BD FACSCelesta Flow Cytometer User manual

23-17147-02
9/2020
For Research Use Only
Becton, Dickinson and Company
BD Biosciences
2350 Qume Drive
San Jose, CA 95131 USA
bdbiosciences.com
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BD Biosciences
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BD FACSCelesta™ Flow
Cytometer
User’s Guide

Copyrights
© 2020, Becton, Dickinson and Company. All rights reserved. No part of this publication may be reproduced,
transmitted, transcribed, stored in retrieval systems, or translated into any language or computer language, in
any form or by any means: electronic, mechanical, magnetic, optical, chemical, manual, or otherwise, without
prior written permission from BD Biosciences.
The information in this guide is subject to change without notice. BD Biosciences reserves the right to change
its products and services at any time to incorporate the latest technological developments. Although this guide
has been prepared with every precaution to ensure accuracy, BD Biosciences assumes no liability for any errors
or omissions, nor for any damages resulting from the application or use of this information. BD Biosciences
welcomes customer input on corrections and suggestions for improvement.
Trademarks
Clorox is a registered trademark of The Clorox Company.
SPHERO is a trademark of Spherotech, Inc.
Microsoft and Windows are registered trademarks of Microsoft Corporation.
Texas Red®and Alexa Fluor®are registered trademarks of Life Technologies Corporation.
Teflon is a registered trademark of E.I. du Pont de Nemours and Company.
Living Colors®(DsRed, mCherry) is a registered trademark of Clontech.
Cy™ is a trademark of GE Healthcare. Cy™ dyes are subject to proprietary rights of GE Healthcare and
Carnegie Mellon University, and are made and sold under license from GE Healthcare only for research and in
vitro diagnostic use. Any other use requires a commercial sublicense from GE Healthcare, 800 Centennial
Avenue, Piscataway, NJ 08855-1327, USA.
Falcon®is a registered trademark of Corning Incorporated.
CF™ is a trademark of Biotium, Inc.
BD, the BD Logo, FACS, FACSCelesta, FACSClean, FACSDiva and FACSFlow are trademarks of Becton,
Dickinson and Company or its affiliates. All other trademarks are the property of their respective owners.
© 2020 BD. All rights reserved.
Patents
APC-Cy7: US 5,714,386
Regulatory information
For Research Use Only. Not for use in diagnostic or therapeutic procedures.
Class 1 Laser Product.

FCC information
WARNING: Changes or modifications to this unit not expressly approved by the party responsible for
compliance could void the user’s authority to operate the equipment.
NOTICE: This equipment has been tested and found to comply with the limits for a Class A digital device,
pursuant to Part 15 of the FCC Rules. These limits are designed to provide reasonable protection against
harmful interference when the equipment is operated in a commercial environment. This equipment generates,
uses, and can radiate radio frequency energy and, if not installed and used in accordance with the instruction
manual, may cause harmful interference to radio communications. Operation of this equipment in a
residential area is likely to cause harmful interference in which case the user will be required to correct the
interference at his or her own expense. Shielded cables must be used with this unit to ensure compliance with
the Class A FCC limits. This Class A digital apparatus meets all requirements of the Canadian Interference-
Causing Equipment Regulations. Cet appareil numérique de la classe A respecte toutes les exigences du
Réglement sur le matériel brouilleur du Canada.
Compliance information
NOTICE: This laboratory equipment has been tested and found to comply with the EMC and the Low
Voltage Directives. This includes FCC, Part 15 compliance for a Class A Digital Device.
CAUTION: Any unauthorized modifications to this laboratory equipment may affect the Regulatory
Compliance items stated above.
History
Revision Date Change made
23-17147-00 11/2015 Initial release
23-17147-01 6/2016 Updated the guide by removing status screen image and air
pressure definition.
23-17147-02 9/2020 Removed references to FACSRinse and added BD Detergent
Solution Concentrate in place of FACSRinse.

Contents
Chapter 1: About this guide 9
What this guide covers . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 10
Conventions . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 11
About the BD FACSCelesta documentation . . . . . . . . . . . . . . . . . . . . . . . . . . . . 11
Instrument technical support . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 13
Chapter 2: Introduction 15
System overview . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 16
Cytometer overview . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 17
Control panel . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 19
Fluidics system . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 20
Sheath and waste containers . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 27
Optics . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 28
Workstation . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 30
Chapter 3: Cytometer setup 31
Starting the cytometer and computer . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 32
Preparing the sheath container . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 33
Removing air bubbles . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 35
Preparing the waste container . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 38
Priming the fluidics . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 40
About the optical filters and mirrors . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 41
Custom configurations and baselines . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 43

BD FACSCelesta Flow Cytometer User’s Guidevi
Chapter 4: Maintenance 45
Maintenance overview . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 46
Cleaning the fluidics . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 47
Shutting down the cytometer . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 49
Flushing the system . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 50
Replacing the waste container cap . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 52
Changing the sheath filter . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 54
Changing the Bal seal . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 56
Changing the sample tube O-ring . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 58
Cleaning or replacing the sheath gasket . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 60
Chapter 5: Optimizing cytometer settings 61
Cytometer settings workflow . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 62
Verifying the configuration and user preferences . . . . . . . . . . . . . . . . . . . . . . . . . 64
Running a performance check . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 67
Setting up an experiment . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 72
Creating application settings . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 77
Recording compensation controls . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 80
Calculating compensation . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 83
Chapter 6: Recording and analyzing data 85
Data recording and analysis workflow . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 86
Preparing the workspace . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 87
Recording data . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 88
Analyzing data . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 91
Reusing an analysis . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 97
Chapter 7: Technical overview 99
About fluidics . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 100
About optics . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 101
About electronics . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 112

Contents vii
Chapter 8: Troubleshooting 117
Cytometer troubleshooting . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 118
Electronics troubleshooting . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 126
Chapter 9: Detector array configurations 127
Fluorescence spectra . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 128
About configuration maps . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 131
About the configuration . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 132
Base configuration polygon maps . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 134
Chapter 10: Supplies and consumables 143
Ordering information . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 144
Beads . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 144
Reagents . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 145
Equipment . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 146
Index 147

1
About this guide
This chapter covers the following topics:
•What this guide covers (page 10)
•Conventions (page 11)
•About the BD FACSCelesta documentation (page 11)
•Instrument technical support (page 13)

BD FACSCelesta Flow Cytometer User’s Guide10
What this guide covers
This guide describes the procedures necessary to operate and
maintain your BD FACSCelesta™ flow cytometer. Because many
cytometer functions are controlled by BD FACSDiva™ software,
this guide also contains information about software features
required for basic cytometer setup and operation.
This guide assumes you have a working knowledge of basic
Microsoft® Windows® operation. If you are not familiar with the
Windows operating system, see the documentation provided with
your computer.

Chapter 1: About this guide 11
Conventions
Introduction The following table lists the safety symbols used in this guide to
alert you to potential hazards.
Safety symbols
About the BD FACSCelesta documentation
Introduction This topic describes the documentation available with the
BD FACSCelesta flow cytometer.
Publication formats This guide is provided in PDF format to provide an eco-friendly
option. All content is also included in the BD FACSDiva software
Help.
Help system The help system installed with BD FACSDiva software includes all
content from this guide and the documents listed below. Access the
BD FACSCelesta help system from the Help menu in the
Symbol Meaning
Caution! Identifies a hazard or unsafe practice that
could result in data loss, material damage, minor
injury, severe injury, or death
Biological hazard
Electrical hazard
Laser hazard

BD FACSCelesta Flow Cytometer User’s Guide12
BD FACSDiva software. Internet access is not required to use the
help system.
The help system is compiled from the following documents:
•BD FACSDiva Software Reference Manual: Includes
instructions or descriptions for installation and setup,
workspace components, acquisition controls, analysis tools,
and data management. Access this manual from the
BD FACSDiva Software Help menu (Help > Documentation >
Reference Manual), or by double-clicking the shortcut on the
desktop.
•BD FACSCelesta Flow Cytometer Site Preparation Guide:
Contains specifications for:
– Cytometer weight and size
– Temperature
– Electrical requirements
•BD High Throughput Sampler User’s Guide: Describes how to
set up and operate the BD™ High Throughput Sampler (HTS)
option. It also contains a description of BD FACSDiva
software features specific to the HTS.
•BD FACSFlow™ Supply System User’s Guide: Describes the
optional automated sheath and waste fluid control system.
Additional
Documentation
BD Cytometer Setup and Tracking Application Guide: Describes
how to use the BD™ Cytometer Setup and Tracking (CS&T)
features in BD FACSDiva software. You can find the document in
the following location bdbiosciences.com/guide.

Chapter 1: About this guide 13
Instrument technical support
Introduction This topic describes how to get technical assistance.
Contacting
technical support
If technical assistance is required, contact your local
BD Biosciences customer support representative or supplier.
When contacting BD Biosciences, have the following information
available:
• Product name, part number, and serial number
• Version of BD FACSDiva software you are using
• Any error messages
• Details of recent system performance
To contact customer support:
1. Go to bdbiosciences.com.
2. Select your region. You will see information in your local
language.
3. Click Go.
4. Click the Support link for details for your local region.

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2
Introduction
This chapter covers the following topics:
•System overview (page 16)
•Control panel (page 19)
•Fluidics system (page 20)
•Sheath and waste containers (page 27)
•Optics (page 28)
•Workstation (page 30)

BD FACSCelesta Flow Cytometer User’s Guide16
System overview
About the system The BD FACSCelesta system includes the BD FACSCelesta flow
cytometer, BD FACSDiva software v8.0.1.1 running on the system
workstation, the optional BD FACSFlow™ supply system (FFSS),
and the optional BD High Throughput Sampler (HTS). Each
component is described in detail in the following sections.
12 3
Number Components
1 Sheath and waste tanks
2 BD FACSCelesta flow cytometer
3 Computer workstation

Chapter 2: Introduction 17
Cytometer overview
Introduction The BD FACSCelesta flow cytometer is an air-cooled multi-laser
benchtop flow cytometer with the ability to acquire 10–12
fluorescent parameters. It uses fixed-alignment lasers that transmit
light through a flow cell to collect and translate the resulting
fluorescence signals into electronic signals. Cytometer electronics
convert these signals into digital data.
Components The following figure shows the main components of the
BD FACSCelesta flow cytometer, which are listed in the table. Each
component is described in detail in the following sections.

BD FACSCelesta Flow Cytometer User’s Guide18
8
23
6
4
5
9
7
1
Number Component
1 Heat ventilation slots (top)
2 Control panel
3 Power button
4 Electrical plug
5 Fluidic sensor ports
6 Sample injection port (SIP)
7 Heat ventilation slots (left side)
8 Air and fluidic ports
9 Optics access door (polygon
detector arrays)

Chapter 2: Introduction 19
Control panel
Overview The following figure shows the components in the control panel,
which are listed in the table.
Caution! Do not place any objects on top of the
instrument. Blocking the ventilation may cause the
instrument to overheat.
Caution: Electrical Hazard! Do not place liquids on top of
the instrument. Any spill of liquid into the ventilation
openings could cause electrical shock or damage to the
instrument.
12
346
5
Number Component
1 System indicators
2 Fluid control buttons
3 Sample flow rate buttons

BD FACSCelesta Flow Cytometer User’s Guide20
More information •Fluidics system (page 20)
•Optics (page 28)
Fluidics system
Introduction The fluidics system carries the sample out of the sample tube and
into the interrogation region of the flow cell. Cells are carried in
the sample core stream in single file and measured individually.
System indicators There are two system indicators (System Status and Activity) on
the control panel.
•System Status. Shows the status of the sheath and waste tank
levels. The following table describes the LED indicators,
conditions that trigger them, and any action that must be
taken.
4 Sample fine adjust buttons
5 Status screen
6 MODE button
Number Component
LED color Status Action Condition
Green Good None Good
Yellow Caution! Sheath
and/or waste tanks
need attention.
Check tank levels Sheath low and/or
waste nearly full
Red Caution! Take
immediate action.
Empty waste tank
Fill sheath tank
Sheath empty and/or
waste full