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

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Instructions for Use
CytoFLEX Platform
CytoFLEX, CytoFLEX S, and CytoFLEX LX Flow Cytometers
For Research Use Only. Not for use in diagnostic procedures.
B49006AS
December 2021
Beckman Coulter, Inc.
250 S. Kraemer Blvd.
Brea, CA 92821 U.S.A.

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  • Page 1 Instructions for Use CytoFLEX Platform CytoFLEX, CytoFLEX S, and CytoFLEX LX Flow Cytometers For Research Use Only. Not for use in diagnostic procedures. B49006AS December 2021 Beckman Coulter, Inc. 250 S. Kraemer Blvd. Brea, CA 92821 U.S.A.
  • Page 2 • In the UK, call us at +44 845 600 1345. • In Ireland, call us at +353 (01) 4073082. • In Italy, call us at +39 0295392 456. • In other locales, contact your local Beckman Coulter Representative. Beckman Coulter (UK) Ltd.

  • 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.beckman.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.beckman.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.beckman.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.beckman.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.beckman.com and download the latest version of the manual or system help for your instrument.
  • Page 8 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.beckman.com and download the latest version of the manual or system help for your instrument.
  • Page 9 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.beckman.com and download the latest version of the manual or system help for your instrument.
  • Page 10 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.beckman.com and download the latest version of the manual or system help for your instrument.
  • Page 11 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.beckman.com and download the latest version of the manual or system help for your instrument.
  • Page 12 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.beckman.com and download the latest version of the manual or system help for your instrument.
  • Page 13 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.beckman.com and download the latest version of the manual or system help for your instrument.
  • Page 14 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.beckman.com and download the latest version of the manual or system help for your instrument.
  • Page 15 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.beckman.com and download the latest version of the manual or system help for your instrument.
  • Page 16 Revision History B49006AS...

  • Page 17: 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 18: Safety Precautions

    • This equipment is used in a manner other than specified. Operate the instrument as instructed in the product manuals. • Installation of software that is not authorized by Beckman Coulter onto your CytoFLEX workstation. Only operate the CytoFLEX with software authorized by Beckman Coulter.
  • Page 19 Safety Notices Symbol Explanations CAUTION Risk of instrument damage. This device is intended for indoor use only. To avoid device damage, do not install the instrument outdoors. WARNING Risk of personal injury. Safety protection can be impaired if used in a manner not specified by the manufacturer.
  • Page 20 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 21 Safety Notices Symbol Explanations Symbol Warning Condition Action California Proposition This product can expose you to chemicals 65 Labeling including phthalates, which are known to the State of California to cause cancer and birth defects or other reproductive harm. For more information go to www.P65Warnings.ca.gov.
  • Page 22 Safety Notices Symbol Explanations xxii B49006AS...

  • Page 23: Table Of Contents

    Sample Tube Holder Positions, 1-22 Plate Loader Components, 1-23 Plate Holder Components, 1-27 System Configuration, 1-31 System Configuration [CytoFLEX and CytoFLEX S], 1-31 System Configuration [CytoFLEX LX], 1-33 Consumables and Supplies, 1-35 Reagents, 1-35 Material Safety Data Sheets (SDS/MSDS), 1-35...
  • Page 24 Category, 1-37 Pollution Degree, 1-38 Acoustic Noise Level, 1-38 Electrical Ratings, 1-38 Cytometer, 1-38 Performance Characteristics, 1-41 Performance Characteristics [CytoFLEX and CytoFLEX S], 1-41 Performance Characteristics [CytoFLEX LX], 1-43 Reagent Limitations, 1-44 CHAPTER 2: Using the CytExpert Software, 2-1 Overview, 2-1...
  • Page 25 Contents Plate Type Library, 2-36 Adding a Plate Type, 2-37 Editing a Plate Type, 2-41 Duplicating a Plate Type, 2-42 Deleting a Plate Type, 2-44 Software Settings, 2-45 Language Settings, 2-48 Setting Up CytExpert Application Programming Interface (API) Test Client, 2-48 CHAPTER 3: Operation Principles, 3-1...
  • Page 26 Preparing the QC Sample, 5-3 Required Materials, 5-3 CytoFLEX Daily QC Fluorospheres Preparation Process, 5-3 CytoFLEX Ready to Use Daily QC Fluorospheres Preparation Process, 5-4 CytoFLEX Daily IR QC Fluorospheres Preparation Process, 5-4 Preparing the QC Sample [With Plate Loader], 5-5...
  • Page 27 Setting Custom Statistics, 6-34 Configuring Acquisition Settings, 6-38 Changing the Tube Name, 6-38 Laser Settings, 6-38 Setting Laser Target Power Settings [CytoFLEX LX Only], 6-40 Adjusting the Gain, 6-41 Adjusting the Threshold, 6-43 Setting Collection Conditions, 6-44 Setting Plot Display...
  • Page 28 Results, 8-9 CHAPTER 9: Daily Shutdown, 9-1 Overview, 9-1 Preparing the Cleaning Solution, 9-1 Shutting Down the Instrument, 9-1 Auto Shutdown [CytoFLEX LX Only], 9-2 CHAPTER 10: Troubleshooting, 10-1 Overview, 10-1 Precautions/Hazards, 10-1 Laser Related Hazards, 10-1 Laser Beam Hazards, 10-2...
  • Page 29 Replacing the Sample Probe Assembly [With Plate Loader], 12-32 Changing the Sample Probe from the Single Tube Sample Station to the Plate Loader [CytoFLEX With Plate Loader], 12-38 Changing the Sample Probe from the Plate Loader to the Single Tube Sample Station [CytoFLEX With Plate...
  • Page 30 Installing the Instrument and Connecting the Equipment [CytoFLEX], A-5 Installing the Instrument and Connecting the Equipment [CytoFLEX LX], A-11 CytExpert Software Installation Options, A-12 Installing the Software [CytoFLEX Platform], A-12 Required Materials, A-13 Installing the CytExpert Software, A-13 Installing the Instrument Configuration...
  • Page 31 Contents Folder Hierarchy Management, B-4 Experiment Related Operations, B-5 Importing an Experiment/Template, B-6 Exporting an Experiment/Template, B-9 Log, B-12 Experiment Operation Log, B-12 System Operation Log, B-15 User Management Operation Log, B-16 Electronic Signature, B-17 Signing Experiments, B-17 Rejecting Experiment, B-20 Setting the Signature Retention Period, B-21 Signature...
  • Page 32 Enabling SMB Digitally Sign Communications, G-92 Enabling Installation Restriction, G-95 Enabling Firewall Defender, G-97 Enabling Network Time Protocol, G-99 APPENDIX H: Table of Hazardous Substances, H-1 Table of Hazardous Substances, H-1 Abbreviations Beckman Coulter, Inc. SOFTWARE END-USER LICENSE AGREEMENT Related Documents xxxii...
  • Page 33 1.21 Calibration Tool for 96-Well Deep Well Plate, 1-30 1.22 System Connections, 1-31 1.23 Back Cover Connections, 1-32 1.24 Front of Cytometer [CytoFLEX Without Plate Loader Shown], 1-32 1.25 System Connections, 1-33 1.26 Back Cover Connections, 1-34 1.27 Front of Cytometer, 1-34 1.28...
  • Page 34 Sample, 7-8 10.1 Laser Warning Label on the Laser Optical Bench [CytoFLEX], 10-3 10.2 Laser Warning Label on the Laser Optical Bench [CytoFLEX LX], 10-4 10.3 Laser Warning Label within the Optical Bench (Located Inside the Cytometer) [CytoFLEX and CytoFLEX S], 10-4 10.4...
  • Page 35 10.10 Biohazard Label Located in the Sample Station and on the Back of the Cytometer [CytoFLEX Shown], 10-7 10.11 Electrical Shock Hazard Label by the Power Switch [CytoFLEX or CytoFLEX S], 10-7 10.12 Caution Labels [CytoFLEX or CytoFLEX S], 10-8 10.13...
  • Page 36 Illustrations xxxvi...
  • Page 37 WDM Optical Filter Mount Color Codes [CytoFLEX S_Violet-Blue-Red-IR], 1-11 WDM Optical Filter Mount Color Codes [CytoFLEX LX without WDM Beam splitter], 1-12 WDM Optical Filter Mount Color Codes [CytoFLEX LX with WDM Beam Splitter], 1-14 Target Power Ranges in the Laser Setting Screen, 6-39 10.1...
  • Page 38 Tables xxxviii...

  • Page 39: Introduction

    CytoFLEX flow cytometer does and the methods guiding operation. It also contains procedures for cleaning and maintenance. • The CytoFLEX Setup Guide provides instructions for unpacking and setting up the CytoFLEX flow cytometer system. • The CytoFLEX S Special Configuration Specifications package insert contains the sections and procedures that are specific to the CytoFLEX S series.

  • Page 40: Sample Injection Mode Control

    CHAPTER 5, 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. Quality control allows you to determine whether your instrument can provide adequate signal strength and precision.

  • Page 41: Conventions Used, Xli

    To access the linked information, select the blue, underlined text. • The information in your Instructions for Use manual applies to the CytoFLEX and CytoFLEX S instruments equipped with and without a plate loader, and the CytoFLEX LX, unless otherwise specified.
  • Page 42 All graphics, including screens and printouts, are for illustration purposes only and must not be used for any other purpose. For example, software screens that show the CytoFLEX system in the background may not depict the latest production version of the system.

  • Page 43: System Overview

    The CytoFLEX S may be ordered in various configurations from 2 lasers, 4 colors to a maximum of 4 lasers, 13 colors. The CytoFLEX LX flow cytometer can perform up to 21 color marker analysis. The system can be ordered in various configurations from a 4 laser, 14 colors, to a maximum of 6 lasers, 21 colors.

  • Page 44: Main Components

    The instrument consists of three main components: Fluid Containers/Cubitainers, Cytometer, and the Workstation. Figure 1.1 Main Components [CytoFLEX Without Plate Loader] 1. Fluid Containers. Accommodates sheath fluid and waste liquids as required for operation of the instrument.

  • Page 45: Main Components [Cytoflex Lx]

    1. Fluid Cubitainers. Accommodates sheath fluid and waste liquids as required for operation of the instrument. NOTE The CytoFLEX LX does not have a fluid container holder. 2. Cytometer. Provides signal generation and collection. 3. Workstation. Displays and manipulates the contents of the Workstation and displays data generated by the Cytometer.

  • Page 46: Optical Components

    Risk of instrument damage. Do not place sample tubes in the optical filter holder. Liquid spills can damage instrument components. Use a tube rack to hold any sample tubes. 4. Optical filter holder. Securely holds additional CytoFLEX platform optical filters. B49006AS...

  • Page 47: 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 WDM and replace the filters inside.

  • Page 48: 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 49: Wdm Optical Filter Mount Color Codes [Cytoflex]

    APC-A700, Alexa 712/25 BP Fluor™700, Cy5.5, DRAQ7™ APC-A750 APC-A750, APC-Cy7, 780/60 BP APC-H7, APC- eFluor™ 780, DRAQ7™ Table 1.2 WDM Optical Filter Mount Color Codes [CytoFLEX S_Violet-Blue-Yellow-Red] CytoFLEX Channel Commonly used Laser Fluorescent Channel Names Fluorescent Dyes PB450 Pacific Blue™dye, V450,...
  • Page 50 System Overview Optical Components Table 1.2 WDM Optical Filter Mount Color Codes [CytoFLEX S_Violet-Blue-Yellow-Red] (Continued) CytoFLEX Channel Commonly used Laser Fluorescent Channel Names Fluorescent Dyes FITC FITC, Alexa Fluor™ 488, 488 nm 525/40 BP CFSE, Fluo-3 PC5.5 PC5.5, PC5, PerCP, 690/50 BP PerCP-Cy5.5, PI, DRAQ7™...

  • Page 51: Wdm Optical Filter Mount Color Codes [Cytoflex S_Nuv-Violet-Blue-Red]

    System Overview Optical Components Table 1.3 WDM Optical Filter Mount Color Codes [CytoFLEX S_NUV-Violet-Blue-Red] CytoFLEX Commonly used Laser Fluorescent Channel Channel Names Fluorescent Dyes NUV450 BUV395, DAPI 375 nm 450/45 BP NUV675 Hoescht Red, BUV661 675/30 BP PB450 Pacific Blue™dye, V450,...
  • Page 52 System Overview Optical Components Table 1.4 WDM Optical Filter Mount Color Codes [CytoFLEX S_NUV-Violet-Blue-Yellow] (Continued) CytoFLEX Channel Commonly used Laser Fluorescent Channel Names Fluorescent Dyes PB450 Pacific Blue™dye, V450, 405 nm 450/45 BP eFluor™ 450, BV421 KO525 Krome Orange, AmCyan,...

  • Page 53: Wdm Optical Filter Mount Color Codes [Cytoflex S_Violet-Blue-Red-Ir]

    Alexa Fluor®790 808 nm 840/20 BP IR885 PromoFluor-840, IR fixable 885/40 BP viability dye NOTE The CytoFLEX and CytoFLEX S have the following additional BP filters supplied within the respective WDM: • 405/10 BP • 638/6 BP • 561/6 BP •...

  • Page 54: Wdm Optical Filter Mount Color Codes [Cytoflex Lx Without Wdm Beam Splitter]

    System Overview Optical Components Table 1.6 WDM Optical Filter Mount Color Codes [CytoFLEX LX without WDM Beam splitter] CytoFLEX Channel Commonly used Laser Fluorescent Channel Names Fluorescent Dyes UV405 BUV395 355 nm 405/30 BP UV525 BUV496 525/40 BP UV675 Hoescht Red, BUV661...
  • Page 55 System Overview Optical Components Table 1.6 WDM Optical Filter Mount Color Codes [CytoFLEX LX without WDM Beam splitter] (Continued) CytoFLEX Channel Commonly used Laser Fluorescent Channel Names Fluorescent Dyes R763-APCA750 APC-A750, APC-Cy7, 638 nm 763/43 BP APC-H7, APC- eFluor™ 780, DRAQ7™...

  • Page 56: Wdm Optical Filter Mount Color Codes [Cytoflex Lx With Wdm Beam Splitter]

    System Overview Optical Components Table 1.7 WDM Optical Filter Mount Color Codes [CytoFLEX LX with WDM Beam Splitter] CytoFLEX Channel Commonly used Laser Fluorescent Channel Names Fluorescent Dyes UV405 BUV395 355 nm 405/30 BP UV525 BUV496 525/40 BP UV675 Hoescht Red, BUV661...
  • Page 57 System Overview Optical Components Table 1.7 WDM Optical Filter Mount Color Codes [CytoFLEX LX with WDM Beam Splitter] (Continued) CytoFLEX Channel Commonly used Laser Fluorescent Channel Names Fluorescent Dyes Y610-mCherry mCherry, ECD, PE-CF594 561 nm 610/20 BP Y763-PC7 763/43 BP...

  • Page 58: Optical Fiber

    PRÉCAUTION LASER CLASSE 3B RADIATION LASER QUAND L ' INSTRUMENT EXT OUVERT ET NON VERROUILLE ÉVITER TOUTE EXPOSITION AU FAISCEAU CytoFLEX CytoFLEX LX 1. Red laser fiber 2. Violet laser fiber 3. Blue laser fiber 4. Infrared laser fiber 5. Blue laser fiber 6.

  • Page 59: 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 60: Fluid Containers/Cubitainers

    The waste cubitainer must be on the same level as the Cytometer. Fluid Containers/Cubitainers On the CytoFLEX and CytoFLEX S, two Fluid Containers are placed in the Fluid Container holder: a sheath fluid container and a waste container. See Figure 1.7.

  • Page 61: Fluidics Module

    CHAPTER 12, Replacement/Adjustment Procedures). Figure 1.9 Fluidics Module View CytoFLEX CytoFLEX LX 1. Alarm. Emits a warning sound when there is a problem with the Fluid Container/Cubitainer capacity or with the performance of certain operations. NOTE When the alarm sounds, the Mute Alerter icon appears in the status bar. The alarm continues for about 30 seconds.

  • Page 62: Fluidic Connections

    6. Sheath fluid in. Connects to the sheath fluid tubing. NOTE Use CytoFLEX Sheath Fluid or other filtered nonionic sheath fluid. Using unfiltered sheath fluid can shorten the service life of the sheath fluid filters and increase noise and debris detection.

  • Page 63: 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 64: 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 by looking directly at the sample tube holder while the Cytometer is processing a sample.

  • Page 65: Plate Loader Components

    System Overview Plate Loader Components Plate Loader Components Figure 1.13 Standard Plate Loader [CytoFLEX Shown] 1. Plate loader door 2. Plate holder stage 3. Plate holder (removable) 1-23 B49006AS...
  • Page 66 System Overview Plate Loader Components Figure 1.14 Plate Loader DW [CytoFLEX Shown] 1. Plate loader door 2. Plate holder stage 3. Plate holder (removable) NOTE The Plate Loader DW supports both standard 96-well plates, and 96-well deep well plates. 1-24...

  • Page 67: Standard Plate Loader

    System Overview Plate Loader Components Figure 1.15 Standard Plate Loader (Front Cover Removed) 1. Plate loader PEEK tubing 2. Plate loader sample probe assembly 3. Plate loader wash station 4. Plate holder 1-25 B49006AS...
  • Page 68 System Overview Plate Loader Components Figure 1.16 Plate Loader DW (Front Cover Removed) 1. Plate loader PEEK tubing 2. Plate loader sample probe assembly 3. Plate loader wash station 4. Plate holder 1-26 B49006AS...

  • Page 69: Plate Holder Components

    System Overview Plate Loader Components Plate Holder Components Figure 1.17 Standard Plate Holder without Groove (Standard Plate Loader) 1. Spring leaves to hold plate 2. Plate holder notches 1-27 B49006AS...

  • Page 70: Plate Holder With Groove (Plate Loader Dw)

    System Overview Plate Loader Components Figure 1.18 Plate Holder with Groove (Plate Loader DW) 1. Spring leaves to hold plate NOTE The plate holder is removable and replaceable. Refer to Replacing the Plate Holder [With Plate Loader] CHAPTER 12, Replacement/Adjustment Procedures.

  • Page 71: Plate Holder Stage (Standard Plate Loader)

    System Overview Plate Loader Components Figure 1.19 Plate Holder Stage (Standard Plate Loader) 1. Pins 2. Position A1 Figure 1.20 Plate Holder Stage (Plate Loader DW) 1. Position A1 2. Spring Lock 1-29 B49006AS...

  • Page 72: Calibration Tool For 96-Well Deep Well Plate

    System Overview Plate Loader Components Figure 1.21 Calibration Tool for 96-Well Deep Well Plate 1. Calibration frame 2. Transparent plate NOTE The calibration frame and the transparent plate are used to assist the calibration in X-axis and Y-axis. They are delivered together with the Plate Loader DW. 1-30 B49006AS...

  • Page 73: System Configuration

    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 and CytoFLEX S] Figure 1.22 System Connections CLASS 1 LASER PRODUCT COMPLIES WITH 21 CFR 1040.10 AND 1040.11...

  • Page 74: Back Cover Connections

    2. Fuse. Protects the internal system from damage by high electrical current. 3. Power line socket. Supplies the power to the Cytometer. Figure 1.24 Front of Cytometer [CytoFLEX Without Plate Loader Shown] 1. Load button. In addition to the software controls, this button can be used for automatic sample loading and data recording.

  • Page 75: 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 76: Front Of Cytometer

    System Overview System Configuration Figure 1.26 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 77: Consumables And Supplies

    The following reagents are available for the CytoFLEX, CytoFLEX S and CytoFLEX LX instrument: CytoFLEX Ready to Use Daily QC Fluorospheres The CytoFLEX Ready to Use Daily QC Fluorospheres are a ready to use formulation of fluorospheres that are used for QC standardization on the CytoFLEX Platform.

  • Page 78: Instrument Specifications

    Plate Loader] Cytometer [With Plate 28.4 kg Loader DW] Refer to Figure 1.28 for the [CytoFLEX and CytoFLEX S] Plate Loader DW dimensions. Figure 1.28 Plate Loader DW Dimensions [CytoFLEX and CytoFLEX S] 25.4 cm (10.0 in) 4.3 cm (1.7 in) 11.5 cm...

  • Page 79: Dimensions [Cytoflex Lx]

    83.6 kg Plate Loader] Cytometer [With Plate 84 kg Loader DW] Refer to Figure 1.29 for the [CytoFLEX LX] Plate Loader DW dimensions. Figure 1.29 Plate Loader DW Dimensions [CytoFLEX LX] 35.3 cm (13.9 in) 5.5 cm (2.2 in) 37.5 cm 19.4 cm...

  • Page 80: Pollution Degree

    QC system, if required. No user intervention is required to ensure optimum system performance. CytoFLEX: The system can be configured with up to three spatially-separated lasers. CytoFLEX S: The system can be configured with up to four spatially-separated lasers.
  • Page 81 Custom Flow Rates: 10 - 240 μL/min in 1 μL increments Fluid capacity CytoFLEX and CytoFLEX S: 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...
  • Page 82 Refer to the related specification of the deep well plates. Volume The 96-well deep well plates are only available for use if the Plate Loader DW is installed. The information below is based on the deep well plates manufactured by Beckman Coulter. Refer to APPENDIX D, Deep Well Plate.

  • Page 83: Performance Characteristics

    <3% resolution The CytoFLEX Flow cytometer is capable of achieving <3% rCV. Using CytoFLEX Daily QC Fluorospheres, CytoFLEX Ready to Use Daily QC Fluorospheres or Daily IR QC Fluorospheres (for 808 nm Laser) for daily QC, the pass criteria is ≤5% for the Violet, Blue, Yellow, and Red lasers while the pass criteria is ≤7% for NUV, and...

  • Page 84: Plate Loader Dw

    10 second acquisition with 3 second mixing and 3 second backflush: <45 min. This performance characteristic is different if you have the Sample Injection Mode Control Kit installed on your CytoFLEX cytometer. Refer to APPENDIX C, Sample Injection Mode Control Kit.

  • Page 85: Performance Characteristics [Cytoflex Lx]

    10 second acquisition with 3 second mixing and 3 second backflush: < 47 min. Loader] This performance characteristic is different if you have the Sample Injection Mode Control Kit installed on your CytoFLEX LX flow cytometer. Refer to APPENDIX C, Sample Injection Mode Control Kit.

  • Page 86: Reagent Limitations

    System Overview Reagent Limitations Reagent Limitations • Only use nonionic sheath fluid with antimicrobial, like CytoFLEX Sheath Fluid. Do not use sheath fluid containing electrolytes. • Do not use organic solvents in the system. 1-44 B49006AS...

  • Page 87: 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 operation, collection of experiment data, and analysis of the results. This chapter will explain the software functions and features. This chapter contains information on: Launching the Software •...

  • Page 88: 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: • . For creating a new experiment. The process creates a file with the .xit New Experiment extension and a folder with the same file name where the raw data (.fcs files) are kept.

  • Page 89: 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 Selecting , or automatically opens New Experiment New Experiment From Template Open Experiment the Acquisition screen.

  • Page 90: 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. B49006AS...

  • Page 91: Collection

    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 92: Collection [With Plate Loader]

    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 93: 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 94: 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 95: Status Bar

    4. Sampler status. Displays the sample injection mode state. There are two sample injection modes: Semi-automatic sample injection mode and manual sample injection mode. NOTE CytoFLEX Cytometers equipped with a plate loader have three sample injection modes: Semi-automatic sample injection mode, manual sample injection mode, and plate loader sample injection mode.

  • Page 96: Drawing Controls Toolbar (Top Of 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. Tube Management Drawing controls (see Figure 2.1) include the multi-data histograms and graphical display data controls.

  • Page 97: Compensation Experiment Screen

    Using the CytExpert Software Main Software Screen Compensation Experiment Screen The Compensation Experiment screen appears when you open or create a new compensation experiment. 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 98: Qc Experiment Screen

    Using the CytExpert Software Main Software Screen Zoom Print 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 1.

  • Page 99: 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 100: Software Menu

    *** These options are only available if the CytExpert Electronic Record Management software option is installed. ††† These options are only available on the CytoFLEX LX flow cytometer. icon is hyper linked to the Cytobank spotlight page where you can login to the Cytobank Premium server, request a 30 day free trial and access additional information.

  • Page 101: Acquisition And Analysis Screen Menu

    ‡‡ These options are only available if either the CytExpert User Management or the CytExpert Electronic Record Management software option is installed. ***These options are only available on the CytoFLEX LX flow cytometer. Acquisition and Analysis Screen Menu CytExpert Default Software Option...
  • Page 102 Using the CytExpert Software Main Software Screen CytExpert Default Software Option CytExpert User Management Software Option 2-16 B49006AS...
  • Page 103 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 104 Using the CytExpert Software Main Software Screen CytExpert User Management Software Option - CytExpert User Management Software Option - Standby state Initialized state 2-18 B49006AS...
  • Page 105 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. CytExpert Default Software Option...
  • Page 106 Using the CytExpert Software Main Software Screen CytExpert User Management Software Option CytExpert Electronic Record Management Software Option QC/Standardization Menu Select from the QC/Standardization menu to start the QC routine. Start QC/Standardization NOTE The QC/Standardization menu is the same for the CytExpert Default, CytExpert User Management, and the CytExpert Electronic Record Management software options.
  • Page 107 Using the CytExpert Software Main Software Screen CytExpert Default Software Option - CytExpert Default Software Option - Plate Loader Semi-Automatic/Manual Sample Injection Mode Sample Injection Mode CytExpert User Management Option - CytExpert User Management Software Option - Semi-Automatic/Manual Sample Injection Mode Plate Loader Sample Injection Mode CytExpert Electronic Record Management CytExpert Electronic Record Management...
  • Page 108 Using the CytExpert Software Main Software Screen Log Menu Used to access logs including the Experiment Operation Log, the System Operation Log, and the User Management Operation Log. CytExpert User Management Software Option CytExpert Electronic Record Management Software Option NOTE The Log menu is not available in the CytExpert Default software option.

  • Page 109: User Management

    Using the CytExpert Software User Management User Management 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 110: User Manager (Grid View)

    Using the CytExpert Software User Management 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. View drop-down: Toggles between Card View NOTE An account locks after 3 failed password (see...

  • Page 111: 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 112 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 113: 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 114: Role Manager

    Using the CytExpert Software Role Management Select . The Role Manager window appears. Refer to Figure 2.7. Account Role Manager > 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 115: 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 116 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 117: Account Policies

    Using the CytExpert Software Account Policies Select Account Policies IMPORTANT Only an Administrator or an account has the Manage user permission 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...

  • Page 118: Account Policies - Account Lockout Policy

    Using the CytExpert Software Account Policies Figure 2.9 Account Policies - Account Lockout Policy NOTE The allowable range for each entry is as follows: • Invalid Login Attempts: 3-10 times • Lockout Duration: 15-1440 minutes Figure 2.10 Account Policies - Application Inactivity Policies NOTE The allowable range for each entry is as follows: •...

  • Page 119: 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 120: 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 121: 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 122: Plate Type Library

    Using the CytExpert Software Plate Type Library Vertical gate a. This gate can be created using the autogate functionality. Refer to Creating and Adjusting Auto Gates CHAPTER 6, Data Acquisition and Sample Analysis Plate Type Library The Plate Type Library is used to manage and calibrate plates. Plates can be added, deleted, duplicated, and edited from the Plate Type Library.

  • Page 123: Adding A Plate Type

    Using the CytExpert Software Plate Type Library Adding a Plate Type Select Select . The Plate Type Library window appears. Advanced > Plate Type Library Select . The Add Plate Type window appears. 2-37 B49006AS...
  • Page 124 Using the CytExpert Software Plate Type Library Enter the plate name in the Name section of the screen. Select the Plate Mode. 2-38 B49006AS...
  • Page 125 Using the CytExpert Software Plate Type Library Enter the Mix time. NOTE The default setting is 5 seconds in Standard Mixing Mode. The default setting is 10 seconds (for 0.5 mL sample) in Deep Well Mixing Mode. You might need custom the Mix time according to the sample volume.
  • Page 126 Using the CytExpert Software Plate Type Library Select . The plate is added into the Plate Type Library. Select 2-40 B49006AS...

  • Page 127: Editing A Plate Type

    Using the CytExpert Software Plate Type Library Editing a Plate Type Select . The Plate Type Library window appears. Advanced > Plate Type Library Select the plate type to be edited, and select . The Edit Plate Type window appears. Edit Enter the Mix setting.

  • Page 128: Duplicating A Plate Type

    Using the CytExpert Software Plate Type Library Optional: Select to calibrate the plate position. Refer to Calibrating the Plate Position Calibrate [With Plate Loader] CHAPTER 12, Replacement/Adjustment Procedures The Calibration can be performed at any time. Select Duplicating a Plate Type Select .
  • Page 129 Using the CytExpert Software Plate Type Library Select the plate type to duplicate, and select . The Duplicate Plate Type window Duplicate appears. Enter the Mix setting. Optional: Enter any remarks. Optional: Select to calibrate the plate position. Refer to Calibrating the Plate Position Calibrate [With Plate Loader]...

  • Page 130: Deleting A Plate Type

    Using the CytExpert Software Plate Type Library Deleting a Plate Type Select . The Plate Type Library window appears. Advanced > Plate Type Library Select the plate type to be deleted, and select . The following message appears: Delete NOTE The default plate types, shown in bold, cannot be deleted from the Plate Type Library.

  • Page 131: Software Settings

    Using the CytExpert Software Software Settings Software Settings Select in the Settings menu to configure the software settings. Options In the experiment settings, you can set the experiment’s default save path. NOTE The Experiment setting is only available if either the CytExpert Default or the CytExpert User Management software option is installed.
  • Page 132 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 133 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. Select to display page boundaries within the Acquisition or Analysis Show page breaks views for simplifying plot arrangement for printing.

  • Page 134: Language Settings

    Setting Up CytExpert Application Programming Interface (API) Test Client The CytExpert API is available for external software to control CytoFLEX Platform. It allows external software to perform operations such as running methods and allows for basic control of the plate loader.

  • Page 135: Operation Principles

    CHAPTER 3 Operation Principles Overview This chapter explains how the Cytometer measures scattered light and fluorescence as cells pass through the laser beam. The illustrations in this chapter are not exact representations of the inside of the Cytometer. They are for explanatory purposes only. This chapter contains information on: •...

  • Page 136: Hydrodynamic Focusing

    Operation Principles Sample Flow (sample peristaltic pump motor) moves the sample peristaltic pump to achieve liquid flow. Refer to Figure 1.12. The sampler module handles a sample tube as follows: • It swings the tube holder out for loading. • It swings the tube holder in and lifts the tube holder. •...

  • Page 137: Laser Beam Shaping

    Operation Principles Laser Beam Shaping Figure 3.1 Flow Cell 1. Sheath stream 2. Sample stream 3. Laser beam 4. Waste out 5. Purge port (Waste) Laser Beam Shaping Before the laser beam reaches the sample stream, lenses focus the beam (see Figure 3.2).

  • Page 138: Cell Illumination

    Operation Principles Cell Illumination Figure 3.2 Laser Beam Shaping 1. Violet laser beam 4. First stage shaping lens 2. Blue laser beam 5. Second stage shaping lens 3. Red laser beam 6. Flow cell Cell Illumination As cells in the sample stream go through the sensing area of the flow cell, the elliptical beam illuminates them.

  • Page 139: Side Scatter And Fluorescent Light Collection

    Operation Principles Light Collection, Separation and Measurement forward angle light is filtered with a 488 nm band pass before it reaches the FS sensor which generates voltage pulse signals. These signals are proportional to the amount of light the sensor receives.

  • Page 140: Signal Processing

    Operation Principles Signal Processing Signal Processing The CytoFLEX is a fully digital system with an active range of 7 logarithmic decades, signal collection speed of 30,000 events/second (includes 15 parameters). Data Storage CAUTION The instrument Workstation is vulnerable to malware, viruses, data corruption, and unauthorized instrument setting changes, or privacy breaches if unauthorized access is gained by malicious personnel.

  • Page 141: Parameters

    Operation Principles Parameters . There are two types of autogates available in the CytExpert for CytoFLEX software: • Auto Gating auto line segment and auto polygon. Parameters TIME Parameter The TIME parameter is the amount of time, in seconds, the instrument acquires data. It is displayed on the plot.

  • Page 142: Statistics

    Operation Principles Statistics Statistics The Statistics Setting window allows you to change the display of the header, statistical elements and cell populations included. B49006AS...

  • Page 143: Daily Startup

    Ô Ô inspection This chapter contains information on: Pre-Startup Inspection • • Turning On the Instrument Logging into the Software • • Initializing the Instrument Pre-Startup Inspection Before using the CytoFLEX platform flow cytometer, perform the following system checks. B49006AS...

  • Page 144: Check Waste And Reagent Levels [4 L Fluid Containers]

    Container holder before filling the sheath fluid container to avoid damage to instrument electronics. If necessary, fill the sheath fluid container with CytoFLEX Sheath Fluid or a similar nonionic sheath fluid while not exceeding the maximum volume indicated (4 L). Refer to...
  • Page 145 Daily Startup Pre-Startup Inspection WARNING Risk of chemical injury from bleach. To avoid contact with the bleach, use barrier protection, including protective eyewear, gloves, and suitable laboratory attire. Refer to the Safety Data Sheet for details about chemical exposure before using the chemical.

  • Page 146: Check Waste And Reagent Levels [10 L Fluid Cubitainers]

    Beckman Coulter CytoFLEX flow cytometers. REACTIVE INGREDIENTS: Anti-microbial compound Confirm that the instrument is in the standby state. If necessary, replace the sheath fluid cubitainer with CytoFLEX Sheath Fluid or a similar nonionic sheath fluid. Refer to Replacing the 10 L Sheath Fluid Cubitainer...

  • Page 147: 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 148: 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 149 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 150 • 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. B49006AS...

  • Page 151: 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 Log out.
  • Page 152 Daily Startup Logging into the Software purposes: running 1.5-mL and 2-mL microcentrifuge sample tubes and a backup mode that allows you to continue to collect data if the Semi-Automatic Injection mode is not working correctly. Using Semi-Automatic Injection Mode Select >...
  • Page 153 Daily Startup Logging into the Software At the flow cell, the sample runs at the designated flow rate and the Cytometer begins to acquire data. NOTE You can also push the load button on the front of the instrument to automatically start the run and record the data.

  • Page 154: Selecting The Plate Loader Sample Injection Mode [With Plate Loader]

    Daily Startup Logging into the Software Select Initialize Load the sample tube. NOTE The sample tube holder accommodates 1.5-mL, 2.0-mL, and 12 x 75 mm sample tubes. Manually swing the sample tube holder gently back to the standby position (see Figure 1.12).
  • Page 155 The restart warning only appears when switching to and from the Plate Loader sample injection mode. Turn the Cytometer's main power switch off. NOTE If you have a CytoFLEX LX instrument, you can turn the Cytometer’s power off by selecting Cytometer Turn Off. >...
  • Page 156 Turn the Cytometer’s main power switch on. The sampler status icon located in the bottom right side of the screen changes to display Plate Loader. NOTE If you have a CytoFLEX LX instrument, you can turn the Cytometer’s power on by selecting Cytometer Turn On. >...
  • Page 157 Daily Startup Logging into the Software [Standard 96-Well Plate in the Plate Holder (With Groove)] [96-Well Deep Well Plate Only] NOTE Ensure that plate well A1 aligns with position A1 on the plate holder. Select to load the plate. Load Select .

  • Page 158: Running The System Startup Program [With The Single Tube Loader]

    8 minutes if the fluidic self-check is not enabled. NOTE The fluidic self-check is an optional feature only available when the sheath damper upgrade is implemented. Select Initialize Select in the Cytometer menu. System Startup Program [CytoFLEX LX Shown] 4-16 B49006AS...
  • Page 159 Daily Startup Logging into the Software If the confirm window appears, select The System Startup Program window appears. Select Initialize. System Startup Program Window in Semi-Automatic Injection Mode System Startup Program Window in Manual Injection Mode 4-17 B49006AS...
  • Page 160 Daily Startup Logging into the Software Wait for the system to initialize. Follow the on screen software prompts, then select Start. The instrument begins to prime or run the fluidic self-check. This process takes about 4 minutes if the fluidic self-check is enabled. This process takes about 1 minute if the fluidic self-check is not enabled.

  • Page 161: 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. When warm up is finished, select to quit the startup program. The system is now Close initialized.
  • Page 162 Daily Startup Logging into the Software If the confirm window appears, select Select from the Cytometer menu to open the System Startup Program System Startup Program window. [CytoFLEX LX Shown] 4-20 B49006AS...
  • Page 163 Daily Startup Logging into the Software The plate loader automatically ejects the plate holder stage and the System Startup Program window appears. 4-21 B49006AS...
  • Page 164 Daily Startup Logging into the Software Select the desired plate type from the Plate Type drop-down menu. NOTE The available plate types included in the drop-down menu depend on the settings selected in the Plate Library. To activate a plate type, refer to Plate Type Library CHAPTER 2, Using the CytExpert Software.
  • Page 165 Daily Startup Logging into the Software Follow the on screen software prompts and select the desired wells and select Set As Deionized 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 166 Daily Startup Logging into the Software Wait for the system to initialize. The instrument begins prime or run the fluidic self-check. This process takes about 1 minute if the fluidic self-check is not enabled. This process takes about 4 minutes if the fluidic self-check is enabled. 4-24 B49006AS...
  • Page 167 Daily Startup Logging into the Software After priming or running the fluidic self-check, the system initializes again. The sample is loaded automatically. This process takes about 1 minute per well. 4-25 B49006AS...
  • Page 168 Daily Startup Logging into the Software When the system finishes acquiring the selected sample well, it uses the remaining time to warm up. 4-26 B49006AS...

  • Page 169: Selecting Experiments From The Start Page

    Daily Startup Logging into the Software When warm up is complete the plate loader ejects the plate holder stage. Select to quit Close 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 170: Initializing The Instrument

    Daily Startup Initializing the Instrument Initializing the Instrument Select in the Data Acquisition Control screen or select in the Cytometer Menu Initialize Initialize to initialize the instrument. NOTE The system prompts you to initialize if the instrument remains in standby for over 24 hours. 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...

  • Page 171: Chapter 5: Instrument Quality Control And Standardization

    Standardization Overview This chapter provides information on performing daily Quality Control (QC) on the CytoFLEX 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 172: Quality Control

    Instrument Quality Control and Standardization Quality Control Obtain target Apply Perform acquisition standardization standardization Ô Ô Ô standardization settings sample settings This chapter contains information on: • Preparing the QC Sample • Preparing the QC Sample [With Plate Loader] Importing Lot-Specific Target Values •...

  • Page 173: Preparing The Qc Sample

    Take one sample tube and label it as the QC sample tube. Add approximately 1 mL of deionized water to the sample tube. Use the vortexer or shake vigorously to thoroughly mix the bottle of CytoFLEX Daily QC Fluorospheres. Add 3 drops of CytoFLEX Daily QC Fluorospheres to the sample tube.

  • Page 174: Cytoflex Ready To Use Daily Qc Fluorospheres Preparation Process

    Process. Take one sample tube and label it as the QC sample tube. Use the vortexer to thoroughly mix the bottle of CytoFLEX Ready to Use Daily QC Fluorospheres for two or three seconds. Add 10 drops of CytoFLEX Ready to Use Daily QC Fluorospheres to the sample tube.

  • Page 175: Preparing The Qc Sample [With Plate Loader]

    CytoFLEX Daily IR QC Fluorospheres Preparation Process. Take one 96-well plate and record the QC sample well position. Use the vortexer or shake vigorously to thoroughly mix the bottle of CytoFLEX Daily QC Fluorospheres. IMPORTANT Do not overfill the sample well.

  • Page 176: Cytoflex Ready To Use Daily Qc Fluorospheres Preparation Process

    Process. Take one 96-well plate and record the QC sample well position. Use the vortexer to thoroughly mix the bottle of CytoFLEX Ready to Use Daily QC Fluorospheres for two or three seconds. Add 3-4 drops of CytoFLEX Ready to Use Daily QC Fluorospheres to sample well.

  • Page 177: Importing Lot-Specific Target Values

    Instrument Quality Control and Standardization Quality Control Importing Lot-Specific Target Values Import lot-specific target values for each new lot of CytoFLEX QC Fluorospheres, CytoFLEX Ready to Use Daily QC Fluorospheres, or CytoFLEX Daily IR QC Fluorospheres. CAUTION Risk of erroneous QC results. Different target value information correspond to different lot numbers.
  • Page 178 Instrument Quality Control and Standardization Quality Control IMPORTANT The Beckman Coulter website may prompt you to select your Region and Country prior to the Beckman Coulter Technical Documents and Software page. Select . The Beckman Coulter Technical Documents and Software Download Target File Downloads page appears.
  • Page 179 Instrument Quality Control and Standardization Quality Control [CytoFLEX QC Fluorospheres Target] [CytoFLEX IR QC Fluorospheres Target] [CytoFLEX Ready to Use Daily QC Fluorospheres Target] Select Search The search results appear below the Search By Lot Number tab. B49006AS...
  • Page 180 Instrument Quality Control and Standardization Quality Control [CytoFLEX QC Fluorospheres Target] [CytoFLEX IR QC Fluorospheres Target] [CytoFLEX Ready to Use Daily QC Fluorospheres Target] 5-10 B49006AS...
  • Page 181 Software Name column. The CytoFLEX QC Fluorospheres Target Values CytoFLEX QC Fluorospheres Target Values page appears. Select under the correct lot number from the CytoFLEX QC Fluorospheres Target Download Values page. If the File Download pop up window appears, select and browse to the desired file path.

  • Page 182: 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 183 Importing Lot-Specific Target Values, then select the proper lot number. Select Initialize Insert the prepared QC sample tube (see CytoFLEX Ready to Use Daily QC Fluorospheres Preparation Process, or CytoFLEX Daily QC Fluorospheres Preparation Process) into the tube holder. Select to load the sample and begin to run the QC procedure.
  • Page 184 QC could fail up to 3 times upon running each new lot number for the first time until target gain values are established. If the lot number of CytoFLEX QC Fluorospheres is NOT new and QC fails, refer to Step CHAPTER 5, Confirming...

  • Page 185: Collecting Qc Data [With Plate Loader]

    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 186 Standardization, then select the proper lot number. Select Initialize Select Eject Insert the prepared QC well plate (see Preparation Process CytoFLEX Daily QC Fluorospheres CytoFLEX Ready to Use Daily QC Fluorospheres Preparation Process CytoFLEX Daily IR QC Fluorospheres Preparation Process) into the plate holder. 5-16 B49006AS...
  • Page 187 Instrument Quality Control and Standardization Quality Control Select . The Plate Settings window appears. IMPORTANT Ensure the well position on the plate matches the well position selected in the software. Select the appropriate QC well. 5-17 B49006AS...
  • Page 188 Instrument Quality Control and Standardization Quality Control Select the desired plate type from the Plate Type dropdown menu. NOTE The available plate types included in the dropdown menu depend on the settings selected in the Plate Library. Refer to Plate Type Library CHAPTER 2, Using the CytExpert Software.
  • Page 189 QC run fails to reach the required event flow rate. This is not considered a QC failure. If this situation occurs, increase the sample concentration by adding one drop of CytoFLEX Daily QC Fluorospheres to the sample tube or prepare a new tube of CytoFLEX Ready to Use Daily QC Fluorospheres and then perform the experiment.

  • Page 190: Confirming Results

    Instrument Quality Control and Standardization Quality Control If the lot number of CytoFLEX QC Fluorospheres is NOT new and QC fails, refer to Step CHAPTER 5, Confirming Results, or CHAPTER 10, Troubleshooting. If QC passes, proceed to Step 16. Run Daily Clean to remove any residual fluorosphere particles. Refer to...
  • Page 191 • The rCV must be within the range. NOTE The CytoFLEX Daily QC Fluorospheres / CytoFLEX Ready to Use Daily QC Fluorospheres / Daily IR QC Fluorospheres (808 Laser) rCV must meet the following criteria to pass: — The rCV for channels of the 405 nm, 488 nm, 561 nm, 638 nm lasers must be ≤5%.
  • Page 192 Instrument Quality Control and Standardization Quality Control 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 193 Instrument Quality Control and Standardization Quality Control Laser-filter combinations that are not part of a default configuration may not have a target value defined in the QC target file and therefore will not generate a result during QC. Use the Target Library in the QC screen of CytExpert Software to view the list of laser-filter combinations that have target values assigned within a target file and therefore will return a result for QC.
  • Page 194 Instrument Quality Control and Standardization Quality Control If QC fails, follow the procedure below: NOTE Select No if the Confirm window appears. a. Verify whether the beads used were within their shelf life and stored in accordance with the appropriate instruction manual. b.
  • Page 195 Instrument Quality Control and Standardization Quality Control IMPORTANT When there are multiple lots, select which lot to create the LJ charts from. Select LJ Chart Settings on the top of the LJ Chart screen. The LJ Chart Settings screen appears. 5-25 B49006AS...
  • Page 196 Instrument Quality Control and Standardization Quality Control Select the tab, and select the power and/or delay checkboxes for each laser as needed. Laser Select the tab, and select each channel checkbox as needed. Channels 5-26 B49006AS...

  • Page 197: Qc Result Manager

    Instrument Quality Control and Standardization Quality Control Select Apply 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 198: Standardization

    Preparing the Standardization Sample Use Beckman Coulter CytoFLEX Daily QC fluorospheres or CytoFLEX Ready to Use Daily QC Fluorospheres or any other reference material that is relevant for your application. 5-28...

  • Page 199: Required Materials

    Required Materials The following materials are required to complete the QC process: • CytoFLEX Daily QC Fluorospheres or CytoFLEX Ready to Use Daily QC Fluorospheres, or other material applicable for your application • CytoFLEX Daily IR QC Fluorospheres (for systems configured with an IR laser) •...
  • Page 200 Instrument Quality Control and Standardization Standardization Change the tube name. Refer to Changing the Tube Name CHAPTER 6, Data Acquisition and Sample Analysis. Select from the File menu to save the experiment. Save As Select to delete all the remaining tubes. 5-30 B49006AS...
  • Page 201 Instrument Quality Control and Standardization Standardization Select . The Compensation Matrix window appears. Select to clear the compensation matrix. The message Are you sure you want to clear the Clear compensation matrix? appears. Select Load the sample tube. NOTE The sample tube holder accommodates 1.5-mL, 2.0-mL, and 12 x 75 mm sample tubes. Select 5-31 B49006AS...
  • Page 202 Instrument Quality Control and Standardization Standardization View the plots and establish the gates. Refer to Creating Plots and Gates CHAPTER 6, Data Acquisition and Sample Analysis. NOTE Use the FSC channel as the trigger channel and select Automatic threshold. NOTE The threshold may need to be adjusted to visualize the QC beads populations.
  • Page 203 Instrument Quality Control and Standardization Standardization Right-click the table and select . The Statistics Setting window appears. Statistics Settings 5-33 B49006AS...
  • Page 204 Instrument Quality Control and Standardization Standardization Select the Population tab and select the relevant population for the tube. 5-34 B49006AS...
  • Page 205 Instrument Quality Control and Standardization Standardization Select the tab then select the Median Fluorescence value for all parameters used. Statistics NOTE The median values are the target settings that will be used for standardization. Right-click the statistics table and select Export tube to CSV file If Excel is not available, manually record all the median values or take a screen shot.

  • Page 206: Adding A New Standardization Item

    Instrument Quality Control and Standardization Standardization Save the experiment. NOTE Rerun the experiment if: • You change the standardization fluorosphere used. • The Lot number for the standardization fluorosphere is changed. Adding a New Standardization Item Select in the QC/Standardization menu to access the Standardization Start QC/Standardization screen.
  • Page 207 Instrument Quality Control and Standardization Standardization Select from the Settings menu. Standardization Target Library 5-37 B49006AS...
  • Page 208 Instrument Quality Control and Standardization Standardization Select . The Add Standardization Target Value window appears. Enter the Item, Lot No., and Expire date from the drop downs located at the top of the Add Standardization Target Value window. NOTE A single Lot No. can include several Items, but you cannot add duplicate Items under the same Lot No..
  • Page 209 Instrument Quality Control and Standardization Standardization Set the channels to be standardized. NOTE The contents of the channel, laser, and filter column come from the current detector configuration setting. Refer to Verifying, Selecting, Editing, and Creating Detector Configuration Data Acquisition and Sample Analysis.

  • Page 210: Performing The Standardization

    Instrument Quality Control and Standardization Standardization Performing the Standardization Select in the QC/Standardization menu to access the Standardization Start QC/Standardization screen. 5-40 B49006AS...
  • Page 211 Instrument Quality Control and Standardization Standardization Select the radio button. Standardization Select the correct Lot No. from the Lot No. dropdown. NOTE Ensure the Lot No. corresponds to the standardization sample that generated the target median values. Select the Items to be standardized. Select the proper detector configuration.
  • Page 212 Instrument Quality Control and Standardization Standardization Select Initialize Select The Process section of the screen displays the process details. Once the process is complete, the Standardization Report displays. NOTE The updated Standardization item is added to the Acquisition Catalog automatically and overwrites the previously existing standardized settings for this item.

  • Page 213: Applying The Standardized Acquisition Settings

    Instrument Quality Control and Standardization Standardization Select from the Cytometer menu to verify the gain settings. The Acq. Acq. Setting Catalog Setting Catalog window appears. NOTE designates test items from Standardization. Run Daily Clean. Refer to Daily Clean CHAPTER 11, Cleaning Procedures.
  • Page 214 Instrument Quality Control and Standardization Standardization Select from the Cytometer menu. The Acq. Setting window appears. Acq.Setting Select . The Acq. Setting Catalog window appears. Import from Catalog 5-44 B49006AS...
  • Page 215 Instrument Quality Control and Standardization Standardization Browse for the item to import and select Import. The standardized settings are applied to the sample tube. The Information window appears to notify of the corresponding channels with the changed gain as a result of the Standardization. 5-45 B49006AS...

  • Page 216: Standardization Target Library

    Instrument Quality Control and Standardization Standardization Select Standardization Target Library Select from the Settings menu. The standardization Target Library Standardization Target Library... window appears. NOTE The Item name displays in the Acquisition Setting Catalog window as the saved acquisition setting name.
  • Page 217 Instrument Quality Control and Standardization Standardization Select to exit the Standardization Target Library window. Exporting a Standardization Item Browse for the Item to export. The available items display on the left panel of the Standardization Target Library window. Select on the Standardization Target Library window. Navigate to the desired file path and select Save NOTE...
  • Page 218 Instrument Quality Control and Standardization Standardization Edit Item, Lot No., Expire date and the parameters for that item and select NOTE Perform a new standardization if the Lot No. of standardization sample is changed. Ensure the item parameters are correct then select and save the file.

  • Page 219: Chapter 6: Data Acquisition And Sample Analysis

    CHAPTER 6 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.
  • Page 220 Data Acquisition and Sample Analysis Creating an Experiment Verify the laser settings. Refer to Laser Settings CHAPTER 6, Data Acquisition and Sample Analysis. Create or open an experiment using one of the following methods: • Create a new experiment: — Select on the Start page, specify the file path, and save the experiment.

  • Page 221: Creating An Experiment [With Plate Loader]

    Data Acquisition and Sample Analysis Creating an Experiment 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. Then select If you are using the CytExpert Electronic Record Management Software Option: Select File >...
  • Page 222 Data Acquisition and Sample Analysis Creating an Experiment Select . The Plate window appears. [CytoFLEX LX Shown] B49006AS...
  • Page 223 Data Acquisition and Sample Analysis Creating an Experiment Select the Add Plate dropdown and select Add Plate [CytoFLEX LX Shown] B49006AS...

  • Page 224: Csv Template

    Data Acquisition and Sample Analysis Creating an Experiment 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. NOTE Select Add Plate from Layout to create a plate from a preset .csv file.
  • Page 225 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. Ensure you set the last sample wells with the appropriate number of cleaning agent wells and deionized water wells.
  • Page 226 Data Acquisition and Sample Analysis Creating an Experiment b. Select NOTE A gray background ( ) indicates auto shutdown is disabled. A yellow background ( indicates auto shutdown is enabled. NOTE Select Save Plate Template to save the acquisition settings as a template in SCTM format. Select Save Plate Layout to save the name, sample ID, or metadata as a layout in CSV format.

  • Page 227: Setting Sample Wells

    Once the plate protocol is created, the plate window appears. Refer to Figure 6.2. Figure 6.2 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. The number labeled at the right bottom shows the order of auto record.
  • Page 228 Data Acquisition and Sample Analysis Creating an Experiment Left-click and drag your mouse to highlight the desired wells or hold the Control key and select each desired well. Select or right-click the selected wells and select . The Set As Sample Set As Sample Wells Wells window appears.
  • Page 229 Data Acquisition and Sample Analysis Creating an Experiment 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. 6-11 B49006AS...
  • Page 230 Data Acquisition and Sample Analysis Creating an Experiment Select the channels and create label names under the Channel tab. Set compensation under the Compensation Matrix tab. Refer to CHAPTER 7, Compensation detailed instructions on setting compensation. 6-12 B49006AS...
  • Page 231 Select Events to record, time to record, or volume to record under the Stopping Rules tab. NOTE Beckman Coulter recommends setting an acquisition time limit to stop the acquisition if the event limit cannot be reached. Create the desired name and value under the Custom Metadata tab.

  • Page 232: Modifying Well Settings

    Data Acquisition and Sample Analysis Creating an Experiment Modifying Well Settings If any well settings require modification, right-click the sample well and select the setting to change. [CytoFLEX LX Shown] 6-14 B49006AS...
  • Page 233 Data Acquisition and Sample Analysis Creating an Experiment Applying Existing Well Settings to Additional Wells Right-click the well with the desired settings and select . The Apply Well Apply Well Settings to Settings of window appears. NOTE The Convert Compensation Matrix by the Gain of Tube checkbox is only available when Acq. Setting is deselected.

  • Page 234: Setting The Channel And Label

    Data Acquisition and Sample Analysis Creating an Experiment Right-click and select Copy 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 235 Data Acquisition and Sample Analysis Creating an Experiment In the Set Channel window, modify which channels are used and how they are displayed. a. Select the channel signal check box, then you can add the reagent name in the Label column.
  • Page 236 Data Acquisition and Sample Analysis Creating an Experiment d. If you only need to modify the label name, select in the Settings menu to make the Set Label 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 237: Creating Plots And Gates

    Data Acquisition and Sample Analysis Creating an Experiment Creating Plots and Gates IMPORTANT The maximum number of elements allowed in an experiment is 200. Elements include plots, statistics tables, and gate hierarchy tables. IMPORTANT The maximum number of gates allowed in an experiment is 200. Use the plotting controls (see Figure 2.1) in the plot area to create plots and gates and to...
  • Page 238 Data Acquisition and Sample Analysis Creating an Experiment 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 239 Data Acquisition and Sample Analysis Creating an Experiment 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. 6-21 B49006AS...
  • Page 240 Data Acquisition and Sample Analysis Creating an Experiment d. Select the tab, and select a channel with the required signal width. 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 from the drop-down Property menu.
  • Page 241 Data Acquisition and Sample Analysis Creating an Experiment 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 242 Data Acquisition and Sample Analysis Creating an Experiment • 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 243 Data Acquisition and Sample Analysis Creating an Experiment 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 244: All Gates - Example Experiment

    Data Acquisition and Sample Analysis Creating an Experiment 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 245 Data Acquisition and Sample Analysis Creating an Experiment b. If necessary, you can select the option from the drop-down menu to Combo Population 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 246: Creating And Adjusting Auto Gates

    Data Acquisition and Sample Analysis Creating an Experiment 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. To change the name of each gate, double-click the name of the desired gate.
  • Page 247 Data Acquisition and Sample Analysis Creating an Experiment 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 from the drop down menu.

  • Page 248: Turning Auto Recalculate On/Off

    Data Acquisition and Sample Analysis Creating an Experiment 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 249: Adjusting Autogate Movement And Extent

    Data Acquisition and Sample Analysis Creating an Experiment 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 250: Movement - Max Setting

    Data Acquisition and Sample Analysis Creating an Experiment Figure 6.6 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 251: Setting Customized Parameters

    Data Acquisition and Sample Analysis Creating an Experiment Figure 6.8 Extent - Maximum Setting Setting Customized Parameters Set custom parameter to create fluorescence calculations. Select from the Settings menu. Or, right-click a test tube from the Set Customized Parameter test tube menu and select .

  • Page 252: Setting Custom Statistics

    Data Acquisition and Sample Analysis Creating an Experiment The new parameter name is displayed in the list of parameters and statistic items. Setting Custom Statistics Set custom statistics to create calculations based on populations of interest. 6-34 B49006AS...
  • Page 253 Data Acquisition and Sample Analysis Creating an Experiment Right-click the statistics table and select . The Statistics Setting window Statistics Setting appears. Select Expression Select . The Expression window appears. Edit 6-35 B49006AS...
  • Page 254 Data Acquisition and Sample Analysis Creating an Experiment Enter the expression name in the Name section and enter the expression using the equation buttons. 6-36 B49006AS...
  • Page 255 Data Acquisition and Sample Analysis Creating an Experiment Select NOTE The equation populates in the Statistics Setting window under the Expression selection. 6-37 B49006AS...

  • Page 256: Configuring Acquisition Settings

    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 ID name in the Tube section of the screen and select or simply double-click the sample Edit Name tube or sample ID name.

  • Page 257: Laser Setting Window [Cytoflex Lx Shown]

    1. Enable/Disable: Enables or disables the laser. 2. Target Power (mW): Used to change the laser target power. NOTE Laser target power can only be adjusted on the CytoFLEX LX system. The power detector has ±1 mW tolerance. Refer to Table 6.1 for the target power ranges allowed in the Laser Setting screen.

  • Page 258: Setting Laser Target Power Settings [Cytoflex Lx Only]

    NOTE Lasers can only be enabled and disabled when the system is in standby mode. Setting Laser Target Power Settings [CytoFLEX LX Only] IMPORTANT When you change the laser target power, it will impact all of your results including QC and standardization.

  • Page 259: Adjusting The Gain

    Data Acquisition and Sample Analysis Configuring Acquisition Settings Standardize your laser target values in the QC Experiment. Refer to Standardization CHAPTER 5, 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 260 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. NOTE Optimize the gain settings according to your own experimental goals. The QC Gain values are only for reference.

  • Page 261: Adjusting The Threshold

    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 262: Setting Collection Conditions

    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 263 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 264: Setting Plot Display Conditions

    Data Acquisition and Sample Analysis Configuring Acquisition Settings Setting Plot Display Conditions Select in the Settings menu. The Events Display Setting window appears. Events Display Setting Three display options are available: Used to view all events on the plot. • Display all events.

  • Page 265: 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 12, Replacement/Adjustment...

  • Page 266: Verifying, Selecting, Editing, And Creating Detector Configuration

    Select the desired configuration. b. Select Set as Current A green checkmark appears in front of the selected configuration. [CytoFLEX Shown] NOTE A configuration is locked when appears to the left of a configuration. A configuration locks for two reasons: •...
  • Page 267 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 to access the Edit Detector Configuration screen. Edit b.
  • Page 268 Data Acquisition and Sample Analysis Load Sample and Record Data d. Ensure the new configuration is highlighted, then select . The Edit Detector Edit 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 269 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. Select Set As Current Select To delete a configuration created in error, select .

  • Page 270: Setting Up Violet Side Scatter (Vssc) Channel

    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 271 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 272 Data Acquisition and Sample Analysis Load Sample and Record Data Select the Default Configuration and select . The Configuration Save As window appears. Save As Name the new configuration VSSC and select Select the VSSC configuration and select . The Edit Detector Configuration window appears. Edit Change the filters and channel names according to the filter order in the violet WDM.

  • Page 273: Sampling And Collecting Data [Without Plate Loader]

    Data Acquisition and Sample Analysis Load Sample and Record Data Right-click the VSSC channel, and select to set it as the Violet SSC channel. Set SSC Select to save the changes and close the Edit Detector Configuration window. Select Set as Current Select to save the changes and close the Detector Configuration window.
  • Page 274 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 275 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: Saves the current tube and creates an additional tube.

  • Page 276: Sampling And Collecting Data [With Plate Loader]

    Data Acquisition and Sample Analysis Load Sample and Record Data Adds new data to the existing data. • Append data to existing file. 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: Creates a new tube in the test tube section of the screen for the data.
  • Page 277 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. [CytoFLEX LX Shown] NOTE Select to remove the auto record setting from the selected wells.

  • Page 278: Creating A Heat Map [With Plate Loader]

    Data Acquisition and Sample Analysis Load Sample and Record Data Creating a Heat Map [with Plate Loader] IMPORTANT The heat map function is only available in plate mode. Select from the Tube management controls (refer to Test Tubes CHAPTER 2, Using the CytExpert Software).
  • Page 279 Data Acquisition and Sample Analysis Load Sample and Record Data 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 280 Data Acquisition and Sample Analysis Load Sample and Record Data Select the desired color mode. NOTE Select Same setting for all parameters to use the same color for all parameters. Select Different settings per parameter to use different colors for different parameters. Select to add a parameter item to the Parameter section of the New Heat Map window.
  • Page 281 Data Acquisition and Sample Analysis Load Sample and Record Data Change the Parameter elements in the Parameter section of the New Heat Map window. a. Change the Label name if needed. b. Select from the Statistic Expression section of the Parameter list. The Expression window appears.
  • Page 282 Data Acquisition and Sample Analysis Load Sample and Record Data 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. All wells are included by default. NOTE designates that a well is excluded.
  • Page 283 Data Acquisition and Sample Analysis Load Sample and Record Data 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 284 Data Acquisition and Sample Analysis Load Sample and Record Data Select to assign colors based on a fixed range specified by the user. The heat Fixed Range map is created directly based on the result of the expression. The color of the heat map displays according to the legend range.

  • Page 285: Refreshing A Heat Map

    Data Acquisition and Sample Analysis Load Sample and Record Data Heat Map with 1 Parameter 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 286: Modifying Existing Heat Map Settings

    Data Acquisition and Sample Analysis Load Sample and Record Data Modifying Existing Heat Map Settings Select from the heat map toolbar to modify existing heat map settings. The Modify Heat Map Settings window appears. 6-68 B49006AS...

  • Page 287: Deleting An Existing Heat Map

    Data Acquisition and Sample Analysis Load Sample and Record Data 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 tool-bar.

  • Page 288: Analyzing And Exporting Data

    Data Acquisition and Sample Analysis Analyzing and Exporting Data Analyzing and Exporting Data Select the sample tube to be analyzed. 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 289 Data Acquisition and Sample Analysis Analyzing and Exporting Data Right-click the plot and select to make the display color of the specified Bring population to front gate appear in front of all other colors, or select to hide the display Send population to back color of the specified gate behind all other colors.
  • Page 290 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. 6-72 B49006AS...
  • Page 291 Data Acquisition and Sample Analysis Analyzing and Exporting Data The final generated plots appear as below. Right-click a plot and select from the drop-down Export to Clipboard Export to Graphic File menu to select an image to export. • copies the plot to the clipboard, allowing you to paste it directly into Export to Clipboard documents in common file formats.
  • Page 292 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 293: Exporting Fcs Files

    Data Acquisition and Sample Analysis Analyzing and Exporting Data Exporting FCS Files Exporting Single Tube Files Right-click the desired tube from the test tube section of the screen and select Export FCS File The Export FCS File window appears. Select the population from the Population dropdown menu. Select either Area Height...
  • Page 294 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 295: Exporting Plots Or The Statistics Table Of Multiple Tubes As Picture Files

    Data Acquisition and Sample Analysis Analyzing and Exporting Data Exporting Multiple FCS Files Select from the File menu. The Export FCS File window appears. Export FCS File Select the tubes to export. Repeat Steps from Exporting Single Tube Files. Exporting Plots or the Statistics Table of Multiple Tubes as Picture Files Right-click on the plot or statistics to export.

  • Page 296: Importing And Exporting Instrument Settings

    Data Acquisition and Sample Analysis Analyzing and Exporting Data Select . The Export Tubes to Graphic Files window appears. Export All Samples to Graphic File Select the desired tubes to export. Select the path. Select NOTE The plots of the selected tubes save as .bmp file. Importing and Exporting Instrument Settings The CytExpert software supports importing and exporting instrument settings to facilitate the experiment process.
  • Page 297 , locate the file with the required instrument settings, or select Import From File Import to import the instrument settings. From Catalog [CytoFLEX LX Shown] Select Close Exporting Instrument Settings Select the desired sample tube to export. Then select Select to export a current set of instrument settings, stored in a file ending in .acq.

  • Page 298: Importing And Exporting Compensation Settings

    Data Acquisition and Sample Analysis Analyzing and Exporting Data Importing and Exporting Compensation Settings The software supports unrestricted importing and exporting of compensation data, regardless of whether the sample tube data has already been acquired. Imported compensation values only cover channels identical with the current instrument configuration.
  • Page 299 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: • Used to access the Preview screen. Print Preview. — Select to select the required format of the file to be exported and to save the file in that format.
  • Page 300 Data Acquisition and Sample Analysis Analyzing and Exporting Data • Used to print data for multiple tubes. Batch Print. 1. Select . The Batch Print window appears. Batch Print 2. Select the tubes to print. 3. Select • Used to print a PDF of the data for multiple tubes. Batch Export to PDF File.

  • Page 301: Saving The Experiment

    Data Acquisition and Sample Analysis Saving the Experiment Saving the Experiment Selecting in the File menu allows you to save the experiment. Save Selecting and saving the experiment under a different name allows you to create a backup. Save As Selecting in the File menu allows you to save the experiment as a template.
  • Page 302 Data Acquisition and Sample Analysis Saving the Experiment 6-84 B49006AS...

  • Page 303: Compensation

    CHAPTER 7 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 304: Creating A Compensation Experiment

    Compensation Creating a Compensation Experiment Workflow: Apply Prepare Create Acquire and Calculate compensation Compensation compensation Ô 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 305 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 306: Preparing The Compensation Sample

    Using Control Samples to Generate the Compensation Matrix The design of CytoFLEX system makes Gain independent compensation possible. CytExpert software automatically recalculates the compensation matrix according to gain. Manual adjustment may be applied but not necessary because it may introduce inaccurate results.
  • Page 307 Compensation Creating a Compensation Experiment Defining the Negative Population Using Unstained Samples Confirm that the instrument has been initialized. Refer to Initializing the Instrument CHAPTER 4, Daily Startup. CAUTION Risk of erroneous results. Calculations based on excessively small volumes of sampled data can be inaccurate.
  • Page 308 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. Select to save the data. Record Running the Single Positive Control Samples Place the single positive tube in sample loading position (see Figure 1.12).

  • Page 309: Positive Population Selected From The Single-Stained Sample

    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 number of positive cells is comparatively low, increase the number of acquisition events to a suitable amount.

  • Page 310: Calculating Compensation Values

    Compensation Creating a Compensation Experiment NOTE Figure 7.4 shows an example of selecting both the positive and negative populations without an unstained sample. Figure 7.4 Positive and Negative Populations Without an Unstained Sample 1. Negative population 2. Positive population Select Record Repeat steps to acquire data from subsequent single positive sample tubes.
  • Page 311 Compensation Creating a Compensation Experiment Select or select in the Compensation menu to calculate the Compensation Calculation 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 312: Creating A Compensation Experiment [With Plate Loader]

    Compensation Creating a Compensation Experiment [With Plate Loader] Select to save the single color compensation values in the Save To Compensation Library 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 313 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 314 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 315: Preparing The Compensation Sample

    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 316: Creating The Compensation Matrix From Previously Acquired Data

    Compensation Creating the Compensation Matrix from Previously Acquired Data Select to save the single color compensation values in the Save To Compensation Library 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 317 Compensation Creating the Compensation Matrix from Previously Acquired Data Right-click on the appropriate test tube and select . Locate the corresponding Import FCS File 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 318: Adjusting Compensation

    Calculating Compensation Values. NOTE The CytoFLEX platform uses the positive-negative method to calculate compensation. Where S(k) denotes the real-time background noise being measured. Where B(k) denotes the background noise being filtered. where B(k+1) denotes the noise background base being filtered at the next point.
  • Page 319 Compensation Adjusting Compensation Both methods allow you to apply the gain independent compensation capability, i.e. importing the compensation values with or without recalculating the compensation matrix based on the gain settings. After opening the desired compensation file, the Import Compensation window appears. Select one of the following: •...

  • Page 320: Importing Compensation Settings From The Compensation Library

    Files available in the compensation library are configuration-specific. The compensation library only displays the files created under the current default configuration. Select to select which compensation values to import from Import From Compensation Library the compensation library. [CytoFLEX LX Shown] 7-18 B49006AS...

  • Page 321: Exporting Compensation Settings

    Compensation Adjusting Compensation 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. Select to import the compensation values. Exporting Compensation Settings Select the desired sample tube to export.
  • Page 322 Compensation Adjusting Compensation [CytoFLEX LX Shown] Select Save NOTE The generated file ends in .comp. 7-20 B49006AS...

  • Page 323: Managing The Compensation Library

    Compensation Adjusting Compensation Managing the Compensation Library Compensation values can be managed in the Compensation Library. Select from the Settings menu. The Compensation Library window Compensation Library 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 324: Adding Channels For Compensation

    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 in the Compensation menu.

  • Page 325: Chapter 8: Data Review

    Exporting Results Copying Experiments and Importing Data Copying a Previously Acquired Experiment Experiments acquired by other CytoFLEX instruments using CytExpert software can be imported to your computer for analysis, provided your computer also uses CytExpert software. Select from the Start page or select...
  • Page 326 Data Review Copying Experiments and Importing Data In the new or opened experiment, select in the File menu to import the data Import FCS File 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 327: 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 328 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 329: 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 under the histogram icon drop-down list to create a new multi-data Histogram Overlay histogram.
  • Page 330 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 331: 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...

  • Page 332: Adjusting Compensation Settings

    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 Data compensation can be carried out at any time. You can select the desired tube in the tube list...

  • Page 333: Exporting Results

    Data Review Exporting Results Exporting Results Refer to CHAPTER 6, Data Acquisition and Sample Analysis. B49006AS...
  • Page 334 Data Review Exporting Results 8-10 B49006AS...

  • Page 335: Chapter 9: Daily Shutdown

    CHAPTER 9 Daily Shutdown Overview This chapter provides procedures for shutting down the CytoFLEX instrument. Workflow: Prepare the cleaning solution Clean the instrument Turn the instrument off Ô Ô This chapter contains information on: Preparing the Cleaning Solution • •...

  • Page 336: 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. Select Standby Exit the software. Optional: Turn the computer off. Optional: Turn the Cytometer's main power switch off. If there are any spills, clean the sample station. Refer to...

  • Page 337: Chapter 10: 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 338: Precautions/Hazards

    Laser Beam Hazards The CytoFLEX Platform flow cytometer can be configured with up to 6 solid-state diode lasers that are capable of producing laser light at the following levels: • 355-nm, 20-mW solid-state diode laser • 375-nm, 60-mW solid-state diode laser •...

  • Page 339: Laser Warning Labels

    If necessary, a cover with a CDRH-approved or IEC compliant label must be removed by a qualified Beckman Coulter Representative only. Refer to the following figures for the locations of the CDRH-approved and IEC compliant labels: Figure 10.1...

  • Page 340: Laser Warning Label On The Laser Optical Bench [Cytoflex]

    Troubleshooting Precautions/Hazards Figure 10.2 Laser Warning Label on the Laser Optical Bench [ CytoFLEX LX Figure 10.3 Laser Warning Label within the Optical Bench (Located Inside the Cytometer) [ CytoFLEX and CytoFLEX S CAUTION CLASS 3B LASER RADIATION WHEN OPEN AND INTERLOCKS DEFEATED...

  • Page 341: Laser Warning Label On The 355-Nm Laser [Cytoflex Lx]

    Troubleshooting Precautions/Hazards Figure 10.5 Laser Warning Label on the 355-nm Laser [ CytoFLEX LX Figure 10.6 Laser Warning Labels on the Cytometer Back Cover [ CytoFLEX and CytoFLEX S 10-5 B49006AS...

  • Page 342: Hazard Labels And Locations

    Troubleshooting Hazard Labels and Locations Figure 10.7 Laser Warning Labels on the Cytometer Back Cover [ CytoFLEX LX CLASS 1 LASER PRODUCT PRODUIT LASER CLASSE 1 Hazard Labels and Locations Carefully read the hazard warning labels on the instrument. The hazard labels are located on the instrument as indicated.

  • Page 343: Electrical Shock Hazard Label And Location

    A nonionic, non-flourescent, and azide free sheath fluid for use on Beckman Coulter CytoFLEX flow cytometers. REACTIVE INGREDIENTS: Anti-microbial compound Figure 10.10 Biohazard Label Located in the Sample Station and on the Back of the Cytometer [CytoFLEX Shown] Electrical Shock Hazard Label and Location Figure 10.11 Electrical Shock Hazard Label by the Power Switch [CytoFLEX or CytoFLEX S]...

  • Page 344: Caution Labels And Location

    Troubleshooting Disposal Precaution Caution Labels and Location Figure 10.12 Caution Labels [CytoFLEX or CytoFLEX S] Plate Loader Hazard Labels and Location Figure 10.13 Plate Loader Hazard Labels Disposal Precaution 10-8 B49006AS...
  • Page 345 Troubleshooting Table Table 10.1 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.”...
  • Page 346 Troubleshooting Troubleshooting Table Table 10.1 Troubleshooting (Continued) Problem Probable Cause Corrective Action The connection indicator • Data connection error 1. Ensure that the USB data cable is light in the lower left securely connected to the back of the • The Cytometer is not turned corner of the software Cytometer and the back of the screen is red and...
  • Page 347 Troubleshooting Troubleshooting Table Table 10.1 Troubleshooting (Continued) Problem Probable Cause Corrective Action The fluid status • Instrument data connection 1. Ensure the sheath fluid/waste harnesses information displays red error. are secured to the correct container. for Sheath and/or Waste • The sensor connection is not 2.
  • Page 348 Troubleshooting Troubleshooting Table Table 10.1 Troubleshooting (Continued) Problem Probable Cause Corrective Action The sampling flow rate is • The threshold setting is too 1. Use the manual threshold setting to too fast. low. increase the threshold. Refer to Adjusting the Threshold CHAPTER 6, •...

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