Expression selection. Analyzing and Exporting Data Select the sample tube to be analyzed. 5-64 B49006AH...
  • Page 219 Data Acquisition and Sample Analysis Analyzing and Exporting Data Establish new gates or adjust the position of existing gates. Refer to Creating Plots and Gates. NOTE Changing a gate’s position does not affect the positions of other gates already established on a given sample tube.
  • Page 220 Data Acquisition and Sample Analysis Analyzing and Exporting Data Bring population to front Right-click the plot and select to make the display color of the specified gate appear in front of all other colors, or select Send population to back to hide the display color of the specified gate behind all other colors.
  • Page 221 Data Acquisition and Sample Analysis Analyzing and Exporting Data The Statistics Setting window allows you to change the display of the header, statistical elements and cell populations included. 5-67 B49006AH...
  • Page 222 Data Acquisition and Sample Analysis Analyzing and Exporting Data The final generated plots appear as below. Right-click a plot and select Export to Clipboard Export to Graphic File from the drop-down menu to select an image to export. • Export to Clipboard copies the plot to the clipboard, allowing you to paste it directly into documents in common file formats.
  • Page 223 Ensure that any storage devices used with the instrument are free from viruses. To guard against data loss, Beckman Coulter recommends backing up data on a frequent and regular basis. Beckman Coulter is not liable for any loss of data resulting from computer viruses or damage to hardware.
  • Page 224 Data Acquisition and Sample Analysis Analyzing and Exporting Data Exporting FCS Files Exporting Single Tube Files Export FCS File Right-click the desired tube from the test tube section of the screen and select The Export FCS File window appears. Select the population from the Population dropdown menu. Area Height Select either...
  • Page 225 Data Acquisition and Sample Analysis Analyzing and Exporting Data Select the FCS format next to File Version. NOTE The default setting is FCS 3.0. If FCS 2.0 is selected, select the parameter type (linear or log) from the parameter type section of the window. NOTE The default CytExpert FCS file contains high auto-fluorescence vector values that may not be recognized by third party software.
  • Page 226 Data Acquisition and Sample Analysis Analyzing and Exporting Data Exporting Multiple FCS Files Export FCS File Select from the File menu. The Export FCS File window appears. Select the tubes to export. Repeat Steps from Exporting Single Tube Files. 5-72 B49006AH...

  • Page 227: Experiment

    Data Acquisition and Sample Analysis Analyzing and Exporting Data Exporting Plots or the Statistics Table of Multiple Tubes as Picture Files Select File > Export FCS File . The Export Tubes to Files window appears. Select the tubes to export. Select the path.

  • Page 228: Importing Instrument

    Data Acquisition and Sample Analysis Analyzing and Exporting Data Importing Instrument Settings Select the desired sample tube to import. Then select NOTE Instrument settings can only be imported into tubes where data has not yet been recorded. Select Import From File , locate the file with the required instrument settings, or select Import From Catalog...

  • Page 229: Importing And Exporting Compensation

    Data Acquisition and Sample Analysis Analyzing and Exporting Data Export To File Select to export a current set of instrument settings, stored in a file ending in .acq. Export To Catalog Select , give a name to the settings to be exported, and export the file to the software’s Acquisition Setting Catalog, then select Select Close...
  • Page 230 Data Acquisition and Sample Analysis Analyzing and Exporting Data Select in the printer control area to print directly. Or, select the print drop-down arrow for the following options: • Print Preview. Used to access the Preview screen. — Select to select the required format of the file to be exported and to save the file in that format.
  • Page 231 Data Acquisition and Sample Analysis Analyzing and Exporting Data • Batch Print. Used to print data for multiple tubes. 1. Select Batch Print . The Batch Print window appears. 2. Select the tubes to print. 3. Select • Batch Export to PDF File. Used to print a PDF of the data for multiple tubes.

  • Page 232: Saving The Experiment

    Data Acquisition and Sample Analysis Saving the Experiment Saving the Experiment Save Selecting in the File menu allows you to save the experiment. Save As Selecting and saving the experiment under a different name allows you to create a backup. Selecting Save As Template in the File menu allows you to save the experiment as a template.

  • Page 233: Chapter 6: Compensation

    CHAPTER 6 Compensation Overview This chapter describes how to create a compensation experiment and automatically calculate compensation values after acquiring the data. It also explains how to use these calculations for other experiments. Compensation involves correction for fluorescence spillover emitted by the primary fluorochrome that is detected by the secondary fluorescent channels.
  • Page 234 Compensation Creating a Compensation Experiment Workflow: Apply Prepare Create Acquire and Calculate compensation Compensation Ô compensation Ô Ô Ô record data compensation to sample for Samples experiment data analysis This chapter contains information on: • Creating a Compensation Experiment • Creating a Compensation Experiment [With Plate Loader] •...
  • Page 235 Compensation Creating a Compensation Experiment If a negative population is not present in each single color tube, then an unstained control tube is recommended. NOTE The default selection is Area. The unstained negative control tube can be selected if needed. NOTE Label and lot number information can be retained in the Compensation Library to facilitate future compensation calculations.
  • Page 236 Compensation Creating a Compensation Experiment Select After confirmation, the software automatically generates the following compensation experiment. NOTE Select Area to calculate compensation based on the Area measured. Alternatively, select Height to calculate compensation based on the Height measured. Preparing the Compensation Sample To perform a compensation experiment, prepare: •...
  • Page 237 Compensation Creating a Compensation Experiment CAUTION Risk of erroneous results. Calculations based on excessively small volumes of sampled data can be inaccurate. Ensure that more than 1,000 positive events and more than 1,000 negative events are sampled. If the ratio of positive cells is comparatively low, increase the number of acquisition events to a suitable amount.
  • Page 238 Compensation Creating a Compensation Experiment Set an appropriate number of cells to save in Events to Record located on the left side of the screen. Record Select to save the data. Running the Single Positive Control Samples Place the single positive tube in sample loading position (see Figure 1.12).

  • Page 239: Positive Population Selected From The Unstained

    Compensation Creating a Compensation Experiment CAUTION Risk of erroneous results. Calculations based on excessively small volumes of sampled data can be inaccurate. Ensure that more than 1,000 positive events and more than 1,000 negative events are sampled. If the ratio of positive cells is comparatively low, increase the number of acquisition events to a suitable amount.

  • Page 240: Positive And Negative Populations Without An Unstained Sample

    Compensation Creating a Compensation Experiment NOTE Figure 6.4 shows an example of selecting both the positive and negative populations without an unstained sample. Figure 6.4 Positive and Negative Populations Without an Unstained Sample 1. Negative population 2. Positive population Record Select Repeat steps to acquire data from subsequent single positive sample tubes.
  • Page 241 Compensation Creating a Compensation Experiment Compensation Calculation Select or select in the Compensation menu to calculate the compensation values. The Compensation Matrix window appears, displaying the calculated compensation values. NOTE The primary fluorescence channels are listed in columns; the secondary fluorescence channels are listed in rows.

  • Page 242: Creating A Compensation

    Compensation Creating a Compensation Experiment [With Plate Loader] Specify the key words and select NOTE The settings stored in the compensation library are specific to the detector configuration. The compensation library can only be applied when the detector configurations are the same. At any time, saved compensation experiments can be reopened and the compensation values recalculated.
  • Page 243 Compensation Creating a Compensation Experiment [With Plate Loader] Select the plate type and sampling sequence located in the top, right of the Compensation Setup window. CAUTION Risk of erroneous results. Select an unstained tube, according to which the fluorescence background will be set. If there is not an unstained tube, then each single marker tube must have a negative population.
  • Page 244 Compensation Creating a Compensation Experiment [With Plate Loader] Assign the well locations. a. Select the fluorochrome. b. Select the desired sample well location for the fluorochrome. c. Select Set As Sample Well NOTE The well location populates in the location column. d.

  • Page 245: Preparing The Compensation

    Compensation Creating a Compensation Experiment [With Plate Loader] Preparing the Compensation Sample To perform a compensation experiment, prepare: • A single positive control well for each color • A negative control well (optional) NOTE A negative control well is required if a single positive control well does not contain a negative population.
  • Page 246 Compensation Creating the Compensation Matrix from Previously Acquired Data Save To Compensation Library Select to save the single color compensation values in the compensation library. Specify the key words and select NOTE The settings stored in the compensation library are specific to the detector configuration. The compensation library can only be applied when the detector configurations are the same.

  • Page 247: Creating The Compensation Matrix From Previously Acquired

    Compensation Creating the Compensation Matrix from Previously Acquired Data Import FCS File Right-click on the appropriate test tube and select . Locate the corresponding data file and import the file. Only files compatible with the detector configuration are supported by the software for importing. in front of a test tube indicates that the corresponding data have been imported.

  • Page 248: Adjusting Compensation

    Compensation Adjusting Compensation Adjusting Compensation Manually Adjusting Compensation The compensation can be manually adjusted in an experiment in two ways: • Select the bivariate plot which the compensation needs to be adjusted. Select from the graphic control area, then click and drag the mouse pointer up and down or left and right inside the plot to adjust compensation.

  • Page 249: Importing Compensation Settings From The Compensation Library

    Compensation Adjusting Compensation After opening the desired compensation file, the Import Compensation window appears. Select one of the following: • Import compensation matrix and convert it with current gains. • Import compensation matrix. • Import compensation matrix and gain. NOTE •...
  • Page 250 Compensation Adjusting Compensation Import From Compensation Library Select to select which compensation values to import from the compensation library. [CytoFLEX LX Shown] In the Keywords column, the corresponding compensation values can be selected for each channel. The compensation values of the same keyword can also be selected using the drop- down menus in the Keywords column.

  • Page 251: Exporting Compensation

    Compensation Adjusting Compensation Exporting Compensation Settings Select the desired sample tube to export. Select Compensation Matrix in the Setting menu. Export Select to specify a path and filename for the compensation file you are saving. [CytoFLEX LX Shown] Select Save NOTE The generated file ends in .comp.

  • Page 252: Managing The Compensation

    Compensation Adjusting Compensation Managing the Compensation Library Compensation values can be managed in the Compensation Library. Select Compensation Library from the Settings menu. The Compensation Library window appears. NOTE The Compensation Library is arranged by fluorescence detection channels. Select the desired single color sample. The compensation information appears on the right side of the window.

  • Page 253: Adding Channels For

    Compensation Adjusting Compensation Adding Channels for Compensation Channels requiring compensation calculations that have not been previously acquired can be added to the compensation experiment by acquiring the necessary positive tubes. In the compensation experiment, select in the compensation controls, or select Compensation Setup in the Compensation menu.
  • Page 254 Compensation Adjusting Compensation 6-22 B49006AH...

  • Page 255: Chapter 7: Data Review

    CHAPTER 7 Data Review Overview This chapter discusses how to use the Analysis screen to analyze data. Data can be analyzed using any computer equipped with the CytExpert software. No online connection is required. Workflow: Import experiment or data Ô Plot and configure statistics Ô...
  • Page 256 Data Review Copying Experiments and Importing Data Import FCS File In the new or opened experiment, select in the File menu to import the data files. Imported data files appear in the Tube screen. symbol in front of each data tube indicates that the data tube is an imported data file. Imported data files are copied and saved in the folder where the current experiment data files are saved.

  • Page 257: Setting The Plots And Statistics

    Data Review Setting the Plots and Statistics Setting the Plots and Statistics Opening the Analysis Screen Select on the left to enter the Analysis screen. Copy plots obtained during data acquisition. a. If you need original plots used during data acquisition, select to access the Acquisition screen.
  • Page 258 Data Review Setting the Plots and Statistics New plots can be created according to need. After selecting the test tubes requiring analysis, use the plotting control buttons at the top of the screen to create a new plot. NOTE Each graph in the Analysis screen may correspond to different data. Pay special attention to each plot’s heading to avoid mistakes during analysis.

  • Page 259: Creating Histogram And Dot Plot Overlays

    Data Review Setting the Plots and Statistics Creating Histogram and Dot Plot Overlays The CytExpert software supports histogram and dot plot data overlay functionality, allowing you to combine data from differing sources onto the same histogram or dot plot. Select Histogram Overlay under the histogram icon drop-down list to create a new multi-data histogram.
  • Page 260 Data Review Setting the Plots and Statistics IMPORTANT A maximum of 10 samples can be overlaid. Select to select samples for overlay display. Or, drag and drop samples from the tube list on the left into the histogram or dot plot overlay. The software automatically assigns different colors to different data.

  • Page 261: Calculating Sample Volume And Concentration

    Data Review Calculating Sample Volume and Concentration Calculating Sample Volume and Concentration The CytoFLEX flow cytometer supports the calculation of the sample concentration based on the volume consumed and/or based on the known concentration of reference beads. NOTE If necessary, calibrate the sample uptake rate (see Calibrating the Sample Flow Rate CHAPTER 11, Replacement/Adjustment...

  • Page 262: Adjusting Compensation Settings

    Data Review Adjusting Compensation Settings Volume In the Statistics Setting screen, select and the concentration item to see the corresponding information in the statistics table. NOTE While collecting samples, instantaneous data calculation can appear inaccurate. Regard the calculation as accurate only after data acquisition has been completed. [CytoFLEX LX Shown] [CytoFLEX LX Shown] Adjusting Compensation Settings...

  • Page 263: Exporting Results

    Data Review Exporting Results Exporting Results Refer to CHAPTER 5, Data Acquisition and Sample Analysis. B49006AH...
  • Page 264 Data Review Exporting Results 7-10 B49006AH...

  • Page 265: Chapter 8: Daily Shutdown

    CHAPTER 8 Daily Shutdown Overview This chapter provides procedures for shutting down the CytoFLEX instrument. Workflow: Prepare the cleaning solution Ô Clean the instrument Ô Turn the instrument off This chapter contains information on: • Preparing the Cleaning Solution • Shutting Down the Instrument •...

  • Page 266: Auto Shutdown [Cytoflex Lx Only]

    Daily Shutdown Auto Shutdown [CytoFLEX LX Only] Remove the sample tube from the instrument and store according to your laboratory procedures. Exit the software. The instrument automatically enters the standby state. Turn the computer off. Turn the Cytometer's main power switch off. If there are any spills, clean the sample station.

  • Page 267: Chapter 9: Troubleshooting

    IMPORTANT In addition to the information stated, never disassemble the instrument or have it repaired by unauthorized personnel. Beckman Coulter bears no responsibility for any problems arising from the unauthorized repair of the instrument. This chapter introduces solutions to common problems. If there is a problem, follow the information in this chapter to carry out self inspection.

  • Page 268: Laser Beam Hazards

    Laser Beam Hazards The CytoFLEX Series flow cytometer can contain up to 6 solid-state diode lasers that are capable of producing laser light at the following levels: • 375-nm, 60-mW solid-state diode laser • 405-nm, 80-mW solid-state diode laser •...

  • Page 269: Laser Warning Label On The Laser Optical Bench

    Troubleshooting Precautions/Hazards Refer to the following figures for the locations of the CDRH-approved and IEC compliant labels: Figure 9.1 Figure 9.2 for the Laser Warning Label on the Cytometer optical bench. Figure 9.3 Figure 9.4 for the Laser Warning Label on the optical bench (located Inside the Cytometer).

  • Page 270: Laser Warning Label Within The Optical Bench

    Troubleshooting Precautions/Hazards Figure 9.3 Laser Warning Label within the Optical Bench (Located Inside the Cytometer) [CytoFLEX] LASER RADIATION AVOID EXPOSURE TO BEAM CLASS 3B LASER PRODUCT LASER RADIATION AVOID EXPOSURE TO BEAM CLASS 3B LASER PRODUCT CAUTION CLASS 3B LASER RADIATION WHEN OPEN AND INTERLOCKS DEFEATED.

  • Page 271: Laser Warning Labels On The Cytometer

    Troubleshooting Precautions/Hazards Figure 9.5 Laser Warning Labels on the Cytometer Back Cover [CytoFLEX] CLASS 1 LASER PRODUCT COMPLIES WITH 21 CFR 1040.10 AND 1040.11 EXCEPT FOR DEVIATIONS PURSUANT TO LASER NOTICE NO. 50 DATED JUNE 24, 2007 MANUFACTURED CLASS 1 LASER PRODUCT COMPLIES WITH 21 CFR 1040.10 AND 1040.11 EXCEPT FOR DEVIATIONS PURSUANT TO LASER NOTICE NO.

  • Page 272: Hazard Labels And Locations

    Sheath Sheath Waste Figure 9.8 Biohazard Label on the 10 L Fluid Cubitainers 10 L Cyto FLEX Sheath Fluid A nonionic, non-flourescent, and azide free sheath fluid for use on Beckman Coulter CytoFLEX flow cytometers. REACTIVE INGREDIENTS: Anti-microbial compound B49006AH...

  • Page 273: Electrical Shock Hazard Label And Location

    Troubleshooting Hazard Labels and Locations Figure 9.9 Biohazard Label Located in the Sample Station and on the Back of the Cytometer [CytoFLEX Shown] CLASS 1 LASER PRODUCT COMPLIES WITH 21 CFR 1040.10 AND 1040.11 EXCEPT FOR DEVIATIONS PURSUANT TO LASER NOTICE NO. 50 DATED JUNE 24, 2007 MANUFACTURED Electrical Shock Hazard Label and Location Figure 9.10 Electrical Shock Hazard Label by the Power Switch [CytoFLEX Shown]...

  • Page 274: Plate Loader Hazard Labels And Location

    European Union. For products under the requirement of WEEE directive, please contact your dealer or local Beckman Coulter office for the proper decontamination information and take back program which will facilitate the proper collection, treatment, recovery, recycling, and safe disposal of device.

  • Page 275: Rohs Notice

    Troubleshooting RoHS Notice RoHS Notice These labels and materials declaration table (the Table of Hazardous Substance's Name and Concentration) are to meet People's Republic of China Electronic Industry Standard SJ/T11364-2006 "Marking for Control of Pollution Caused by Electronic Information Products" requirements. RoHS Caution Label This label indicates that this electronic information product contains certain toxic or hazardous substances.

  • Page 276: Troubleshooting Table

    Troubleshooting Troubleshooting Table Troubleshooting Table Table 9.1 lists problems that you could encounter while running the CytoFLEX flow cytometer, the probable causes of each problem, and the corrective actions. These problems are listed alphabetically in the Index, under the primary entry “troubleshooting.” Table 9.1 Troubleshooting Problem Probable Cause...
  • Page 277 Troubleshooting Troubleshooting Table Table 9.1 Troubleshooting (Continued) Problem Probable Cause Corrective Action • The alarm is not working. 1. Ensure the sheath fluid/waste harnesses The alarm does not sound when the waste are secured to the correct container. • Instrument data connection container is full or the 2.
  • Page 278 Troubleshooting Troubleshooting Table Table 9.1 Troubleshooting (Continued) Problem Probable Cause Corrective Action • Instrument data connection 1. Ensure the sheath fluid/waste harnesses The fluid status information displays red error. are secured to the correct container. for Sheath and/or Waste • The sensor connection is not 2.
  • Page 279 Troubleshooting Troubleshooting Table Table 9.1 Troubleshooting (Continued) Problem Probable Cause Corrective Action • The threshold setting is too 1. Use the manual threshold setting to The sampling flow rate is too fast. low. increase the threshold. Refer to Adjusting the Threshold CHAPTER 5, Data •...
  • Page 280 Troubleshooting Troubleshooting Table Table 9.1 Troubleshooting (Continued) Problem Probable Cause Corrective Action • Air bubbles are in the flow 1. Run Prime. Refer to Population amplitude is Priming the Flow Cell decreasing and CV cell. CHAPTER 11, Replacement/ values are increasing. Adjustment Procedures.
  • Page 281 Troubleshooting Troubleshooting Table Table 9.1 Troubleshooting (Continued) Problem Probable Cause Corrective Action • The threshold setting is too 1. Decrease the threshold setting. Refer to No data acquisition. high. Adjusting the Threshold CHAPTER 5, Data Acquisition and Sample Analysis. • The gain setting is too low. 2.
  • Page 282 Troubleshooting Troubleshooting Table Table 9.1 Troubleshooting (Continued) Problem Probable Cause Corrective Action • The detector configuration 1. Ensure that the detector configuration is Data populations are not where they are expected. setting is incorrect. set correctly. Refer to Verifying, Selecting, Editing, and Creating Detector •...
  • Page 283 Troubleshooting Troubleshooting Table Table 9.1 Troubleshooting (Continued) Problem Probable Cause Corrective Action • Erroneous data acquisition. 1. Ensure that the corresponding negative The calculation of the automatic compensation control tube and the individual positive • The gate is not set on the experiment is incorrect.
  • Page 284 Troubleshooting Troubleshooting Table Table 9.1 Troubleshooting (Continued) Problem Probable Cause Corrective Action • The sample concentration is 1. Ensure that the pipette used in sample The concentration calculation is incorrect. not within the specified processing is calibrated. range. 2. Verify that the concentration of the •...
  • Page 285 Troubleshooting Troubleshooting Table Table 9.1 Troubleshooting (Continued) Problem Probable Cause Corrective Action • Sample mixing is disabled in 1. Ensure sample mixing is enabled in the The mixer is not functioning. the software. software. Refer to Changing Sample Mixing and Backflush Settings •...
  • Page 286 Troubleshooting Troubleshooting Table Table 9.1 Troubleshooting (Continued) Problem Probable Cause Corrective Action • The diluted CytoFLEX Daily 1. Add 1 drop of CytoFLEX Daily QC QC aborted due to low event rate. QC Fluorosphere or CytoFLEX Fluorospheres to the QC solution. Then, IR Daily QC Fluorosphere rerun QC.
  • Page 287 Troubleshooting Troubleshooting Table Table 9.1 Troubleshooting (Continued) Problem Probable Cause Corrective Action • The median fluorescence 1. Rerun QC. Refer to QC failed. CHAPTER 4, fails to meet the target Instrument Quality Control and specification. Standardization. • The QC gain value does not 2.
  • Page 288 Troubleshooting Troubleshooting Table Table 9.2 Troubleshooting [With Plate Loader] Problem Probable Cause Corrective Action • Incorrect plate type selected. 1. Verify that the correct plate type is The sample probe comes in contact with the selected in the Plate window. •...

  • Page 289: Backup And Restore

    Troubleshooting Backup and Restore Backup and Restore IMPORTANT If you have the Electronic Record Management software option installed, ensure the following prior to attempting to restore data: • The target system includes all of the disk volumes that each experiment directory is saved to. •...
  • Page 290 Troubleshooting Backup and Restore On the Workstation that is being backed up, go to C:\ProgramData\Beckman Coulter\CytExpert\Database\. NOTE If the Program Data folder is not visible: 1. Select the Windows Start menu and search for Folder Options. The Folder Options window appears.

  • Page 291: Restore

    Troubleshooting Backup and Restore 3. Select the Show hidden files, folders, and drives radio button. Copy Right-click on the file called CytExpert.db and select from the pop-up menu. Paste the file in your desired backup location. If you have the 21CFR Tools software option installed, proceed to Step 6. If you have the 21CFR Tools software option installed, go to the location of each experiment directory as noted in Step 1, and copy the folder to the backup location.
  • Page 292 Troubleshooting Backup and Restore Go to C:\ProgramData\Beckman Coulter\CytExpert\Database\ on the system being restored and paste the CytExpert.db file. If you have the 21CFR option installed, proceed to Step 5. If you have the 21CFR Tools software option installed: a. Go to your backup location and copy the user directory's backup folder.

  • Page 293: Chapter 10: Cleaning Procedures

    CHAPTER 10 Cleaning Procedures Overview This chapter describes how to carry out certain routine and nonscheduled cleaning procedures. Proper cleaning can help extend the service life of the instrument and ensure experimental accuracy. When conducting any cleaning, take all necessary biosafety precautions and use proper personal protective equipment.
  • Page 294 Cleaning Procedures Routine Cleaning Daily Clean Select in the Cytometer menu. Add 2 mL of FlowClean solution to an unused sample tube. Add 3 mL of DI water to an unused sample tube. 10-2 B49006AH...
  • Page 295 Cleaning Procedures Routine Cleaning Insert the sample tube with 2 mL of FlowClean solution into the sample holder and select NOTE The default cleaning time is 3 minutes. Remove the Flow Clean tube. 10-3 B49006AH...
  • Page 296 Cleaning Procedures Routine Cleaning Insert the sample tube with 3 mL of DI water into the sample holder and select to perform the second step of the cleaning process. NOTE The default cleaning time is 5 minutes. After the process has been completed, remove the sample tube and close the Daily Clean window.

  • Page 297: Daily Clean [With Plate Loader]

    Cleaning Procedures Routine Cleaning Daily Clean [With Plate Loader] Daily Clean should be performed during instrument startup and instrument shutdown to clean the sample line. After sampling an excessively large sample or a sample that can easily clog the sample probe, it is recommended to perform the Daily Clean procedure.
  • Page 298 Cleaning Procedures Routine Cleaning Follow the on screen software prompts and select the desired wells for cleaning agent and deionized water. [CytoFLEX LX Shown] IMPORTANT You must select at least one cleaning solution well and one water well. a. Select the desired wells for the cleaning agent and select Set As Cleaning Agent Well.

  • Page 299: Cleaning The Sample Station

    Cleaning Procedures Routine Cleaning Cleaning the Sample Station Carry out semi-automatic cleaning for the sample injection device once a week. Ensure that the system has been shut down properly. Refer to Shutting Down the Instrument CHAPTER 8, Daily Shutdown. WARNING Risk of chemical injury from bleach.

  • Page 300: Cleaning The Sample Probe

    Cleaning Procedures Routine Cleaning Cleaning the Sample Probe When problems such as blockage of the sample probe occur, it is required to replace or clean the sample probe. Confirm that the instrument is in the standby state or that the power supply is turned off. WARNING Risk of biohazardous contamination if you have skin contact with the sample probe or the sample peristaltic pump tubing.
  • Page 301 Cleaning Procedures Routine Cleaning Place the instrument in standby state. Remove the right-side cover. Refer to Right-Side Cover Removal and Reinstallation CHAPTER 11, Replacement/Adjustment Procedures. Confirm that the Deep Clean solution volume in the bottle located inside the Fluidics module is sufficient.

  • Page 302: Cleaning The 4 L Sheath Fluid

    Cleaning Procedures Routine Cleaning Allow the cleaning solution to remain in the flow cell for approximately 30 minutes. If you are required to postpone the cleaning time, do not exceed 24 hours. During the Deep Clean cycle, the power supply of the unit can be turned off, but the instrument cannot be initialized. Prime Select in the Cytometer menu.

  • Page 303: Cleaning The 4 L Waste Container

    Cleaning Procedures Routine Cleaning Remove the sheath fluid harness from the sheath fluid container. Empty the residual sheath fluid from the sheath fluid container. Add about 50 to 100 mL of CytoFLEX Sheath Fluid to the sheath fluid container. Insert the sheath fluid harness back into the sheath fluid container and tightly close the sheath fluid container cap.
  • Page 304 Cleaning Procedures Routine Cleaning WARNING Risk of biohazardous contamination if you have skin contact with the waste container, its contents, and its associated tubing. The waste container and its associated tubing could contain residual biological material and must be handled with care.

  • Page 305: Surface Cleaning And Disinfection

    Place the waste container back into the Fluid Container holder. Nonscheduled Cleaning Surface Cleaning and Disinfection Wipe the CytoFLEX Series label of the instrument clean with water and wipe dry immediately. If you have a plate loader installed on your instrument, remove the front cover. Refer to Front...

  • Page 306: Preparing The Instrument For Transport Or

    Cleaning Procedures Nonscheduled Cleaning WARNING Risk of chemical injury from bleach. To avoid contact with the bleach, use barrier protection, including protective eyeware, gloves, and suitable laboratory attire. Refer to the Safety Data sheet for details about chemical exposure before using the chemical.
  • Page 307 Cleaning Procedures Nonscheduled Cleaning Clean the Sample Station. Refer to Cleaning the Sample Station. Empty the sheath fluid container and the waste container (see Emptying the 4 L Waste Container [CytoFLEX] CHAPTER 11, Replacement/Adjustment Procedures). Clean and disinfect all surfaces. Refer to Surface Cleaning and Disinfection.
  • Page 308 Cleaning Procedures Nonscheduled Cleaning CAUTION Risk of instrument damage. The Cytometer can suffer irreparable damage if it is exposed to subfreezing temperatures while it still contains liquid. Always drain the flow cell after cleaning the Cytometer if the Cytometer will be stored or transported in subfreezing temperatures.
  • Page 309 Cleaning Procedures Nonscheduled Cleaning Disconnect the yellow-labeled tubing connected to the liquid damper to vent the flow cell, allowing it to drain. 1. Liquid damper Verify that liquid has stopped dripping from the blue-labeled tubing. NOTE The flow cell is empty when liquid stops dripping from the blue-labeled tubing. Dispose of the absorbent material used to collect the liquid in accordance with the local regulations and acceptable laboratory procedures and cleanup any spills.

  • Page 310: Lifting And Carrying

    Cleaning Procedures Nonscheduled Cleaning If the instrument is to be transported or stored, put the instrument and the Plate Loader module (if applicable) into the Beckman Coulter supplied packing and comply with the requirements in Instrument Transportation and Storage APPENDIX A, Instrument Installation, regarding correct placement during transportation and storage.
  • Page 311 Cleaning Procedures Nonscheduled Cleaning WARNING Risk of personal injury. Use caution when lowering the Cytometer to avoid pinching fingers. Lower the Cytometer to its designated location. 10-19 B49006AH...
  • Page 312 Cleaning Procedures Nonscheduled Cleaning 10-20 B49006AH...

  • Page 313: Replacement/Adjustment Procedures

    Beckman Coulter to ensure proper functioning of the instrument. Never disassemble any part of the instrument without prior authorization. Beckman Coulter assumes no responsibility for any instrument problems resulting from the use of any part not authorized by Beckman Coulter for use with the instrument.

  • Page 314: Replacement/Adjustment

    Replacement/Adjustment Procedures Routine Replacement/Adjustment — Calibrating the Sample Flow Rate [With Plate Loader] — Setting Laser Delay — Replacing the Optical Filter — Replacing the Fuse — Replacing the Sheath Fluid Harness and/or Waste Harness — Changing Sample Mixing and Backflush Settings —...

  • Page 315: Reinstallation

    Replacement/Adjustment Procedures Routine Replacement/Adjustment Remove the two screws securing the front cover and pull the front cover forward. 1. Securing screws 2. Front cover Lift the front cover up and out of the slots in the frame. Reinstallation Slide the tabs on the bottom of the front cover into the slots in the bottom of the frame. Push in the latches on the front cover to retract the pins, push the front cover into place, and release the latches to secure the cover.

  • Page 316: Right-Side Cover Removal

    Replacement/Adjustment Procedures Routine Replacement/Adjustment Right-Side Cover Removal and Reinstallation IMPORTANT The CytoFLEX LX flow cytometer only requires you to unfasten the thumbscrews the first time. There is a retaining clip that is used to secure the right-side cover without the need for the thumbscrews, if desired.

  • Page 317: Reinstallation

    Replacement/Adjustment Procedures Routine Replacement/Adjustment Unfasten the two captive thumbscrews for the right-side cover. 1. Captive thumbscrews 2. Right-side cover 3. Tabs Lift the right-side cover up and out of the slots in the frame. Reinstallation Slide the tabs on the bottom of the right-side cover into the slots in the bottom of the frame and push the cover into place.

  • Page 318: Filling The 4 L Sheath Fluid Container

    Replacement/Adjustment Procedures Routine Replacement/Adjustment Filling the 4 L Sheath Fluid Container [CytoFLEX] Confirm that the instrument is turned off or is in the standby state. If necessary, remove any cardboard cutouts from the new CytoFLEX Sheath Fluid cubitainer. If you do not need a new cubitainer, skip to Step 7. Locate the spigot inside the cardboard cutout.

  • Page 319: Replacing The 10 L Sheath Fluid

    Replacement/Adjustment Procedures Routine Replacement/Adjustment Replacing the 10 L Sheath Fluid Cubitainer Confirm that the instrument is turned off or is in the standby state. If necessary, remove any cardboard cutouts from the new CytoFLEX Sheath Fluid cubitainer. Remove the cap and seal from the new sheath fluid cubitainer. Be sure to completely remove the foil seal.
  • Page 320 Replacement/Adjustment Procedures Routine Replacement/Adjustment CAUTION Misleading results could occur if you contaminate the sheath fluid. Be careful not to contaminate the sheath fluid. Do not let your fingers, paper towels, or other objects touch the pickup tube assembly. Unscrew the plastic cap that secures the pickup tube assembly into the old sheath fluid cubitainer and lay it on a leakproof disposable container, such as a glove or beaker.

  • Page 321: Emptying The 4 L Waste Container

    Replacement/Adjustment Procedures Routine Replacement/Adjustment Carefully insert the pickup tube assembly straight into the new sheath fluid cubitainer. Tighten the cap. Place the 10 L sheath fluid cubitainer to the left of the instrument. Put the cap from the new container onto the old container and dispose of the container properly.

  • Page 322: Emptying The 10 L Waste

    Replacement/Adjustment Procedures Routine Replacement/Adjustment Remove the waste harness (see Figure 1.7). The waste harness from the Cytometer is connected to a 4-L waste container. Remove the waste container from the Fluid Container holder. WARNING Risk of biohazardous contamination if you have skin contact with the waste container, its contents, and its associated tubing.
  • Page 323 Replacement/Adjustment Procedures Routine Replacement/Adjustment Lift the waste cubitainer and swirl it before removing the cap. 11-11 B49006AH...
  • Page 324 Replacement/Adjustment Procedures Routine Replacement/Adjustment WARNING Risk of biohazardous contamination if you have skin contact with the waste cubitainer, its contents, and its associated tubing. The waste cubitainer and it associated tubing might container residual biological material and must be handled with care. Clean up spills immediately. Dispose of the contents of the waste cubitainer in accordance with your local regulations and acceptable laboratory procedures.
  • Page 325 Replacement/Adjustment Procedures Routine Replacement/Adjustment WARNING Risk of chemical injury from bleach. To avoid contact with the bleach, use barrier protection, including protective eyeware, gloves, and suitable laboratory attire. Refer to the Safety Data Sheet for details about chemical exposure before using the chemical.

  • Page 326: Managing The Maintenance

    Replacement/Adjustment Procedures Routine Replacement/Adjustment Managing the Maintenance Reminder The maintenance reminder tracks the last maintenance date and initiates a reminder to complete maintenance for the following three items: • Refill Deep Clean solution bottle (cleaning solution) • Replace sheath fluid filter •...
  • Page 327 Replacement/Adjustment Procedures Routine Replacement/Adjustment Detail Select . The Refill Deep Clean Solution Bottle window appears. Refill Select . A pop-up window appears to reset the maintenance date as the current date that the maintenance was performed. Detail Select . The Replace Sheath Fluid Filter window appears. 11-15 B49006AH...
  • Page 328 Replacement/Adjustment Procedures Routine Replacement/Adjustment Replace Select . A pop-up window appears to reset the maintenance date as the current date that the maintenance was performed. Detail Select . The Replace Peristaltic Pump Tubing window appears. Replace Select . A pop-up window appears to reset the maintenance date as the current date that the maintenance was performed.

  • Page 329: Adding The Deep Clean Solution

    Replacement/Adjustment Procedures Routine Replacement/Adjustment Adding the Deep Clean Solution Check occasionally whether the Deep Clean solution in the Deep Clean solution bottle is sufficient. Replace the Deep Clean solution when the maintenance reminder prompts you to do so. WARNING Risk of chemical injury from Contrad® 70 reagent. To avoid contact with the Contrad®...

  • Page 330: Replacing The Sheath Fluid

    Replacement/Adjustment Procedures Routine Replacement/Adjustment Tighten the cap, and attach the Deep Clean solution bottle to the bracket. Reinstall the right-side cover (see Right-Side Cover Removal and Reinstallation), and fasten the thumbscrews. Reset the maintenance reminder tracker. Refer to Managing the Maintenance Reminder.
  • Page 331 Replacement/Adjustment Procedures Routine Replacement/Adjustment Press the spring piece on the quick connector and the Cytometer on the upper side of the filter, and disconnect the quick connector. Repeat Step for the quick connector behind the quick connector removed in the previous step. NOTE The spring is on the opposite side.
  • Page 332 Replacement/Adjustment Procedures Routine Replacement/Adjustment Attach the filter to the filter bracket. Remove and set aside the vent cap and the rubber insert. Turn on the Cytometer and open the software. Prime Select in the Cytometer menu. 11-20 B49006AH...
  • Page 333 Replacement/Adjustment Procedures Routine Replacement/Adjustment CAUTION Risk of instrument damage. If the vent cap is not sealed tightly, unstable flow rate and leakage of the sheath fluid can occur. Observe the liquid level in the filter during the prime cycle. When the liquid level reaches the upper section of the filter, reinstall the rubber insert, and lock the vent cap to prevent air leakage.
  • Page 334 If you need to replace the single tube sample probe assembly and you have the Sample Injection Mode Control Kit installed on your CytoFLEX Series instrument, pull the probe out of the CytoFLEX Sample Injection Mode Control Kit and push the new probe into place.
  • Page 335 Replacement/Adjustment Procedures Routine Replacement/Adjustment Remove the sample pump cover thumbscrew and the sample pump cover. Take out the sample peristaltic pump tubing. 11-23 B49006AH...
  • Page 336 Replacement/Adjustment Procedures Routine Replacement/Adjustment CAUTION Risk of optical misalignment. Removing the PEEK tubing from the bottom of the flow cell could cause misalignment of the optical components. Do not remove the PEEK tubing from the bottom of the flow cell. Remove the sample PEEK tubing from the sample peristaltic pump tubing.
  • Page 337 Replacement/Adjustment Procedures Routine Replacement/Adjustment Remove the sample peristaltic pump tubing from the sample probe. Dispose of the old sample probe and/or sample peristaltic pump tubing in accordance with your local regulations and acceptable laboratory procedures. Connect the sample peristaltic pump tubing to the sample probe. 11-25 B49006AH...
  • Page 338 Replacement/Adjustment Procedures Routine Replacement/Adjustment Connect the sample PEEK tubing to the sample peristaltic pump tubing. Slide the sample probe into the wash station. 11-26 B49006AH...
  • Page 339 Replacement/Adjustment Procedures Routine Replacement/Adjustment Install the sample peristaltic pump tubing, taking care not to use any sharp tools, ensuring that the tube is fully inserted into the groove. NOTE You can use the sample pump cover thumbscrew to install the sample peristaltic pump tubing. Install the sample pump cover.

  • Page 340: Replacing The Sample Probe Assembly

    Replacement/Adjustment Procedures Routine Replacement/Adjustment Replacing the Sample Probe Assembly [With Plate Loader] IMPORTANT If you have the Sample Injection Mode Control Kit installed on your CytoFLEX Series instrument, contact us to replace the sample probe assembly. WARNING Risk of biohazardous contamination if you have skin contact with the sample probe or the sample peristaltic pump tubing.
  • Page 341 Replacement/Adjustment Procedures Routine Replacement/Adjustment CAUTION Risk of optical misalignment. Removing the PEEK tubing from the bottom of the flow cell could cause misalignment of the optical components. Do not remove the PEEK tubing from the bottom of the flow cell. Remove the plate loader PEEK tubing from the sample peristaltic pump tubing.
  • Page 342 Replacement/Adjustment Procedures Routine Replacement/Adjustment CAUTION The plate loader PEEK tubing can be deformed, which could affect sample flow. When routing the plate loader PEEK tubing to and from the Sample Station, be careful not to pinch, crimp, stretch, or break the tubing. Pull the plate loader PEEK tubing through the slot so the tubing sits inside the instrument.
  • Page 343 Replacement/Adjustment Procedures Routine Replacement/Adjustment Lift the plate loader sample probe assembly straight up to remove it from the probe holder. Align the tongue on the plate loader sample probe assembly with the groove in the probe holder and slide the probe down until it is flush with the probe holder. 11-31 B49006AH...
  • Page 344 Replacement/Adjustment Procedures Routine Replacement/Adjustment Tighten the white plastic thumbscrew. CAUTION The plate loader PEEK tubing can be deformed which could affect sample flow. When routing the plate loader PEEK tubing to and from the Sample Station, be careful not to pinch, crimp, stretch, or break the tubing. Slide the new plate loader PEEK tubing through the slot into the single tube sample station area.
  • Page 345 Replacement/Adjustment Procedures Routine Replacement/Adjustment Connect the sample peristaltic pump tubing to the plate loader PEEK tubing. Install the sample pump cover. NOTE To ensure the proper placement of the sample probe, push the sample pump cover up while installing the thumbscrew. Reinstall the right-side cover (refer to Right-Side Cover Removal and Reinstallation CHAPTER 11, Replacement/Adjustment...
  • Page 346 Clean up spills immediately. NOTE If you have the Sample Injection Mode Control Kit installed on your CytoFLEX Series instrument, refer Switching the Sample Probe from the Single Tube Sample Station to the Plate Loader [With Sample...

  • Page 347: Tube Sample Station

    Replacement/Adjustment Procedures Routine Replacement/Adjustment Remove the sample pump cover thumbscrew and the sample pump cover. Remove the sample probe from the single tube sample station. 11-35 B49006AH...
  • Page 348 Replacement/Adjustment Procedures Routine Replacement/Adjustment Place the sample probe in the white clip located on the right side of the sample station. 11-36 B49006AH...
  • Page 349 Replacement/Adjustment Procedures Routine Replacement/Adjustment CAUTION The plate loader PEEK tubing can be deformed which could affect sample flow. When removing the plate loader PEEK tubing from the white clips, be careful not to pinch, crimp, stretch, or break the tubing. Remove the plate loader PEEK tubing from the two white clips located at the top of the sample station.
  • Page 350 Replacement/Adjustment Procedures Routine Replacement/Adjustment Connect the plate loader PEEK tubing to the sample peristaltic pump tubing. Reinstall the sample pump cover. NOTE To ensure the proper placement of the sample probe, push the sample pump cover up while installing the thumbscrew. Reinstall the right-side cover (refer to Right-Side Cover Removal and Reinstallation CHAPTER 11, Replacement/Adjustment...
  • Page 351 Clean up spills immediately. NOTE If you have the Sample Injection Mode Control Kit installed on your CytoFLEX Series instrument, refer Switching the Sample Probe from the Plate Loader to the Single Tube Sample Station [With Sample...
  • Page 352 Replacement/Adjustment Procedures Routine Replacement/Adjustment Remove the sample pump cover thumbscrew and the sample pump cover. Remove the plate loader PEEK tubing from the sample peristaltic pump tubing. 11-40 B49006AH...
  • Page 353 Replacement/Adjustment Procedures Routine Replacement/Adjustment CAUTION The plate loader PEEK tubing can be deformed, which could affect sample flow. When placing the plate loader PEEK tubing in the white clips, be careful not to pinch, crimp, stretch, or break the tubing. Place the plate loader PEEK tubing in the two white clips located at the top of the sample station.

  • Page 354: Inspecting The Liquid Flow Path For Priming The Flow Cell

    Replacement/Adjustment Procedures Routine Replacement/Adjustment Connect the sample probe to the sample peristaltic pump tubing. Daily Clean CHAPTER 10, Cleaning Procedures. Inspecting the Liquid Flow Path for Leaks WARNING The liquid flow tubing can aged and cracked or the connector can be loosened. Liquid leakage can lead to biological harm.
  • Page 355 Replacement/Adjustment Procedures Routine Replacement/Adjustment Check the connectors and tubes in the Fluidics module, and check whether any liquid leaks out. Check the sheath fluid, backflush, and waste liquid connector on the back of the Cytometer, and check whether any liquid leaks out. Place the instrument in standby state, complete the priming procedure, and check whether the Fluidics module has any liquid leakage.
  • Page 356 Replacement/Adjustment Procedures Routine Replacement/Adjustment Prime Select from the Cytometer Menu to prime the flow cell. Wait for the beep and for the Instructions window to close. Otherwise, look for the status bar to display that priming has completed. Run Daily Clean to clean the sample line. Refer to Daily Clean CHAPTER 10, Cleaning Procedures...

  • Page 357: Replacing The Plate Holder

    Replacement/Adjustment Procedures Routine Replacement/Adjustment Replacing the Plate Holder [With Plate Loader] Figure 11.1 Plate Holder Stage 1. Pins 2. Position A1 CAUTION The plate holder must be secured tightly to the plate holder stage with position A1 located in the top, left corner of the plate holder stage (refer to Figure 11.1) to prevent instrument damage.

  • Page 358: Plate Loader Module Removal And Reinstallation

    Replacement/Adjustment Procedures Routine Replacement/Adjustment Plate Loader Module Removal and Reinstallation [With Plate Loader] WARNING Risk of contamination from biohazardous material. Always wear PPE when performing this procedure as you can contact components with blood residue. Removal Power down the instrument and disconnect the power cable from the wall. Lift the top cover and remove the front cover.
  • Page 359 Replacement/Adjustment Procedures Routine Replacement/Adjustment In the Sample Station, disconnect that end of the plate loader PEEK tubing. • If the plate loader PEEK tubing is attached to the sample peristaltic pump tubing, remove the sample pump cover and disconnect the plate loader PEEK tubing. 11-47 B49006AH...
  • Page 360 Replacement/Adjustment Procedures Routine Replacement/Adjustment • If the plate loader PEEK tubing is in the plate loader PEEK tubing clips, remove the tubing from the clips. CAUTION The PEEK tubing can be deformed which could affect sample flow. When routing the Plate Loader PEEK tubing to and from the Sampling Station, be careful not to pinch, crimp, stretch, or break the tubing.

  • Page 361: Removing The Tubings From The Fluidics Module To The Plate Loader

    Replacement/Adjustment Procedures Routine Replacement/Adjustment Remove the red and black marked tubes that are attached to the left and right connectors in the Plate Loader module, respectively, as shown in Figure 11.2. Figure 11.2 Removing the Tubings from the Fluidics Module to the Plate Loader Screw 1.

  • Page 362: Installation

    Replacement/Adjustment Procedures Routine Replacement/Adjustment Remove the two countersunk M4 x 10 Phillips-head screws in the Plate Loader module. Refer to Figure 11.3. Figure 11.3 Plate Loader Module Securing Screws 1. Securing screws Slide the Plate Loader module out of the Cytometer. Refer to Figure 11.4.

  • Page 363: Plate Loader Module Securing

    Replacement/Adjustment Procedures Routine Replacement/Adjustment Remove the right-side cover. Refer to Right-Side Cover Removal and Reinstallation. Slide the Plate Loader module in to the Cytometer. Refer to Figure 11.5. Figure 11.5 Installing the Plate Loader in to the Cytometer Install the two countersunk M4 x 10 Phillips-head screws in the Plate Loader module. Refer to Figure 11.6.

  • Page 364: Connecting The Tubings From The Fluidics Module To The Plate Loader

    Replacement/Adjustment Procedures Routine Replacement/Adjustment Install the red and black marked tubings that are attached to the left and right connectors in the Plate Loader module, respectively, as shown in Figure 11.7. Figure 11.7 Connecting the Tubings from the Fluidics Module to the Plate Loader Screw 1.
  • Page 365 Replacement/Adjustment Procedures Routine Replacement/Adjustment CAUTION The PEEK tubing can be deformed, which could affect sample flow. When routing the Plate Loader PEEK tubing to and from the Sampling Station, be careful not to pinch, crimp, stretch, or break the tubing. Slide the new plate loader PEEK tubing through the slot into the single tube sample station area.
  • Page 366 Replacement/Adjustment Procedures Routine Replacement/Adjustment Connect the plate loader PEEK tubing to the sample peristaltic pump tubing. Reinstall the sample pump cover. NOTE To ensure the proper placement of the sample probe, push the sample pump cover up while installing the thumbscrew. Replace the right-side cover (refer to Right-Side Cover Removal and Reinstallation CHAPTER 11, Replacement/Adjustment...

  • Page 367: Changing The Event Rate Setting

    Replacement/Adjustment Procedures Routine Replacement/Adjustment Close the top cover. Turn the Cytometer's main power switch on. Verify that the system is operating correctly by running QC in the Plate Loader mode. Refer to Preparing the QC Sample [With Plate Loader] Collecting QC Data [With Plate Loader] CHAPTER 4, Instrument Quality Control and Standardization.

  • Page 368: Replacement/Adjustment

    Replacement/Adjustment Procedures Nonscheduled Replacement/Adjustment Nonscheduled Replacement/Adjustment Calibrating the Sample Flow Rate Calibrate the sample flow rate: • After replacing the sample peristaltic pump tubing. • If a precise volumetric measurement is required. The accuracy of concentration calculations can be affected by the sample flow rate. Verify that the instrument is in the initialized state.
  • Page 369 Replacement/Adjustment Procedures Nonscheduled Replacement/Adjustment Use a calibrated electric balance to measure the weight of one sample tube containing at least 1 mL clean deionized water. Record the data and enter it into the software. Next Select and put the sample tube in the sample loading position (see Figure 1.12).
  • Page 370 Replacement/Adjustment Procedures Nonscheduled Replacement/Adjustment Select to begin acquiring the sample. Wait for the sample run to finish, remove the sample tube, and use the electronic balance to measure the weight and record the value. 11-58 B49006AH...

  • Page 371: Calibrating The Sample Flow

    Replacement/Adjustment Procedures Nonscheduled Replacement/Adjustment Next Select to determine whether the current flow rate is required to be calibrated. According to need, select whether any other flow rate is required to be calibrated. If no calibration is required, the current setting is kept. If deviation occurs, the setting is automatically corrected.
  • Page 372 Replacement/Adjustment Procedures Nonscheduled Replacement/Adjustment Calibrate Sample Flow Rate Select in the Cytometer menu. The plate loader automatically ejects the plate holder stage. Calibrate Select the sampling speed that needs to be calibrated and select . The Calibrate Sample Flow Rate window appears. Follow the onscreen software prompts, then weigh the sample plate.
  • Page 373 Replacement/Adjustment Procedures Nonscheduled Replacement/Adjustment Enter the weight of the sample plate and set the acquisition time. NOTE Do not exceed three minutes when using the fast sampling rate. Select Plate Settings to set the sample well and plate type. Select to save the settings.
  • Page 374 Replacement/Adjustment Procedures Nonscheduled Replacement/Adjustment Next Select to proceed to the next step. The plate loader automatically ejects the plate holder stage. Place the sample plate onto the plate loader. Verify the settings and select Load to load the plate. Select .

  • Page 375: Setting Laser Delay

    Replacement/Adjustment Procedures Nonscheduled Replacement/Adjustment Next Enter the remaining weight value and select to confirm the calibration. Select Setting Laser Delay Laser delay is preset for QC. Only change the laser delay if the software prompts you that there is a difference in the actual delay and the default delay.

  • Page 376: Replacing The Optical Filter

    For the specific part number of the optical filter, consult your Beckman Coulter Representative or your local dealer. Confirm that the instrument is in the standby state or that the instrument is turned off.
  • Page 377 Replacement/Adjustment Procedures Nonscheduled Replacement/Adjustment Press the spring piece of the WDM cap corresponding to the laser, and open the WDM cap. 11-65 B49006AH...
  • Page 378 Replacement/Adjustment Procedures Nonscheduled Replacement/Adjustment CAUTION Risk of damage to the optical filter. Do not touch the optical filter glass piece. Touching the optical filter glass piece can obscure and/or scratch the optical filter glass piece. CAUTION Risk of damage to the optical filter. Pull the filter straight up when removing it from the WDM.

  • Page 379: Replacing The Fuse

    Replacement/Adjustment Procedures Nonscheduled Replacement/Adjustment Close the WDM cap and the instrument top cover. Turn on the Cytometer and open the software. Detector Configuration Select in the Cytometer menu and create a new instrument configuration based on the settings of the new optical filter. Refer to Verifying, Selecting, Editing, and Creating Detector Configuration CHAPTER 5, Data Acquisition and Sample...
  • Page 380 Check whether the fuse installed is blown, and replace the blown fuse with a new one. The specifications of the fuse required are: T 5 AL 250 VAC, delay blow fuse, 5A, 250 VAC, 5 x 20 mm. Beckman Coulter recommends using SCHURTER 0034.3124. Insert the fuse holder.

  • Page 381: Replacing The Sheath Fluid Harness And/Or Waste

    Replacement/Adjustment Procedures Nonscheduled Replacement/Adjustment Reconnect the power cable. Replacing the Sheath Fluid Harness and/or Waste Harness Replace the sheath fluid harness and/or the waste harness if you have a faulty sheath fluid sensor and/or waste sensor. Confirm that the instrument is turned off or is in the standby state. Remove sheath and/or waste pickup tubing from the appropriate container.
  • Page 382 Replacement/Adjustment Procedures Nonscheduled Replacement/Adjustment Disconnect the waste and/or sheath level sensors. Dispose of the sheath fluid harness and/or the waste harness according to your laboratory procedures. Insert the new sheath and/or waste pickup tubing into the appropriate container. Connect the blue harness from the sheath fluid container and/or the yellow harness from the waste container...

  • Page 383: Changing Sample Mixing And Backflush Settings

    Replacement/Adjustment Procedures Nonscheduled Replacement/Adjustment Connect the waste and/or sheath level sensors. Changing Sample Mixing and Backflush Settings WARNING Risk of biohazardous contamination. Enabling sample mixing for 1.5-mL and 2-mL sample tubes in the semi-automatic sample injection mode can result in sample splashing.
  • Page 384 Replacement/Adjustment Procedures Nonscheduled Replacement/Adjustment Cytometer Configuration Select in the Cytometer menu. The Cytometer Configuration window appears. Select the Sample Mixing checkbox to enable sample mixing. Deselect the Sample Mixing checkbox to disable sample mixing. Change the sample mixing duration to the desired time. NOTE The default setting is 1 second.

  • Page 385: Calibrating The Plate Position [With Plate Loader]

    Replacement/Adjustment Procedures Nonscheduled Replacement/Adjustment Select Calibrating the Plate Position [With Plate Loader] Use the following procedure to calibrate the plate position and the sample probe position: • Upon installation • After a new plate type is defined for first use •...
  • Page 386 Replacement/Adjustment Procedures Nonscheduled Replacement/Adjustment Select the plate type and place the plate on the plate holder. NOTE Ensure plate well A1 aligns with position A1 on the plate holder. Calibrate Select . The message Are you sure the plate is loaded? appears. Select .
  • Page 387 Replacement/Adjustment Procedures Nonscheduled Replacement/Adjustment CAUTION Risk of instrument damage. Do not crash the probe into the bottom of the plate as this will cause irreparable damage to the probe. Move the probe one step at a time when lowering the probe on the Z-axis toward the bottom of the plate. You will hear a click when the probe makes contact with the bottom of the well, this is the correct position for the Z-axis of the probe.
  • Page 388 Replacement/Adjustment Procedures Nonscheduled Replacement/Adjustment Next Select to move to the next well. Repeat Steps for wells A12, H1, and H12. 11-76 B49006AH...
  • Page 389 Replacement/Adjustment Procedures Nonscheduled Replacement/Adjustment Select additional plate types to be calibrated on the Calibration completed window, if necessary. Place the corresponding plate on the plate holder and repeat Steps 5-8. Select Finish to save the settings. 11-77 B49006AH...
  • Page 390 Replacement/Adjustment Procedures Nonscheduled Replacement/Adjustment Close Select to quit the program. Reinstall the front cover. Refer to Front Cover Removal and Reinstallation CHAPTER 11, Replacement/Adjustment Procedures. 11-78 B49006AH...

  • Page 391: Appendix A: Instrument Installation

    Ô Ô environment instrument [CytoFLEX LX]: The CytoFLEX LX is installed by your Beckman Coulter Service Representative. Do not open the box or crate. Wait for a qualified Beckman Coulter Service Representative. This chapter contains information on: • Instrument Transportation and Storage •...

  • Page 392: Installation Environment Validation

    Instrument Installation Installation Environment Validation Installation Environment Validation IMPORTANT This instrument is intended for indoor use only. Verify whether the installation environment satisfies the following requirements: Worktable CAUTION Risk of instrument damage. Place the instrument on a level surface. Failing to do so places the system is in danger of toppling and can result in damage.

  • Page 393: Power Source

    Verify that the output voltage of the power outlet conforms to the system requirements and that a 5 A, time delay, T 5 AL 250 VAC fuse is installed. To prevent personal injury, Beckman Coulter recommends using a power source designed to protect against electrical shock.

  • Page 394: Temperature And Humidity

    Instrument Installation Unpacking the Instrument and Inspecting the Materials for Defects or Omissions [CytoFLEX] Temperature and Humidity CAUTION Risk of instrument damage and/or erroneous results. To ensure reliability, the system must be operated in the specified environment, within the required temperature and humidity ranges.

  • Page 395: Installing The Instrument And Connecting The Equipment

    Instrument Installation Unpacking the Instrument and Inspecting the Materials for Defects or Omissions [CytoFLEX] • Mouse • Keyboard • Monitor • Fluid Container holder [CytoFLEX] • Sheath fluid container • Waste container • Sheath fluid tubing • Waste tubing • USB configuration key •...
  • Page 396 Instrument Installation Unpacking the Instrument and Inspecting the Materials for Defects or Omissions [CytoFLEX] After removing the monitor and attaching the base, place them on the worktable with the computer. [CytoFLEX Without Plate Loader Shown] 1. Fluid Containers. Place on the left side of the Cytometer. 2.
  • Page 397 Instrument Installation Unpacking the Instrument and Inspecting the Materials for Defects or Omissions [CytoFLEX] Remove any shipping tape and open the cap of each WDM. Verify that the light filters inside are in place. Insert sheath and waste pickup tubing into the appropriate container. Connect the blue harness from the sheath fluid container and the yellow harness...
  • Page 398 Instrument Installation Unpacking the Instrument and Inspecting the Materials for Defects or Omissions [CytoFLEX] Connect the waste and sheath level sensors. Place the sheath fluid container and the waste container in the Fluid Container holder Sheath Sheath Waste B49006AH...
  • Page 399 Instrument Installation Unpacking the Instrument and Inspecting the Materials for Defects or Omissions [CytoFLEX] Set up the supplied computer and connect the USB cable from the back of the Cytometer to a USB port on the back of the computer. CLASS 1 LASER PRODUCT COMPLIES WITH 21 CFR 1040.10 AND 1040.11 EXCEPT FOR DEVIATIONS PURSUANT TO...
  • Page 400 Instrument Installation Unpacking the Instrument and Inspecting the Materials for Defects or Omissions [CytoFLEX] Remove the foam support holding the sample loader in the front-right corner of the instrument. [CytoFLEX Without Plate Loader Shown] Add Deep Clean solution to the Deep Clean solution bottle. Refer to Adding the Deep Clean Solution CHAPTER 11, Replacement/Adjustment...

  • Page 401: Installing The Instrument And Connecting The Equipment

    Within five business days, activate your Warranty by contacting us and providing them with your latest QC run results. Installing the Instrument and Connecting the Equipment [CytoFLEX LX] The CytoFLEX LX is installed by Beckman Coulter service. A-11 B49006AH...

  • Page 402: Cytexpert Software Installation Options

    Instrument Installation CytExpert Software Installation Options CytExpert Software Installation Options CytExpert software version 2.0 and higher has three installation options upon install. • CytExpert Default software option. User Login is not required to run the system. • CytExpert User Management software option. User Login is required to run the system. Contains features and functionality that facilitates user and role management.

  • Page 403: Required Materials

    The following materials are required to install the CytExpert software: • CytoFLEX Series flow cytometer. • CytoFLEX Series Workstation. • CytExpert software installation USB. • Authorized Beckman Coulter USB configuration key. Installing the CytExpert Software IMPORTANT Follow this procedure when installing the CytExpert software for the first time.
  • Page 404 Instrument Installation Installing the Software [CytoFLEX] Select . The CytExpert Setup Welcome window appears. Next Select Select both support program checkboxes in the CytExpert Setup Prerequisites window. A-14 B49006AH...
  • Page 405 Instrument Installation Installing the Software [CytoFLEX] Next Select . The Microsoft .NET Framework window appears. Select the I have read and accept the license terms checkbox. Install Select . The Installation is Complete window appears when installation has finished. A-15 B49006AH...
  • Page 406 Instrument Installation Installing the Software [CytoFLEX] Finish Select . The Microsoft Visual C++ 2010 Redistributable Setup window appears. Select the I have read and accept the license terms checkbox. Install Select . The Setup Successful window appears when installation has finished. A-16 B49006AH...
  • Page 407 Instrument Installation Installing the Software [CytoFLEX] Close Select . The Welcome to the CytExpert Setup Wizard window appears. Select Next . The Beckman Coulter License Terms window appears. Read the Beckman Coulter Customer End User License Agreement. A-17 B49006AH...
  • Page 408 Instrument Installation Installing the Software [CytoFLEX] Select the I accept the terms of this agreement checkbox. NOTE The checkbox is not selectable until you scroll all the way to the end of the agreement. Select Next . The Choose a file location window appears. Select Next .
  • Page 409 Instrument Installation Installing the Software [CytoFLEX] Select the desired installation option. Refer to CytExpert Software Installation Options. Select Next . The Begin installation of CytExpert window appears. Install Select to begin installing the software. The Installation Progress window appears. NOTE The software will install into the default file path provided unless otherwise specified.
  • Page 410 Instrument Installation Installing the Software [CytoFLEX] If a Windows Security window appears. Select Install this driver software anyway to install the USB drive. The following software prompt appears. Select NOTE The term “device” in this message refers to the cytometer. Wait for the software to finish installing.

  • Page 411: Installing The Instrument Configuration File

    Instrument Installation Installing the Software [CytoFLEX] Close Select to finish the CytExpert software installation. Install the instrument configuration file. Refer to Installing the Instrument Configuration File. Installing the Instrument Configuration File CAUTION Risk of erroneous results or instrument damage. Only install the configuration file that matches your instrument.
  • Page 412 Instrument Installation Installing the Software [CytoFLEX] Select . The CytExpert Config Welcome window appears. Next Select . The CytExpert Config Ready to Install window appears. Verify that the serial name displayed at the top of the window is correct. A-22 B49006AH...
  • Page 413 Instrument Installation Installing the Software [CytoFLEX] Install Select . When the installation has finished, the Completing the CytExpert Config Setup Wizard window appears. Finish Select If you get the following message, select . When you launch CytExpert, the system will automatically upgrade your configuration file.

  • Page 414: Starting The Software

    Instrument Installation Upgrading the CytExpert Software Launch CytExpert. A System Warning message appears. Select to update the configuration file. Starting the Software IMPORTANT The default username is admin. The default password is password. Insert the USB configuration key into the USB port of the computer. Start the software.
  • Page 415 Instrument Installation Upgrading the CytExpert Software CAUTION Risk of data loss. Reinstalling the CytExpert Software could overwrite your database. Ensure you backup your database prior to software reinstallation. Insert the software USB into the computer. NOTE If the Autoplay window appears, select Open folder to view files. CytExpert_X.X_Setup.exe Select .
  • Page 416 Instrument Installation Upgrading the CytExpert Software Select . The Welcome to CytExpert Setup Wizard window appears. Select Next . The Beckman Coulter License Terms window appears. Read the Beckman Coulter Customer End User License Agreement. A-26 B49006AH...
  • Page 417 Instrument Installation Upgrading the CytExpert Software Select the I accept the terms of this agreement checkbox. NOTE The checkbox is not selectable until you scroll all the way to the end of the agreement. Select Next . The Installation Options window appears. A-27 B49006AH...
  • Page 418 Instrument Installation Upgrading the CytExpert Software Next Select . If you are upgrading your software from a previous version and the previous version is not uninstalled first, the following window appears. Select Install to begin installing the software. The Installation Progress window appears. A-28 B49006AH...

  • Page 419: Reinstalling The Cytexpert Software

    Instrument Installation Reinstalling the CytExpert Software If the following software prompt appears, select NOTE The term “device” in this message refers to the cytometer. Wait for the software to finish installing. The install complete window appears. Select Close to finish the CytExpert software installation. Reinstalling the CytExpert Software Use this procedure to: •...
  • Page 420 Instrument Installation Reinstalling the CytExpert Software • Reinstall the same version of software. • Upgrade your software to a version newer than software version 2.0. CAUTION Risk of data loss. Reinstalling the CytExpert Software could overwrite your database. Ensure you backup your database prior to software reinstallation. Backup your CytExpert data if you previously had a software option other than the Default software option installed.
  • Page 421 Instrument Installation Reinstalling the CytExpert Software Select . The Welcome to CytExpert Setup Wizard window appears. Select Next . The Change your installation of CytExpert screen appears. A-31 B49006AH...
  • Page 422 Instrument Installation Reinstalling the CytExpert Software Remove Select . The Begin Remove of CytExpert screen appears. IMPORTANT If the Remove CytExpert settings, CytExpert database, cytometer configuration, and temporary files checkbox is checked, your settings, database, configuration files, and temporary files will be overwritten and you will need to reinstall your cytometer configuration and restore any databases you might have.
  • Page 423 Instrument Installation Reinstalling the CytExpert Software Remove Select . When software removal is complete, the software displays the message CytExpert has been successfully configured. Close Select Navigate back to the software USB folder. CytExpert_X.X_Setup.exe Select . The User Account Control window appears. A-33 B49006AH...
  • Page 424 Instrument Installation Reinstalling the CytExpert Software Select . The Welcome to CytExpert Setup Wizard window appears. Select Next . The Beckman Coulter License Terms window appears. Read the Beckman Coulter Customer End User License Agreement. A-34 B49006AH...
  • Page 425 Instrument Installation Reinstalling the CytExpert Software Select the I accept the terms of this agreement checkbox. NOTE The checkbox is not selectable until you scroll all the way to the end of the agreement. Select Next . The Choose a file location window appears. Select Next .
  • Page 426 Instrument Installation Reinstalling the CytExpert Software Select the desired installation option. Refer to CytExpert Software Installation Options. Select Next . The Begin installation of CytExpert window appears. Install Select to begin installing the software. The Installation Progress window appears. A-36 B49006AH...
  • Page 427 Instrument Installation Reinstalling the CytExpert Software If the following software prompt appears, select NOTE The term “device” in this message refers to the cytometer. Wait for the software to finish installing. The install complete window appears. Select Close to finish the CytExpert software installation. A-37 B49006AH...
  • Page 428 Instrument Installation Reinstalling the CytExpert Software A-38 B49006AH...

  • Page 429: Appendix B: Cytexpert Electronic Record Management

    APPENDIX A, Instrument Installation. Beckman Coulter's CytoFLEX with CytExpert software version 2.0 and higher contains features and functionality that facilitates compliance with 21 CFR Part 11 guidelines for Electronic Records and Signatures. This electronic record management includes controls for user identification, permissions, electronic signatures, data integrity, operation and experiment logs and audit trails.

  • Page 430: Experiment Management

    CytExpert Electronic Record Management Experiment Management Experiment Management CAUTION Risk of file corruption. Do not add, delete, or modify data from the Windows Explorer directory. Manage all data changes using Experiment Explorer to ensure file indexing remains intact. Closed File System The closed file system provides audit trail capability for CytExpert experiment files.

  • Page 431: Experiment Directory Management

    CytExpert Electronic Record Management Experiment Management Experiment Directory Management When launching CytExpert for the first time after installing the software, the Administrator must setup at least one experiment directory for experiment files. NOTE Only one experiment directory can be created per disk. Setting Up the Experiment Directory Settings >...

  • Page 432: Folder Hierarchy Management

    CytExpert Electronic Record Management Experiment Management Select . The specified folder appears in the Set Experiment Directory window. NOTE Select Rename to rename the experiment directory. Select Delete to delete an experiment directory. Folder Hierarchy Management File > Experiment Explorer New Folder Select to view Experiment Explorer.

  • Page 433: Experiment Related Operations

    CytExpert Electronic Record Management Experiment Management Experiment Related Operations The Experiment Explorer dialog appears in place of the Windows File Explorer in the following operations: New/Open Experiment, New/Open Compensation Experiment, Save As, Save Experiment As Template, Recent Template, and New Experiment from Template. B49006AH...

  • Page 434: Importing An Experiment/Template

    CytExpert Electronic Record Management Experiment Management Importing an Experiment/Template Use the following procedure to import experiment (.xit), compensation (.xitc), or experiment from template (.xitm) files into the system. Select File > Experiment Explorer . The Experiment Explorer window appears. Import Experiment from the right-click drop down menu or from the Experiment Explorer Select toolbar.
  • Page 435 CytExpert Electronic Record Management Experiment Management Open Browse to the desired file path to import and select A progress bar appears when importing files. B49006AH...

  • Page 436: Exporting An Experiment/Template

    CytExpert Electronic Record Management Experiment Management Once the import is complete, the imported files display in the Experiment Explorer. NOTE Users can import .xit, .xitc, and .xitm files individually or multiple files at a time. Exporting an Experiment/Template Select File > Experiment Explorer .
  • Page 437 CytExpert Electronic Record Management Experiment Management To export a single experiment: a. Select Export Experiment from the right-click drop down menu or from the Experiment Explorer toolbar. b. Browse to the desired file path for export and select Save A progress bar appears when exporting files. Once the export is complete, the exported file displays in the target folder.
  • Page 438 CytExpert Electronic Record Management Experiment Management To export multiple experiments: a. Select all of the experiments to be exported then select Export Experiment from the right-click drop down menu or from the Experiment Explorer toolbar. b. Browse to the desired file path for export and select B-10 B49006AH...

  • Page 439: Log

    CytExpert Electronic Record Management A progress bar appears when exporting files. Once the export is complete, the exported file displays in the target folder. The CytExpert Electronic Record Management software option includes three logs: Experiment Operation Log — The experiment operation log lists the audit trail records related to the experiment operations based on query criteria.
  • Page 440 CytExpert Electronic Record Management Figure B.1 Experiment Operation Log Window 1. Experiment: Used to specify the experiment file 6. Query display: Displays the queried results. criteria. : Displays the print and export NOTE preview dialog. Refer to Figure B.3. If an experiment is open, the current experiment displays in the Experiment : Selects an experiment from the section.
  • Page 441 CytExpert Electronic Record Management Figure B.2 Select Experiment Profile Window 1. Keyword search: Used to search for keywords in the experiment display list. 2. Clear: Clears the keyword search. 3. Experiment display list: Displays the experiments in a list. 4. Select: Selects an experiment. 5.

  • Page 442: System Operation Log

    CytExpert Electronic Record Management Figure B.3 Print and Export Preview Window 1. Print: Prints the report. 2. Export to PDF: Exports the report as a PDF. 3. Report View: Displays the detailed report view. System Operation Log The System Operation Log generates system log records for settings, configuration, maintenance, and QC.

  • Page 443: User Management Operation Log

    CytExpert Electronic Record Management Figure B.4 System Operation Log Window NOTE The System Operation Log uses the same query criteria and functions as the Experiment Operation Log. Refer to Figure B.1. User Management Operation Log The User Management Operation Log generates audit log records for user management, role management, account policies, password changes, and login/logout records.

  • Page 444: Electronic Signature

    CytExpert Electronic Record Management Electronic Signature Figure B.5 User Management Operation Log Window NOTE The User Management Operation Log uses the same query criteria and functions as the Experiment Operation Log. Refer to Figure B.1. Electronic Signature Signing an Experiment Select Signature >...

  • Page 445: Retracting An Experiment Signature

    CytExpert Electronic Record Management Electronic Signature Enter your password. Select The signature status displays in the top left corner of the main UI. Select Signature > Electronic Signature Details to view the electronic signature details. Retracting an Experiment Signature Signature > Electronic Signature Details Select to view the electronic signature details.

  • Page 446: User Management

    CytExpert Electronic Record Management User Management Select . The following message appears: Are you sure you want to remove the electronic signature? Select to confirm the retraction of your signature or to cancel. The following message appears after editing and saving a signed experiment: Saving the experiment will retract the electronic signature of this experiment, continue? Select to remove the signature or...
  • Page 447 APPENDIX C Sample Injection Mode Control Kit Overview The Sample Injection Mode Control Kit is a mechanical knob installed by your service engineer that enables users to switch between the Plate Loader sample injection mode and the Semi-Automatic or manual sample injection mode. The switch eliminates the need to manually re-route the tubing. This chapter contains information on: •...

  • Page 448: Appendix C: Sample Injection Mode Control Kit

    Sample Injection Mode Control Kit Sample Injection Mode Control Kit Components Sample Injection Mode Control Kit Components Figure C.1 Sample Injection Mode Control Knob 1. Switch module knob NOTE The P indicates the instrument is set to Plate Loader sample injection mode. The T indicates the instrument is set to the Semi-Automatic or manual sample injection mode.
  • Page 449 Sample Injection Mode Control Kit Switching the Sample Probe from the Single Tube Sample Station to the Plate Loader [With Sample Injection Mode Control Knob] CAUTION Risk of instrument damage. The sample probe can easily become damaged or deformed. To avoid damage to the sample probe, turn the switch module knob carefully and avoid contacting the sample probe.
  • Page 450 Sample Injection Mode Control Kit Switching the Sample Probe from the Plate Loader to the Single Tube Sample Station [With Sample Injection Mode Control Knob] Switching the Sample Probe from the Plate Loader to the Single Tube Sample Station [With Sample Injection Mode Control Knob] WARNING Risk of biohazardous contamination if you have skin contact with the sample probe or the sample peristaltic pump tubing.
  • Page 451 Sample Injection Mode Control Kit Switching the Sample Probe from the Plate Loader to the Single Tube Sample Station [With Sample Injection Mode Control Knob] Turn the switch module knob to the left (counterclockwise) until the flat side of knob is perpendicular to the front panel and the T is facing you.
  • Page 452 Sample Injection Mode Control Kit Switching the Sample Probe from the Plate Loader to the Single Tube Sample Station [With Sample Injection Mode Control Knob] B49006AH...
  • Page 453 APPENDIX D Table of Hazardous Substances Table of Hazardous Substances The Hazardous Substances Names and Concentration are shown in Table D.1 Table D.2. B49006AH...

  • Page 454: Table Of Hazardous Substances Name And Concentration

    Table D.1 Table of Hazardous Substances Name and Concentration [CytoFLEX] 电子电气产品号码 产品名称 Product Name: CytoFLEX EEP Part Number: A00-1-1102 产品型号 Product Model Number: A00-1-1102 部件名称 有毒有害物质或元素 Hazardous Substances Name Component Name 铅 汞 镉 六价铬 多溴联苯 多溴二苯醚 (Pb) (Hg) (Cd) (PBB) (PBDE) 印刷电路板组件...

  • Page 455: Table Of Hazardous Substances Name And Concentration

    Table D.2 Table of Hazardous Substances Name and Concentration [CytoFLEX LX] 电子电气产品号码 产品名称 Product Name: CytoFLEX LX EEP Part Number: B90883 产品型号 Product Model Number: B90883 部件名称 有毒有害物质或元素 Hazardous Substances Name Component Name 铅 汞 镉 六价铬 多溴联苯 多溴二苯醚 (Pb) (Hg) (Cd) (PBB)
  • Page 456 Table of Hazardous Substances Table of Hazardous Substances B49006AH...
  • Page 457 Gr Wt — gross weight APC-Cy7 — Allophycocyanin-Cyanin 7 H — humidity API — Application Programming Interface Hz — hertz Acq. — Acquisition IEC — International Electrotechnical Commission BCI — Beckman Coulter Incorporated IR — infrared BMP — bitmap kg — kilograms Abbreviations-1 B49006AH...
  • Page 458 Abbreviations KO — Krome Orange rCV — robust coefficient of variation LED — light emitting diode RH — relative humidity L — liter RoHS — Restriction of Hazardous Substances Directive LJ — Levey-Jennings RPTM — real-time messaging protocol LWH — length, width, height S/N —...
  • Page 459 Index Символи ’ define, Abbreviations-1 define, Abbreviations-1 " define, Abbreviations-1 define, Abbreviations-1 °C account define, Abbreviations-1 unlock, 2-26 °F account menu, 2-21 define, Abbreviations-1 account policies, 2-30 μ acoustic noise level, 1-25 define, Abbreviations-1 Acq. μL define, Abbreviations-1 define, Abbreviations-1 acquisition screen, μm collection,...
  • Page 460 Index alarm does not sound when the waste container bottle is full or the sheath fluid container is low Deep Clean solution, 1-12 and the software status display is red, 9-11 define, Abbreviations-1 analysis screen, analysis screen, calculating to open, compensation values, 6-8, 6-13 analyzing...
  • Page 461 Index 4 L waste container, 10-11 adjust, nonscheduled, 10-13 exporting, 6-19 routine, 10-1 importing and exporting, 5-75 sample probe, 10-8 compensation settings from compensation sample station, 10-7 library surfaces, 10-13 to import, 6-17 cleaning and ventilation compensation settings from compensation installation environment validation, matrix files cleaning solution...
  • Page 462 Index compensation matrix from previously data acquired data, 6-14 analyzing, 5-64 detector configuration, 5-25 collecting, 5-32 dot plot overlays, exporting, 5-64 heat map (plate loader), 5-15 sampling, 5-32 histogram overlays, to import, Levey-Jennings charts, 4-19 data populations are normal on one laser, but new experiment, too low on another laser, 9-15...
  • Page 463 Index to edit, 5-25 define, Abbreviations-1 to select, 5-25 emptying to verify, 5-25 10 L waste cubitainer (CytoFLEX LX), 11-10 detector configuration default, 5-26 4 L waste container (CytoFLEX), 11-9 detector unit wavelength division multiplexor (WDM), define, Abbreviations-1 disabling lasers, 5-54 enabling lasers, 5-54...
  • Page 464 Index statistics table of multiple tubes as a picture fluidics module file, 5-73 Deep Clean solution bottle, 1-12 user logs, 2-31 fluidics system, 1-12 exporting and experiment/template fluidics system component, 1-10 electronic record management, sheath fluid filters, 1-12 extent fluidics system autogate, 5-52 components,...
  • Page 465 Index graphic and gating styles, 2-32 inspecting materials graphics unpacking, printing, 5-75 inspection liquid flow tubing, 11-42 pre-boot, inspection when unpacking, installation define, Abbreviations-1 environment validation, hazard instrument, laser, software, A-12 hazards installation category, 1-25 laser beam, installation options radiation, CytExpert, A-12 hazards/precautions,...
  • Page 466 Index define, Abbreviations-1 items define, Abbreviations-2 adding for standardization, 4-24 Levey-Jennings charts deleting for standardization, 4-26 to create, 4-19 editing for standardization, 4-25 line segment gates importing for standardization, 4-26 for single-parameter plots, 2-33 liquid flow tubing inspection, 11-42 liquid flow tubing inspection routine maintenance, 11-42 define,...
  • Page 467 Index menu alarm, 1-12 account, 2-21 mute alerter icon, 1-12 acquisition and analysis screen, 2-15 advanced, 2-20 define, Abbreviations-2 cytometer, 2-17 files, 2-16 help, 2-22 log, 2-21 define, Abbreviations-2 QC/Standardization, 2-20 NaClO settings, 2-19 define, Abbreviations-2 signature, 2-22 NaN3 software, 2-14 define, Abbreviations-2...
  • Page 468 Index analysis screen, picture files optical to export, 5-73 components, plate holder optical bench components, 1-18 optical component, plate holder with plate loader optical components replacing, 11-45 detector (WDM), plate loader optical bench, components, 1-16 optical fiber, Plate Loader module optical fiber, installing, 11-46...
  • Page 469 Index cubitainers), QC data check waste and reagent levels (4 L fluid collecting, 4-10 containers), QC data with plate loader power source inspection, collecting, 4-12 workstation connections inspection, QC experiment screen, 2-12, 2-13 precaution QC failed, 9-21 disposal, QC menu tree precautions software, 2-15...
  • Page 470 Index CytExpert software, A-29 define, Abbreviations-2 reinstallation procedures right-side cover right-side cover, 11-5 reinstallation, 11-5 removal procedures removal, 11-4 front cover, 11-2 RoHS right-side cover, 11-4 caution label, removing define, Abbreviations-2 Plate Loader module, 11-46 environmental label, rename role management, 2-26 experiment directory (electronic record roles...
  • Page 471 Index calibrating, 11-56 automatically, 9-12 sample flow rate is unstable, 9-12 sample tube holder position sample flow rate with plate loader sample acquisition position, 1-15 calibrating, 11-59 sample loading position, 1-15 sample injection control knob standby position, 1-15 performance characteristics, sample wells with plate loader sample injection mode setting,...
  • Page 472 Index setup to start, A-24 CytExpert API test client, 2-37 software installation, A-12 experiment directory (electronic record installing CytExpert software, A-13 management), installing instrument configuration sheath fluid container, 1-10 file, A-21 to fill, required materials, A-13 sheath fluid container 4 L (CytoFLEX) starting the software, A-24 filling,...
  • Page 473 Index startup daily concentration calculation is incorrect, 9-18 routine maintenance, 10-1 connection indicator light in the lower left startup daily with plate loader corner of the software screen is red and routine maintenance, 10-5 displays Disconnected and Error, 9-10 station Cytometer cannot be turned on, 9-10 sample,...
  • Page 474 Index user account, 2-26 viewing unpacking inspection, user logs, 2-31 check instrument configuration, violet side scatter channel (VSSC) installing the instrument and connecting to set up, 5-29 the equipment, A-5, A-11 VSSC unpacking the instrument, define, Abbreviations-2 unstained samples to define the negative population, upgrade CytExpert software, A-24...
  • Page 475 Index samples, 5-13 sample, 5-13 sample injection mode selecting, 3-12 selecting proper sample injection mode, 3-12 Workstation, 1-2, 1-3, component, Workstation cannot be turned on, 9-10 workstation connections inspection pre-boot, worktable installation environment validation, Index-17...
  • Page 476 Index Index-18...
  • Page 477 Beckman Coulter Product software. Because some states do not allow the exclusion or limitation of liability for consequential damages, the above limitation might not apply to you.
  • Page 478 Beckman Coulter, Inc. Customer End User License Agreement Warranty-2 B49006AH...
  • Page 480 • CytExpert Electronic Record Management • Sample Injection Mode Control Kit • Table of Hazardous Substances • Abbreviations CytoFLEX Flow Cytometer Quick Start Guide PN B49008 CytoFLEX Setup Guide PN B53767 www.beckmancoulter.com © 2017 Beckman Coulter, Inc. All Rights Reserved...

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