Thermo Scientific Applied Biosystems QuantStudio Manuale utente

For Research Use Only. Not for use in diagnostic procedures.

QuantStudio™3D Digital PCR System

USER GUIDE

for use with:

QuantStudio™3D Digital PCR Instrument

ProFlex™2 × Flat PCR System or Dual Flat Block GeneAmp™PCR

System 9700

QuantStudio™3D Digital PCR 20K Chip v2

QuantStudio™3D Digital PCR Master Mix v2

QuantStudio™3D Digital PCR Chip Loader

Catalog Number A29154

Publication Number MAN0007720

Revision E.0

Manufacturer:

Life Technologies Holdings Pte Ltd |

Block 33 |

Marsiling Industrial Estate Road 3 |

#07-06, Singapore 739256

Products:

QuantStudio™3D Digital PCR Instrument

ProFlex™2 × Flat PCR System

Dual Flat Block GeneAmp™PCR System 9700

QuantStudio™3D Digital PCR Chip Loader

Manufacturer:

Life Technologies Corporation |

6055 Sunol Blvd |

Pleasanton, CA 94566

Products:

QuantStudio™3D Digital PCR 20K Chip v2

Life Technologies Corporation |

7335 Executive Way | Frederick, MD

21704 | USA

Products:

QuantStudio™3D Digital PCR Master Mix v2

For descriptions of symbols on product labels or product documents, go to thermoﬁsher.com/symbols-deﬁnition.

The information in this guide is subject to change without notice.

DISCLAIMER: TO THE EXTENT ALLOWED BY LAW, THERMO FISHER SCIENTIFIC INC. AND/OR ITS AFFILIATE(S) WILL NOT BE

LIABLE FOR SPECIAL, INCIDENTAL, INDIRECT, PUNITIVE, MULTIPLE, OR CONSEQUENTIAL DAMAGES IN CONNECTION WITH OR

ARISING FROM THIS DOCUMENT, INCLUDING YOUR USE OF IT.

Revision history: Pub. No. MAN0007720

Revision Date Description

E.0 18 March 2020 Removed the v1 chip and master mix. Updated template and legal

information. Updated “Prepare the dPCR reaction mix for each sample”

on page 23. Updated thermal cycling parameters. Reformatted “Install

the GeneAmp™PCR System 9700” on page 73.

D.0 15 July 2015 Updated for v2 chips, lids, and master mix. Also updated with changes

to instrument rmware, including adjusting for the different well volumes

between v1 and v2 chips. Includes improvements to the chip loading and

imaging workows.

C.0 February 2015 Updated for rmware and software revisions, including changes to

instrument networking, thermal cycling, chip analysis, maintenance,

software access, and computer requirements. Updated to latest

corporate boilerplate. Updated general sample and chip preparation,

system installation, troubleshooting, and parts and materials.

B.0 April 2014 Added support for the ProFlex™PCR System and revised the chip

loading instructions.

A.0 December 2013 Updated the manual chip preparation and added procedures for chip

preparation using the QuantStudio™3D Digital PCR Chip Loader and

wireless network installation.

02 June 2013 Updated general chip preparation and instrument networking.

01 March 2013 Initial version

Important Licensing Information: These products may be covered by one or more Limited Use Label Licenses. By use of these

products, you accept the terms and conditions of all applicable Limited Use Label Licenses.

Trademarks: All trademarks are the property of Thermo Fisher Scientiﬁc and its subsidiaries unless otherwise speciﬁed. TaqMan is a

registered trademark of Roche Molecular Systems, Inc., used under permission and license.

Contents

■CHAPTER 1 Product information ....................................... 8

About the QuantStudio™3D Digital PCR System .................................... 8

Instruments, kits, consumables, and accessories ................................... 9

Required materials not supplied ................................................. 10

Power line regulator ........................................................ 12

Surge protector ............................................................ 12

About the instrument ........................................................... 12

Data collection ............................................................ 13

Supported dyes ........................................................... 13

About the software ............................................................. 13

System requirements ....................................................... 13

About the chip ................................................................. 14

■CHAPTER 2 Getting Started ........................................... 15

Operational workow ........................................................... 16

Connect the QuantStudio™3D Digital PCR Instrument to a network .................. 17

Connect the QuantStudio™3D AnalysisSuite™Software ............................ 17

Choosing a thermal cycler ....................................................... 17

Compatible thermal cyclers ................................................. 17

■CHAPTER 3 Prepare the digital PCR reaction and load

the chips .................................................................. 19

Guidelines for loading and sealing chips .......................................... 19

Prepare the DNA samples ....................................................... 20

Quality of DNA ............................................................ 20

Quantity of DNA ........................................................... 20

Sample dilution ............................................................ 21

QuantStudio™3D Digital PCR System User Guide 3

Determine the optimal dilution when the target is known ........................ 21

Determine the optimal dilution when the target is unknown ...................... 22

Prepare the digital PCR reactions ................................................ 22

Required materials ......................................................... 22

Guidelines for dPCR sample preparation ..................................... 22

Recommended DNA volume and concentration ............................... 23

Prepare the dPCR reaction mix for each sample ............................... 23

Load the chips using the QuantStudio™3D Digital PCR Chip Loader ................. 24

Materials required ......................................................... 24

Chip loader status light ..................................................... 25

Prepare the chip loading workspace ......................................... 25

Prepare the syringe containing the Immersion Fluid ............................ 26

Load and seal the chips .................................................... 27

Fill the chip case with Immersion Fluid ....................................... 30

Seal the chip case with the label ............................................. 31

■CHAPTER 4 Thermal cycle the chips ................................. 32

Materials required .............................................................. 32

Prepare the thermal cycler ...................................................... 32

Guidelines for handling loaded chips ............................................. 35

Perform the PCR ............................................................... 36

Unload the thermal cycler ....................................................... 38

■CHAPTER 5 Image and analyze the chips ........................... 40

About imaging and primary analysis .............................................. 40

QuantStudio™3D System data ow .......................................... 40

About the instrument interface ................................................... 41

Chip imaging .................................................................. 42

Materials required ......................................................... 42

Guidelines for imaging multiple chips ......................................... 42

Specify the data destination ................................................. 42

Image the chips ........................................................... 43

Primary analysis ............................................................... 46

About imaging data and primary analysis ..................................... 46

About the results screen .................................................... 47

Data quality ags .......................................................... 48

About data les ............................................................ 48

Transfer the results ......................................................... 49

Using the QuantStudio™3D AnalysisSuite™Software .............................. 51

Access the software from a Connect cloud-based platform account ............. 51

Access the software from the server ......................................... 51

Obtain information from the Help system ..................................... 53

Contents

4QuantStudio™3D Digital PCR System User Guide

■CHAPTER 6 Troubleshooting .......................................... 54

Troubleshooting chip images using the Chip View .................................. 54

■APPENDIX A Install the QuantStudio™3D Digital PCR System .... 61

Time required for installation ..................................................... 61

Materials required .............................................................. 61

Before you begin ............................................................... 61

Plan the laboratory layout ....................................................... 62

Choose the instrument locations ................................................. 62

Install the QuantStudio™3D Digital PCR Instrument ................................ 63

Install the ProFlex™2 × Flat PCR System ......................................... 68

Unpack and place the ProFlex™PCR System Base Module ..................... 68

Prepare the workspace and instrument ....................................... 69

Install the Tilt Base ......................................................... 69

Unpack and install the sample block module .................................. 71

Set up the ProFlex™PCR System ............................................ 71

Install the GeneAmp™PCR System 9700 .......................................... 73

Unpack and place the GeneAmp™PCR System 9700 Base Module .............. 73

Unpack and install the Dual Flat Block Module ................................ 74

Install the Tilt Base and Chip Adapter ........................................ 75

Set up the GeneAmp™PCR System 9700 ..................................... 75

■APPENDIX B Specications and Layout ............................. 79

Component dimensions and weights ............................................. 79

QuantStudio™3D Digital PCR Instrument layout and connections .................... 80

Instrument clearances .......................................................... 80

Electrical specications ......................................................... 81

Environmental requirements ..................................................... 82

Network requirements .......................................................... 83

■APPENDIX C Maintenance ............................................ 84

Cleaning the chip tray and sample block .......................................... 84

Materials required ......................................................... 84

Clean the chip tray or sample block .......................................... 85

Congure the instrument settings ................................................ 86

(Optional) Set the instrument name .......................................... 86

Set the date and time ...................................................... 87

Check the instrument rmware .................................................. 87

Update the instrument rmware ............................................. 88

Returning your instrument for service ............................................. 89

Contents

QuantStudio™3D Digital PCR System User Guide 5

Replace the instrument fuses .................................................... 90

Materials required ......................................................... 90

Replace the fuses .......................................................... 90

Calibrate the touchscreen ....................................................... 91

View the instrument log ......................................................... 92

■APPENDIX D Networking .............................................. 94

Networking overview ........................................................... 94

About the network port and wireless adaptor .................................. 94

Networking guidelines and best practices ..................................... 95

Connect the instrument to the network ........................................... 95

Network setup workow .................................................... 96

Materials required ......................................................... 96

Required network information ............................................... 96

Connect the QuantStudio™3D Digital PCR Instrument to a network .............. 98

Create a network destination to receive run data ............................. 103

■APPENDIX E Load chips manually .................................. 106

Guidelines for loading and sealing chips ......................................... 106

Materials required ............................................................. 107

Prepare the syringe containing the Immersion Fluid ............................... 107

Load and seal the chips manually ............................................... 108

Fill the chip case with Immersion Fluid ........................................... 111

Seal the chip case with the label ................................................ 112

■APPENDIX F Safety ................................................... 113

Symbols on this instrument .................................................... 113

Conformity symbols ...................................................... 114

Safety alerts on this instrument ................................................. 115

Location of safety alerts on the instrument ................................... 115

Safety information for instruments not manufactured by Thermo Fisher Scientic ..... 116

Instrument safety ............................................................. 116

General ................................................................. 116

Electrical safety .......................................................... 117

Cleaning and decontamination ............................................. 118

Safety and electromagnetic compatibility (EMC) standards ......................... 118

Safety ................................................................... 119

EMC .................................................................... 119

Environmental design ..................................................... 119

Chemical safety .............................................................. 120

Biological hazard safety ....................................................... 122

Contents

6QuantStudio™3D Digital PCR System User Guide

■Documentation and support ........................................... 123

Related documentation ........................................................ 123

Customer and technical support ................................................ 123

Limited product warranty ...................................................... 124

Contents

QuantStudio™3D Digital PCR System User Guide 7

Product information

About the QuantStudio™3D Digital PCR System

The QuantStudio™3D Digital PCR System provides quantitative and qualitative

detection of target nucleic acid sequences (targets) using post-PCR (endpoint)

analysis.

Use the QuantStudio™3D Digital PCR Instrument to perform imaging and preliminary

analysis of a QuantStudio™3D Digital PCR 20K Chip v2. The chip must have been

loaded with uorescent-labeled quantitative PCR reagents (TaqMan™probe-based

assays or SYBR™Green primer-based assays) and thermal cycled using a ProFlex™

2 × Flat PCR System (or Dual Flat Block GeneAmp™PCR System 9700). The

QuantStudio™3D AnalysisSuite™Software performs subsequent analysis and post-

processing to yield relative or absolute quantication results from the raw imaging

data.

212

Figure 1 Components of the QuantStudio™3D Digital PCR System

1QuantStudio™3D Digital PCR Thermal Pads

2QuantStudio™3D Digital PCR Chip Adapters

3ProFlex™2 × Flat PCR System (or Dual Flat Block GeneAmp™PCR System 9700)

4QuantStudio™3D Tilt Base

5QuantStudio™3D Digital PCR Instrument

6QuantStudio™3D Digital PCR Chip Lid v2

7QuantStudio™3D Digital PCR Sample Loading Blade

8QuantStudio™3D Digital PCR 20K Chip v2

9(Optional) QuantStudio™3D Digital PCR Chip Loader

10 Immersion Fluid syringe

11 QuantStudio™3D Digital PCR Master Mix v2

12 Fluorescent-labeled quantitative PCR reagents (TaqMan™Assays or SYBR™Green primers)

13 (Not shown) QuantStudio™3D AnalysisSuite™Software

8QuantStudio™3D Digital PCR System User Guide

Instruments, kits, consumables, and accessories

The following tables describe the products covered in this user guide.

Instrument system

Item Catalog No. Amount

QuantStudio™3D Digital PCR System package

with v2 chips and master mix, includes:

•QuantStudio™3D Digital PCR Instrument

•QuantStudio™3D Digital PCR Chip Loader

•QuantStudio™3D Digital PCR 20K Chips v2

•QuantStudio™3D Digital PCR Master Mix

•ProFlex™2 × Flat PCR System

A29154 1 system

QuantStudio™3D Digital PCR Instrument with

power cord

4489084 1 instrument

QuantStudio™3D Digital PCR Tilt Base for

ProFlex™Thermal Cycler

A24898 1 base

QuantStudio™3D Digital PCR Tilt Base for

GeneAmp™PCR System 9700

4486414 1 base

QuantStudio™3D Digital PCR Chip Adapter Kit

for Flat Block Thermal Cycler

4485513 1 adapter

QuantStudio™3D Extended Arm WiFi Adapter A28598 1 wireless adapter

QuantStudio™3D Digital PCR 20K Chip Kit v2

and Master Mix v2, includes:

•8 × 12 packs of v2 chips, chip lids, and

QuantStudio™3D Digital PCR Sample

Loading Blades

•24 syringes of Immersion Fluid, plus tips

•1.5 mL of QuantStudio™3D Digital PCR

Master Mix v2

A26317 1 kit (96 chips)

QuantStudio™3D Digital PCR 20K Chip Kit v2,

includes:

•12 v2 chips, chip lids, and

QuantStudio™3D Digital PCR Sample

Loading Blades

•3 syringes of Immersion Fluid, plus tips

A26316 1 kit (12 chips)

QuantStudio™3D Digital PCR Immersion Fluid,

1.6 mL per syringe

A27322 3 syringes

Chapter 1 Product information

Instruments, kits, consumables, and accessories 1

QuantStudio™3D Digital PCR System User Guide 9

Instrument system

Item Catalog No. Amount

QuantStudio™3D Digital PCR Chip Loader 4482592 1 loader

QuantStudio™3D Digital PCR Chip Lids v2 and

QuantStudio™3D Digital PCR Sample Loading

Blades

A29247 12 each

QuantStudio™3D Digital PCR Master Mix v2 A26358

A26359

1.5 mL

5.0 mL

QuantStudio™3D Digital PCR Reagent Kit,

includes:

•Assays and reagents for system installation

and verication, or training of operators on

the use of the system

A26360 1 kit

QuantStudio™3D Digital PCR Starter Kit,

includes:

•QuantStudio™3D Digital PCR Reagent Kit

• QuantStudio™3D Digital PCR

20K Chip Kit v2 (12 chips)

A26361 1 kit

Required materials not supplied

This system uses common molecular biology equipment, supplies, and reagents.

Unless otherwise indicated, all materials are available through thermoﬁsher.com.

"MLS" indicates that the material is available from ﬁsherscientiﬁc.com or another

major laboratory supplier.

Item Source

Clean room-grade, low-lint polyester wipes Fisher Scientic

(Cat. No. NC0375992) or MLS

Gloves, powder-free, nitrile MLS

Microcentrifuge MLS

Pipette tips, 10 to 1000 μL MLS

Pipettes, P10 to P1000 MLS

Reaction tubes, DNase/RNase-free, non-stick or

low-binding: 0.5-mL or 1.5-mL

Fisher Scientic

(Cat. No. 13-864-253 or

13-864-254) or MLS

Vortexer MLS

Chapter 1 Product information

Required materials not supplied

10 QuantStudio™3D Digital PCR System User Guide

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