ion 520 & ion 530 ext kit −chef user guide (pub. no...

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

Ion 520™ & Ion 530™ ExT Kit – ChefUSER GUIDE

for use with:Ion Chef™ SystemIon S5™ SystemIon S5™ XL System

Catalog Number A30670Publication Number MAN0015805

Revision C.0

The information in this guide is subject to change without notice.DISCLAIMER: TO THE EXTENT ALLOWED BY LAW, LIFE TECHNOLOGIES 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 YOURUSE OF IT.

Revision history: Pub. No. MAN0015805

Revision Date DescriptionC.0 22 February 2017 Volume of Ion S5™ ExT Calibration Standard provided in ordered item (Cat. No. A30738)

changed from 24 µL to 16 µL.

B.0 26 January 2017 • Number of flows recommended for the instrument reset run (formerly "purge andclean") procedure changed from 500 to 200, and recommended time to perform theprocedure after sequencer initialization changed from 27 hours to 24 hours (see “Perform an instrument reset run with an initialized, unused Sequencing Reagentscartridge“ on page 64)

• Web links updated

A.0 13 October 2016 New user guide with detailed instructions for use of the Ion 520™ & Ion 530™ ExT Kit –Chef with the Ion Chef™ and the Ion S5™/Ion S5™ XL Systems.

Important Licensing Information: This product may be covered by one or more Limited Use Label Licenses. By use of this product, you accept theterms and conditions of all applicable Limited Use Label Licenses.Corporate entity: Life Technologies Corporation | Carlsbad, CA 92008 USA | Toll Free in USA 1 800 955 6288

Trademarks: All trademarks are the property of Thermo Fisher Scientific and its subsidiaries unless otherwise specified. Guava and easyCyte aretrademarks of Guava Technologies, Inc. Agilent and Bioanalyzer are trademarks of Agilent Technologies, Inc.

©2017 Thermo Fisher Scientific Inc. All rights reserved.

Contents

■ CHAPTER 1 Product information . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 7

Product description . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 7Library compatibility . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 7Software compatibility . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 7

Kit contents and storage . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 8Kit summary . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 8Kit components . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 8

Compatible Ion Chip kits . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 9

Ion S5™ ExT Calibration Standard . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 9

Ion S5™ Controls Kit Plus . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 9

E. coli DH10B Control 600 Library . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 10

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

About the Ion Chef™ System . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 11Ion Chef™ Instrument components . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 11Interior hardware and consumables . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 12About the Ion Chef™ System touchscreen interface . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 13

Precautions . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 14Avoid nucleic acid contamination . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 14Avoid chip damage . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 14Guidelines for using Ion Chef™ consumables and reagents . . . . . . . . . . . . . . . . . . . . . . 14

Workflow . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 15

■ CHAPTER 2 Before you begin . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 16

Create a Planned Run . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 16About Planned Runs . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 16Create a Planned Run . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 16Planned Run wizard key fields . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 20

Dilute the library . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 21Guidelines for library dilution . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 21Dilute the sample library . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 21Dilute the control library . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 21

Prepare the Ion Chef™ System for use . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 22

Ion 520™ & Ion 530™ ExT Kit − Chef User Guide 3

■ CHAPTER 3 Run the Ion Chef™ System .. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 23

Materials required . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 23

Prepare the library and consumables . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 23

Load the Ion Chef™ System . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 25Load the tip racks and PCR plate . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 26Load the Reagents and Solutions cartridges . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 28Load the Recovery Tubes and Enrichment Cartridge v2 . . . . . . . . . . . . . . . . . . . . . . . . . 29Load the Chip-loading centrifuge . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 31Confirm that consumables are correctly installed . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 33Ion Chef™ pre-run checklist . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 34

Start the Ion Chef™ run . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 35

Unload the chip for sequencing . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 40

■ CHAPTER 4 Initialize the sequencer . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 42

Ion S5™ System component positions . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 42

Before you begin . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 43

When a manual cleaning of the sequencer is required . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 43

Initialize the sequencer . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 44

Reagent consumables disposal . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 45CO2 scrubber removal and disposal . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 45Recycle Ion S5™ Wash and Cleaning solution bottles . . . . . . . . . . . . . . . . . . . . . . . . . . . . 46

■ CHAPTER 5 Start the sequencing run . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 47

Chip handling guidelines . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 47

Start the sequencing run . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 48

Maintain the sequencer . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 50Required materials . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 50Clean or decontaminate the sequencer . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 50

■ CHAPTER 6 Clean the Ion Chef™ System . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 51

About the cleaning protocol . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 51

Materials required . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 51

Clean the Ion Chef™ Instrument . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 52Remove and dispose of used consumables . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 53Inspect and clean the Recovery centrifuges and buckets . . . . . . . . . . . . . . . . . . . . . . . . 54Start the cleaning . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 55

Contents

4 Ion 520™ & Ion 530™ ExT Kit − Chef User Guide

■ APPENDIX A Troubleshooting . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 57

Ion Chef™ System setup and operation . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 57

Ion S5™ System setup and operation . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 60Ion S5™ Sequencer alarms and events . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 60Initialization—General errors . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 61

■ APPENDIX B Supplemental procedures . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 62

Quality control of Ion Sphere™ Particles . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 62Quality control using the Guava™ easyCyte™ 5 Flow Cytometer . . . . . . . . . . . . . . . . . . . 62Quality control using the Attune™ Acoustic Focusing Cytometer . . . . . . . . . . . . . . . . . . 62

Perform a manual cleaning of the sequencer . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 63

Perform an instrument reset run with an initialized, unused SequencingReagents cartridge . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 64

Maintain the Ion Chef™ System . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 65Install a firmware update . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 65Change the instrument name . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 66

■ APPENDIX C Ion Chef™ and Torrent Server network setup . . . . . . . . . 67

Networking overview . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 67About the Ion Chef™ instrument . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 68About the Torrent Server . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 68Networking guidelines and best practices . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 68Network setup workflow . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 68Network setup examples . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 68Collect the required network information . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 68Materials required . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 69General requirements . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 69

Ion Chef™ Networking Configuration Options . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 71Target the Torrent Server IP Address . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 71

Editing the Torrent Server firewall tables to allow the LAN port IP . . . . . . . . . . . . . . . . . . . 75Create an exception . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 75

Enable Planned Run sharing among Torrent Servers . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 77Set up Server Network (admin action) . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 78

Transfer a Planned Run . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 79

Undo a Planned Run transfer (administrator) . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 81

Undo a Planned Run transfer (user) . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 81

Contents


■ APPENDIX D Safety . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 82

Symbols on this instrument . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 82Conformity symbols . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 83

Location of safety labels on this instrument . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 84

Safety information for instruments not manufactured by Thermo Fisher Scientific . . . . . 84

Instrument safety . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 84General . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 84Physical injury . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 84Electrical . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 85Cleaning and decontamination . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 85Ultraviolet (UV) Safety . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 85

Safety and electromagnetic compatibility (EMC) standards . . . . . . . . . . . . . . . . . . . . . . . . . . 85Safety compliance . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 86EMC . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 86Environmental design . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 86

Instrument safety — Ion S5™/Ion S5™ XL System . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 86

Chemical safety . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 87

Biological hazard safety . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 88

■ Documentation and support . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 89

Customer and technical support . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 89

Limited product warranty . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 89

Contents


Product information

■ Product description . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 7

■ Kit contents and storage . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 8

■ Compatible Ion Chip kits . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 9

■ Ion S5™ ExT Calibration Standard . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 9

■ Ion S5™ Controls Kit Plus . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 9

■ E. coli DH10B Control 600 Library . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 10

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

■ About the Ion Chef™ System . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 11

■ Precautions . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 14

■ Workflow . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 15

IMPORTANT! Before using this product, read and understand the information in the“Safety” appendix in this document.

Product description

The Ion 520™ & Ion 530™ ExT Kit – Chef (Cat. No. A30670) includes reagents andmaterials for preparing and loading template-positive Ion Sphere™ Particles in Ion520™ or Ion 530™ Chips in 4 single-chip Ion Chef™ runs. The Ion Chef™ Instrumentuses automated isothermal amplification technology with the kit to prepare 400 to600-base-read templates with greatly reduced run times compared to emulsion PCR-based templating. The kit also includes reagents and materials to sequence the loadedchips on the Ion S5™ or Ion S5™ XL Sequencer.

The Ion 520™ & Ion 530™ ExT Kit – Chef can be used with 400 to 600-base-readlibraries of any type prepared using any available Ion library kit.

The Ion 520™ & Ion 530™ ExT Kit – Chef is compatible with Torrent Suite™ Softwarev5.2.1 and later. Be sure to update your Torrent Server to the latest available version ofTorrent Suite™ Software before using this kit.

1

Librarycompatibility

Softwarecompatibility


Kit contents and storage

IMPORTANT! Do not substitute components from any other Ion sequencing kits. Wehave verified this protocol using these specific materials. Substitution may adverselyaffect system performance.

IMPORTANT! Store all consumables and cartridges under the recommendedconditions and in an upright position. Do NOT store the Ion S5™ ExT SequencingReagents (Part No. A30522) on dry ice or in a closed environment where dry ice ispresent.

The Ion 520™ & Ion 530™ ExT Kit – Chef (Cat. No. A30670) supports 4 initializations,and contains all materials required for preparing and sequencing 4 Ion 520™ or Ion530™ Chips. Upon arrival, inspect all consumables and contact Technical Support ifany of the products have been damaged during shipping.

Component Part No. Quantity per kit

Ion S5™ ExT Chef Supplies A30673 4 boxes

Ion S5™ Chef Solutions A27754 1 box

Ion S5™ ExT Chef Reagents A30672 1 box

Ion S5™ ExT Sequencing Solutions A30521 1 box

Ion S5™ ExT Sequencing Reagents A30522 1 box

Contents Amount / box Storage

Ion S5™ ExT Chef Supplies (Part No. A30673)

Chip Adapter 1 15°C to 30°C

Enrichment Cartridge v2 1

Ion S5™ ExT Tip Cartridge 1

PCR Plate and Frame Seal v2 1 each

Recovery Station Disposable Lid v2 2

Recovery Tube 12

Ion S5™ Chef Solutions (Part No. A27754)

Ion S5™ Chef Solutions 4 cartridges 15°C to 30°C

Ion S5™ ExT Chef Reagents (Part No. A30672)

Ion S5™ ExT Chef Reagents 4 cartridges –30°C to –10°C

Kit summary

Kit components

Chapter 1 Product informationKit contents and storage1


Contents Amount / box Storage

Ion S5™ ExT Sequencing Solutions (Part No. A30521)

Ion S5™ ExT Wash Solution 4 × 1.5 L 15°C to 30°C

Ion S5™ Cleaning Solution 250 mL

Ion S5™ ExT Sequencing Reagents (Part No. A30522)

Ion S5™ ExT Sequencing Reagents 4 cartridges –30°C to –10°C

IMPORTANT! Do not store the Ion S5™ ExT Sequencing Reagents (Part No. A30522)on dry ice or in a closed environment containing dry ice.

Compatible Ion Chip kits

The following Ion Chip kits, ordered separately, are compatible with the Ion 520™ &Ion 530™ ExT Kit – Chef.

Description Cat. No. Quantity Storage

Ion 520™ Chip Kit (4-pack) A27761 4 chips15°C to 30°C

Ion 530™ Chip Kit (4-pack) A27763 4 chips

Ion S5™ ExT Calibration Standard

For de novo sequencing applications that do not include a reference BAM file, the IonS5™ ExT Calibration Standard can be added to libraries to increase base-callingaccuracy. The Ion S5™ ExT Calibration Standard (Cat. No. A30738) is orderedseparately.

Contents Amount Storage

Ion S5™ ExT Calibration Standard 16 µL –30°C to –10°C

Ion S5™ Controls Kit Plus

The E. coli DH10B Control 400 Library includes sufficient material for 3 templatingruns on the Ion Chef™ System. The Ion S5™ Controls Kit Plus (Cat. No. A30729) isordered separately.


Human CEPH Genomic DNA Control (red cap) 30 µL –30°C to –10°C

Human CEPH Control 200 Library (yellow cap) 12 µL

E. coli DH10B Control 400 Library (orange cap) 16 µL

Chapter 1 Product informationCompatible Ion Chip kits 1


E. coli DH10B Control 600 Library

The E. coli DH10B Control 600 Library includes sufficient material for 4 templatingruns on the Ion Chef™ System. The E. coli DH10B Control 600 Library (Cat. No.A32470) is ordered separately.


E. coli DH10B Control 600 Library (lilac cap) 20 µL –30°C to –10°C

Required materials not supplied

Unless otherwise indicated, all materials are available through thermofisher.com.MLS: Fisher Scientific (fisherscientific.com) or other major laboratory supplier.

Item Source

Non-interruptible Power Supply (UPS) [1] MLS

Thermal cycler with a heated lid MLS

Microcentrifuge[2] MLS

P2, P10, P20, P200, P1000 pipettor set and filtered tips MLS

0.2-mL PCR tubes, flat cap (Axygen™ MAXYMum Recovery™ PCRTube

Fisher Scientific14-222-283

or MLS

Microcentrifuge tubes, 1.5-mL or 1.7-mL MLS

Vortex mixer with a rubber platform MLS

Gloves, powder-free nitrile MLS

Ice buckets and ice —

Nuclease-free water, molecular biology grade MLS

Isopropyl alcohol, 70% solution MLS

Wipes, disposable lint-free MLS

[1] For laboratories that experience frequent power outages or line voltage fluctuations, we recommend that you use a non-interruptible power supply that is compatible with 2500 W output or higher.

[2] Must fit standard 0.2- and 1.5‑mL microcentrifuge tubes and generate 21,000 × g. To convert the RPMs of your centrifuge to RCF in units of gravity, see tools.thermofisher.com/content/sfs/brochures/TR0040-Centrifuge-speed.pdf.

Chapter 1 Product informationE. coli DH10B Control 600 Library1


http://www.thermofisher.com

http://fisherscientific.com

https://tools.thermofisher.com/content/sfs/brochures/TR0040-Centrifuge-speed.pdf

https://tools.thermofisher.com/content/sfs/brochures/TR0040-Centrifuge-speed.pdf

About the Ion Chef™ System

The Ion Chef™ System provides automated, high-throughput template preparationand chip loading for use with the Ion S5™ or Ion S5™ XL Sequencer. The system uses aset of cartridge-based consumables and reagents that enables a user to load an Ion520™ Chip, or an Ion 530™ Chip in about 7 hours with less than 15 minutes of hands-on time. The Ion Chef™ System features network integration with the Torrent Suite™

Software to enable sample and reagent traceability throughout the chip preparationworkflow.

The following figure illustrates the major external and internal features of theIon Chef™ Instrument.

4

1 2 3

5 6 7 8

1 Door – Provides access to the interior ofthe instrument. The door is locked in theclosed position during operation.

2 Micropipettor – A mechanical positive-displacement pipettor that performs allfluid transfers during sample and chippreparation.

3 Robotic arm – Performs fluid transferduring chip loading. The arm alsocontains an optical sensor that reads thebarcodes of instrument reagents andconsumables.

4 Touchscreen – Provides access to allinstrument functions for operation,maintenance, and troubleshooting.

5 Power button – Power switch for theIon Chef™ Instrument, where the statesare on (illuminated) and off.

6 Power port – A 100–240 VAC port thatprovides power to the Ion Chef™

Instrument.7 Ethernet port – An RJ45 port that provides

Ethernet (100 Mbit) communication withthe Ion Chef™ Instrument.

8 USB port – Provides USB communicationwith the Ion Chef™ Instrument. Used toupdate the instrument firmware and totransfer data during service ormaintenance.

Ion Chef™

Instrumentcomponents

Chapter 1 Product informationAbout the Ion Chef™ System 1


The following figure illustrates the interior of the Ion Chef™ Instrument and describesthe stations involved in the preparation of chips for sequencing on the Ion S5™ or IonS5™ XL System.

A

B

C

D

E

F

G

H

1 2 3 4 5 6 7 8 9 10 11 12

530

DABD02226

Ion Chef™S5 Series

Chip Balance

1

5

2 3

4

6

7

9 8

1 New pipette tips – The position of the rackcontaining unused pipette tips.

2 Automated heated cover – Transfers theplate cover to the PCR reaction platewithin the sample block. During thermalcycling, the heated cover appliescompression to seal the reaction plateand heats the cover to preventcondensation.

3 Waste pipette tip rack – The position ofthe rack containing waste (used) pipettetips.

4 Thermal cycler sample block – Performsthermal cycling of the sequencingreactions on a 96-well PCR reaction plate.

5 Reagents station – The position on theinstrument deck of the diluted libraries,NaOH, and the Ion S5™ ExT Chef Reagentscartridge.

6 Solutions station – The position on theinstrument deck of the Ion S5™ ChefSolutions cartridge, which is maintainedat room temperature.

7 Recovery centrifuges – Twin stations thatperform centrifugation of the ISPs duringthe recovery phase of templatepreparation.

8 Enrichment station – The position of therack containing consumables for theenrichment of the template-positive ISPs.

9 Chip-loading centrifuge – Performscentrifugation of the sequencing chip thathas been mounted to a chip-loadingadapter and loaded with template-positiveISPs, and a chip balance loaded oppositeto the sequencing chip.

Interior hardwareand consumables

Chapter 1 Product informationAbout the Ion Chef™ System1


The Ion Chef™ System features a simple interface for loading chips, cleaning theinstrument, and performing system maintenance and configuration tasks.

2

1

3 4 5

1 Set up Run button – Set up the Ion Chef™

template preparation and chip-loadingroutine. Choose Step by Step to have theinstrument lead you stepwise throughinstallation of reagents and consumables,or choose Quick Start to proceed if youhave already installed the consumables.

2 Open Door button.3 Notifications button – View notifications

about instrument status during andbetween runs.

4 Quick Start button – Proceed directly tothe Quick Start instrument setup mode.User verifies the loading of a new pipettetip cartridge and an empty pipette tip rackto hold waste tips generated during therun, before proceeding to Deck Scan.

5 Settings button – Advance to a screenproviding the following options:

• Notifications: view notifications aboutinstrument status during and betweenruns

• Instrument Settings: view currentsettings and network configuration,set instrument name, adjust time zone

• Service tools: access screens forservice-related maintenance andinstrument diagnostics

• Torrent Server: add and manageTorrent Server connections

• Clean Ion Chef: proceed directly to theinstrument cleaning routine

• Check for updates: check availabilityof system software updates

About the IonChef™ Systemtouchscreeninterface

Chapter 1 Product informationAbout the Ion Chef™ System 1


Precautions

WARNING! The protection that is provided by the equipment can be impairedif the instrument is operated outside the environment and use specifications,the user provides inadequate maintenance, or the equipment is used in amanner that is not specified by the manufacturer.

IMPORTANT! A primary source of contamination is spurious DNA fragments fromprevious sample processing steps. Do not introduce amplified DNA into the librarypreparation laboratory or work area.

IMPORTANT! To avoid possible damage to the chip due to electrostatic discharge,ground yourself before picking up a chip or placing a chip on a surface such as a labbench. For example, touch the metal trim on the chip compartment before inserting orremoving a chip from the chip clamp.

To ensure the proper function of the Ion Chef™ System, handle the associatedconsumables and reagents according to the following guidelines:

• Store all Ion Chef™ System consumables and cartridges under the recommendedconditions and in an upright position.

• Inspect all Ion Chef™ System consumables and cartridges for damage uponarrival and again before use.

• Hold sequencing chips by gently gripping them by their edges. Do not weargloves when handling chips.

• When the Ion Chef™ System is not in use, remove all consumables and reagentsfrom the deck and close the instrument door.

• Except for the pipette tip cartridge, do not reuse any of the Ion Chef™ Systemconsumables or reagents. After each run, transfer the empty Ion S5™ ExT TipCartridge to the waste tip station for use as the waste container for the next run.

IMPORTANT! All Ion Chef™ Instrument components are single-use only.

• Use only Ion Torrent™ kits and supplies with the Ion Chef™ Instrument. The useof third-party reagents and supplies can adversely affect the performance of theIon Chef™ Instrument and chips prepared.

• Remove and sequence chips within 1 hour after the Ion Chef™ System finishesloading them. If you cannot sequence a loaded chip immediately, store it within achip storage container at 4°C until you are ready to run it (up to 6–8 hoursmaximum).

Note: If you choose to store a loaded chip, remove the chip from 4°C storage (butkeep it in the chip storage container) at least 20 minutes prior to running it,allowing the chip to warm to room temperature.

Avoid nucleic acidcontamination

Avoid chip damage

Guidelines forusing Ion Chef™

consumables andreagents

Chapter 1 Product informationPrecautions1


Workflow

The following workflow illustrates how to prepare and a load sample using the Ion520™ & Ion 530™ ExT Kit – Chef, and Ion Chef™ System for sequencing on the Ion S5™

or Ion S5™ XL Sequencer.

Create a Planned Run

q

Dilute the sample library

q

Prepare the library and consumables

q

Load the Ion Chef™ System

q

Start the Ion Chef™ run

q

Unload the chip for sequencing

q

Initialize the sequencer

q

Start the sequencing run

Chapter 1 Product informationWorkflow 1


Before you begin

■ Create a Planned Run . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 16

■ Dilute the library . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 21

■ Prepare the Ion Chef™ System for use . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 22

Create a Planned Run

Planned Runs contain all the settings that are used in a sequencing run, includingnumber of flows, kit types, barcodes, sample information, and reference files (if any).They are used to track samples, chips, and reagents throughout the sequencingworkflow, from template preparation on the Ion Chef™ Instrument throughsequencing on the Ion S5™ or Ion S5™ XL Sequencer and subsequent data analysis.

1. Log in to the Torrent Server via the Torrent Browser.

2. Select the Plan tab, click Templates, select the application that you want to run(such as Whole Genome), then click either:

• Plan New Run to plan a new run using the generic template for the selectedapplication.

2

About PlannedRuns

Create a PlannedRun


• Plan Run in the dropdown menu under the Settings tab to the right ofthe existing template you select from the template list.

3. In the Planned Run wizard, review the IonReporter and Application tabs, thenmake selections appropriate to your run. On the Kits screen, make the followingselections:

a. Select Ion S5™ System from the Instrument dropdown list.

b. Select the appropriate chip type in the Chip Type dropdown list.

c. Select Ion Chef to the right of Template Kit, then select Ion 520 & Ion 530ExT Kit-Chef from the Template Kit dropdown list.

d. Select Ion S5 ExT Sequencing Kit from the Sequencing Kit dropdown list.

e. Select the appropriate barcode set, if used, from the Barcode Set dropdownlist.

Chapter 2 Before you beginCreate a Planned Run 2


f. Ensure that the appropriate number of flows is entered after selecting thechip: 1350 flows are sufficient for 600-base-read sequencing on either an Ion520™ Chip, or an Ion 530™ Chip.

IMPORTANT! Do not exceed 1400 total flows for one Ion S5™ SequencingReagents Cartridge.

g. Select or edit the optional information fields appropriately for your run.

IMPORTANT! For de novo sequencing applications that do not include areference BAM file, select Enable Calibration Standard from the BaseCalibration Mode dropdown list. Select this option only if you add Ion S5™

ExT Calibration Standard to the Ion Chef™ Library Sample Tube. Selectingthis option and adding the Ion S5™ ExT Calibration Standard to your libraryallows greater accuracy of base-calling in libraries for which a referenceBAM file does not exist.

h. Click Next.

4. Review the Plugins and Projects screens, then make selections appropriate toyour run.

Chapter 2 Before you beginCreate a Planned Run2


5. On the Plan screen, enter or scan the barcode of the Ion Chef™ Library SampleTube into the Sample Tube Label field. Enter "1" in the Number of chips field,then click the check mark button to the right of this field. The Chip ID field willpopulate after deck scanning.

Note: If you selected a Barcode Set in the Kits tab, fields appear in the Plan tabwhere you enter the number of barcodes used, and the corresponding SampleTube Label. Then enter a sample name for each barcode used in the Samplestable.

Note: For a complete description of the fields of the Torrent Browser application,see the Torrent Suite™ Software Help, available at thermofisher.com/ion-s5-docs.

6. When you have completed your selections, click Plan Run at the bottom right ofthe Plan screen to save the run.

The run is listed on the Planned Runs page under the name that you specified, and isautomatically used by the Ion Chef™ System when the associated sample is loaded.

Chapter 2 Before you beginCreate a Planned Run 2


http://www.thermofisher.com/ion-s5-docs.html

Field name Description

Application Select the sequencing application you are performing.

Template Kit Select Ion 520 & Ion 530 ExT Kit-Chef

Sequencing Kit Select Ion S5 ExT Sequencing Kit

Chip Type Select Ion 520™ Chip, or Ion 530™ Chip

Barcode Set(optional)

If you are using barcodes with:

• DNA libraries: Select the appropriate barcode set used.

• RNA libraries prepared using the Ion Total RNA-Seq Kit v2:Select the IonXpressRNA barcode set, which contains all16 barcodes in the Ion Xpress™ RNA BC01-16 Kit(Cat. No. 4475485).

If you are not using barcodes with:

• DNA libraries: Leave the Barcode field blank.

• RNA libraries prepared using the Ion Total RNA-Seq Kit v2:Select RNA_Barcode_None from the dropdown list. This willensure that the proper trimming is performed on theresulting sequence when the RNA library does not have abarcode.

Flows Enter the appropriate number of flows for the read length. Forexample, enter

• 850 flows for 400‑base read sequencing

• 1350 flows for 600‑base read sequencing.

Plugins Select the appropriate plugins for your application.

Projects Select or add a project within which to group your run data. Youcan include runs in multiple projects, and remove runs from aproject at any time.

Run Plan Name(required)

Enter a name for the run.

Reference Library Select a reference library uploaded to the Torrent Server, if any.

BED files Select the Target Regions or HotSpot Regions BED file on the IonTorrent Server, if any.

Sample Name(Required)

Is Ion Reporter™ Uploader enabled?

• Yes – Enter each sample name and select the appropriatevalues for workflow, relation, relation role, and set ID.

• No – Scan the barcode of the library tube into the "SampleName" and "Sample Tube Label" fields for the specificsample.

Planned Runwizard key fields

Chapter 2 Before you beginCreate a Planned Run2


Field name Description

Sample Tube Label Scan or enter the barcode located on the Ion Chef™ Library SampleTube that you intend to use to load the sample into the Ion Chef™

Instrument.

MonitoringThresholds

Set thresholds for Bead Loading, Usable Sequence, and KeySignal. In the Torrent Browser Monitor4Runs in Progress tab, analert is displayed if the values for a run fall below the selectedthresholds.

Dilute the library

• The sample library dilution recommendations below represent optimal inputconcentrations for control libraries. Users who have already optimized a libraryconcentration for other Ion Chef™ reagent kits or the manual Ion PGM™ TemplateIA 500 Kit may need to re-optimize library concentration for the Ion 520™ & Ion530™ ExT Kit – Chef.

• Recommendations are based on qPCR quantification. If libraries are quantifiedwith an Agilent™ 2100 Bioanalyzer™ instrument, a higher calculatedconcentration may need to be used for equivalent input.

• Prepare a fresh dilution of each library before use with the Ion Chef™ System, anduse the library dilutions within 48 hours.

IMPORTANT! Before proceeding, dilute the Ion library to the optimal inputconcentration. The quality of your sequencing data relies on achieving the correctconcentration of starting library.

Dilute stock libraries according to the following table. Use polyclonality and low-quality filter results from a sequencing run performed with ISPs templated at thestarting concentration, then titrate up or down to achieve optimal concentrations, ifneeded.

Library read length Recommendedconcentration Molecules per input volume

400−600 bp 60–100 pM 1.8−3.0 × 109

If you are running a control library for troubleshooting purposes, dilute theappropriate library obtained from one of the controls kits, according to the followingtable:

Control library Volume of library Volume of Nuclease-freeWater

E. coli DH10B Control 400Library

5 µL 45 µL

E. coli DH10B Control 600Library

5 µL 45 µL

Guidelines forlibrary dilution

Dilute the samplelibrary

Dilute the controllibrary

Chapter 2 Before you beginDilute the library 2


Prepare the Ion Chef™ System for use

Before you use the Ion Chef™ Instrument:

• Ensure that the Ion Chef™ Instrument has been cleaned following the previousrun. If not, clean the instrument before loading it with consumables.

Note: For more information on the cleaning procedure, see Chapter 6, “Clean theIon Chef™ System“.

• Inspect the empty compartments of the Reagents and Solutions stations forcondensation. Condensate can collect in these compartments, depending ontemperature and humidity conditions. Before loading consumables into theinstrument, wipe the compartments dry with a laboratory wipe or absorbentcloth, if needed.

• Thaw the Ion S5™ ExT Chef Reagents cartridge at room temperature for45 minutes before use.

• Ensure that the Ion Chef™ Instrument has a connection to your Torrent Server. Onthe Ion Chef™ home touchscreen, touch Settings, then Torrent Server to view theconnection status of your instrument.

Note: If the instrument is not connected, see Appendix C, “Ion Chef™ andTorrent Server network setup“ for instructions on how to configure a direct orindirect network connection of the Ion Chef™ Instrument to a Torrent Server.

Chapter 2 Before you beginPrepare the Ion Chef™ System for use2


Run the Ion Chef™ System

■ Materials required . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 23

■ Prepare the library and consumables . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 23

■ Load the Ion Chef™ System . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 25

■ Start the Ion Chef™ run . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 35

■ Unload the chip for sequencing . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 40

This chapter describes:• Setting up the Ion Chef™ Instrument for use by diluting the two Ion libraries, and

loading the instrument with all of the required reagents and consumables• Starting an Ion Chef™ run• Unloading the Ion Chef™ Instrument

Materials required

• Ion 520™ & Ion 530™ ExT Kit – Chef (Cat. No. A30670)• Ion 520™ Chip or Ion 530™ Chip• Molecular-biology grade Nuclease-free Water• P200 pipettor and filtered tips• Waste container

Prepare the library and consumables

1. Unbox the Ion S5™ ExT Chef Reagents cartridge 45 minutes before use, thenallow it to warm to room temperature.

IMPORTANT! The Reagents cartridge must sit at room temperature for45 minutes before use.

2. Pipet 50 µL of the diluted library (see “Dilute the sample library“ on page 21)into the Ion Chef™ Library Sample Tube (barcoded tube) from Position A of theReagents cartridge.

Note: If you are performing a de novo sequencing experiment that does notinclude a reference BAM file, add 4 µL of the Ion S5™ ExT Calibration Standardto the diluted library in the Ion Chef™ Library Sample Tube. Cap the tube, vortex

3


to mix, then centrifuge for 1–2 seconds to collect contents at the bottom of thetube.

Note: If running the E. coli DH10B Control 400 Library, or the E. coli DH10BControl 600 Library, dilute the stock library with Nuclease-free Water followingthe recommendations in “Dilute the control library“ on page 21, then pipet 50 µLof the diluted library into the Ion Chef™ Library Sample Tube.

3. Cap and store the diluted DNA library on ice until you are ready to load the tubeinto the Reagents cartridge and the Ion Chef™ Instrument.

4. Remove the other cartridges and consumables from their packaging, then placethem on the bench next to the Ion Chef™ Instrument.Prepare the following:

• Chip Adapter and Chip Balance• Enrichment Cartridge v2• Ion S5™ ExT Tip Cartridge• PCR Plate and Frame Seal v2• Recovery Station Disposable Lid v2 (2)• Recovery Tube (12)• Ion S5™ Chef Solutions cartridge• Ion S5™ ExT Chef Reagents cartridge (from step 1)

IMPORTANT! Before use, gently tap the Reagents cartridge and Solutionscartridge on the bench to force the reagents to the bottoms of the tubes.

Note: When stored under normal conditions, a precipitate can form in sometubes of the Ion S5™ ExT Chef Reagents cartridge. If present, load the cartridge asdirected. The precipitate dissolves when the reagents are mixed duringinstrument operation.

Chapter 3 Run the Ion Chef™ SystemPrepare the library and consumables3


Load the Ion Chef™ System

IMPORTANT!· Rated centrifuge speeds are intended for operation only with the provided buckets

and approved consumable chips, tubes, and sample preparation reagents.· The Chip-loading centrifuge is rated to operate at the listed rotational frequencies

with the chip buckets, chip, adapter, and chip balance. The centrifuge must be load-balanced. Proper care must be taken to load the bucket properly. If excessivevibrations arise, check to ensure that items are installed properly and rotors areload-balanced.

· Use only the materials supplied in the Ion 520™ & Ion 530™ ExT Kit – Chef to runthe centrifuges at the rated speeds. Do not remove or change the rotors. Inspect thebuckets before each use to assure normal operation.

· Ensure that the instrument is powered on and has been cleaned following the lastuse.

· Ensure that all components are clean and dry before loading them onto theIon Chef™ Instrument.

· Ensure that the Reagents and Solutions station compartments are dry and free ofcondensate before loading components.

Follow the procedure below to load the Ion Chef™ Instrument. A completely loadedinstrument is shown in the following figure:

A

B

C

D

E

F

G

H

1 2 3 4 5 6 7 8 9 10 11 12

530

DABD02226


Chip Balance

5

9

31 2

4

6

7

8

1 New Ion S5™ ExT Tip Cartridge2 Frame Seal v2 (inserted)3 Empty tip rack (move from new Tip

Cartridge position)4 PCR plate5 Ion S5™ ExT Chef Reagents cartridge

6 Ion S5™ Chef Solutions cartridge7 Recovery centrifuges: Recovery Tubes and

Recovery Station Disposable Lid v28 Enrichment Cartridge v29 Chip-loading centrifuge: Chip

Adapter/Chip assembly and Chip Balance

Chapter 3 Run the Ion Chef™ SystemLoad the Ion Chef™ System 3


1. Touch (Open Door) in the instrument touchscreen to open the instrumentdoor, then wait for the latch to open.

2. Lift the instrument door to the top of the travel until the latch mechanismengages.

1

1 Hold here, then lift

3. Load an empty pipette tip rack to the Used (Waste) Pipette Tip Position, thenchange gloves.

11

1 Used Pipette Tip Position

IMPORTANT!· Confirm that the pipette tip rack in the Used (Waste) Pipette Tip Position does

not contain any tips. The instrument will abort the run if tips are present in theused position.

· To prevent contamination, change gloves immediately after moving the emptypipette tip rack to the Used (Waste) Pipette Tip Position.

Note: A small amount of dried residue may be present in the tub of the emptypipette tip rack after a run. This will not affect the next run in the Used PipetteTip Position.

4. Unwrap a new Ion S5™ ExT Tip Cartridge and remove the cover to expose thepipette tips, then load it in the New Pipette Tip Position.

Note: Two Ion Chef™ Piercing Tips are pre-loaded into tip positions G7 and H7on the Ion S5™ ExT Tip Cartridge.

Note: Keep the cover for later use.

Load the tip racksand PCR plate

Chapter 3 Run the Ion Chef™ SystemLoad the Ion Chef™ System3


5. Slide the catch forward to allow the locking bracket to pivot upward. Load theIon S5™ ExT Tip Cartridge into the New Pipette Tip Position, pull the bracketdownward, then push the catch backward to lock the cartridge in place.

2

3

1

4

1 New Pipette Tip Position2 Bracket3 Catch4 New Ion S5™ ExT Tip Cartridge

6. Load a new PCR plate into the thermal cycler sample block, then insert a newFrame Seal v2 underneath the automated heated cover.

IMPORTANT! When the Frame Seal v2 is positioned correctly, its tabs projectupward and contact the heated cover.

A

B

C

D

E

F

G

H

1 2 3 4 5 6 7 8 9 10 11 12

A

B

C

D

E

F

G

H

1 2 3 4 5 6 7 8 9 10 11 12 41

3

2

1 Thermal cycler sample block2 Well A13 Cover4 Keyed corner



IMPORTANT! Thaw the Reagents cartridge at room temperature for 45 minutesbefore use.

1. Gently tap the Ion S5™ ExT Chef Reagents cartridge on the bench to force thereagents to the bottoms of the tubes.

2. Load the cartridge into the Reagents station so that it snaps into place and is levelon the deck.

IMPORTANT! Do not force the Ion Chef™ cartridges into place. Each cartridgefits only one location on the deck and in one orientation. If a cartridge does notfit, confirm that you are loading the correct cartridge in the correct orientation.

A

B

C

D

E

F

G

H

1 2 3 4 5 6 7 8 9 10 11 12

12

1 Reagents station (4°C)2 Ion S5™ ExT Chef Reagents cartridge

3. Uncap, then load the Library Sample Tubes containing 50 µL of diluted libraryinto Position A on the Reagents cartridge.

A00012216

1

D 4

C 3

B 2

1 Position A (Library Sample Tube)2 Position B (2M NaOH)3 Position C (Ion S5™ ExT Pellet)4 Position D (Ion S5™ ExT Pellet)

IMPORTANT! Orient the Library Sample Tubes so that the barcode is visible andoriented to the right.

Load the Reagentsand Solutionscartridges



4. Uncap the tube of 2M NaOH in Position B and the tubes in Positions C and D,each containing an Ion S5™ ExT Pellet.

IMPORTANT! When the Reagents cartridge is loaded:

· Press down on the Library Sample Tubes to ensure that it is firmly seated inthe cartridge.

· Ensure that all four tubes are uncapped.

5. Gently tap the Ion S5™ Chef Solutions cartridge on the bench to force the reagentsto the bottoms of the tubes.

6. Load the Solutions cartridge into the Solutions station until it snaps into placeand is level on the deck.

A

B

C

D

E

F

G

H

1 2 3 4 5 6 7 8 9 10 11 12

2

1

1 Solutions station (room temperature)2 Ion S5™ Chef Solutions cartridge

1. Load six Recovery Tubes (v2) into each Recovery centrifuge.

A

B

C

D

E

F

G

H

1 2 3 4 5 6 7 8 9 10 11 12

1 2

1 Recovery centrifuges2 Recovery Tube

Before sealing each centrifuge, confirm that:• The centrifuge is load-balanced with all required consumables.

IMPORTANT! The centrifuge must be load-balanced.

• The buckets are securely seated in the centrifuge rotors.• The buckets are oriented correctly in the centrifuge so that they pivot

outwards.

Load the RecoveryTubes andEnrichmentCartridge v2



2. Place a Recovery Station Disposable Lid v2 over each centrifuge by lining up thetab with the depression on the deck, then snap into place. Ensure that the lidssnap completely into place by applying firm downward pressure along the lidperimeter.

3. Close the hinged cover of the Recovery centrifuges. Confirm that the port of eachdisposable lid is positioned toward the rear of the instrument.

1 32

4

4

1 Recovery Tubes installed2 Recovery Station Disposable Lids installed3 Recovery centrifuge cover closed4 Port

IMPORTANT! Do not obstruct or place any object on top of the Recoverycentrifuge cover.

IMPORTANT! Use only the supplied materials, including buckets anddisposables, to run the centrifuges at the rated speeds. Do not remove or changethe rotors. To ensure normal operation, inspect the buckets before each use.

4. Load the Enrichment Cartridge v2, then press down on the cartridge to ensurethat it is level with the instrument deck.

IMPORTANT! Ensure that the Enrichment Cartridge v2 is loaded so that thelettering is positioned on the right side of the cartridge.

A

B

C

D

E

F

G

H

1 2 3 4 5 6 7 8 9 10 11 12

A

B

C

D

E

F

G

H

1

2

3

1 Enrichment station2 Enrichment Cartridge v23 Lettering



1. Load the sequencing chip into acentrifuge bucket, then attach a ChipAdapter to the assembly.

Note: (Optional) Label the tops ofchips to distinguish them. Do notobstruct or overwrite the chipbarcode with the label.

a. Place the chip in the chip-loading bucket, then align thewells of the Chip Adapter to thewells of the chip, orienting theadapter onto the chip so that thechip barcode is visible.

b. Place the adapter onto the chip,then insert the stationary tabs atthe reservoir end of the adapter into the slots of the bucket.

c. Gently squeeze the flexible tabs at the other end of the adapter into thebucket slots until the adapter locks into place.

d. Confirm that the tabs at all four corners of the adapter are fitted into theslots in the centrifuge bucket. Loading can fail if the adapter is not attachedsecurely.

2. Load the adapter/chip/bucket assembly into Position 1 of the Chip-loadingcentrifuge.

A

B

C

D

E

F

G

H

1 2 3 4 5 6 7 8 9 10 11 12

14

2

5

3

1 Chip-loading centrifuge2 Position 13 Position 2 marker holes4 Position 25 Position 1 marker hole

Note: Position 1 of the Chip-loading centrifuge is the position 90° clockwisefrom the single hole in the rotor bucket cover at rest. The chip loaded in Position1 is loaded with ISPs prepared from the DNA library in the Library Sample Tubeloaded in Position A of the Reagents cartridge.

Load the Chip-loading centrifuge

530

CBDD02226

1

2

3

6

5

7

4

1 Chip Adaptor2 Ion chip3 Bucket4 Ports (align with chip)5 Tabs6 Keyed corner (align with bucket)7 Slots



3. Load an Ion Chef™ S5 Series Chip Balance into Position 2 of the Chip-loadingcentrifuge.

530DABD02226


Chip Balance 1 3

4

2


Chip Balance

1 Position 1 (Chip)2 Position 2 marker holes3 Position 2 (Chip Balance)4 Position 1 marker hole

IMPORTANT! When the Chip-loading centrifuge is loaded, ensure that the ChipAdapter is firmly attached to its bucket, and that each bucket is securely seated inthe centrifuge rotors.

4. Ensure the centrifuge is load-balanced, and the chip buckets are securely seatedand oriented correctly in the centrifuge so that they pivot 90° outwards whentouched. Then close the lid of the Chip-loading centrifuge.

IMPORTANT! Do not obstruct or place any object on top of the lid.



Before continuing:• Confirm that each cartridge is at the correct location and in the correct

orientation.• Press down on all cartridges to confirm that they are firmly pressed into place.• Confirm that all tubes in the Ion S5™ ExT Chef Reagents cartridge, including the

tube of NaOH in Position C, are uncapped and firmly pressed into place.• Confirm that the centrifuge lids are installed correctly so that the port is oriented

toward the rear of the instrument.• Confirm that the tube and chip buckets are seated securely in the rotor arms of

the Chip-loading and Recovery centrifuges, and that the consumables theycontain are correctly installed.

CAUTION! To ensure correct and safe instrument operation, you must confirmthat all consumables are installed correctly to the deck before you start a run.The Ion Chef™ Instrument does not verify all aspects of the consumable setupprior to beginning each run.

Confirm thatconsumables arecorrectly installed



A

B

C

D

E

F

G

H

1 2 3 4 5 6 7 8 9 10 11 12

530

DABD02226

Ion Che f ™ S5 Series Chip Balance

1) A new Ion S5™ ExT Tip Cartridge is loaded in the New Pipette Tip position.2) Bracket is pulled downward to lock the tip cartridge in place.

1) A new PCR Plate is loaded in the thermal cycler sample block.2) A Frame Seal v2 is in place and oriented under the heated cover. 1) The tip rack from previous run is transferred

from the New Pipette Tip position to the Used Pipette Tip position.

1) Recovery Tubes (v2) are securely seated in the Recovery centrifuge buckets.2) Recovery centrifuges are load-balanced.3) Recovery Station Disposable Lids (v2) are positioned so that the port is correctly oriented: port should not be located in the position facing the front of the instrument.

1) Enrichment Cartridge v2 is pressed into place at the Enrichment station so that the cartridge is firmly seated and level with the deck.2) Lettering on the cartridge is right-side up and positioned to the right of the enrichment tubes.1) Adapter/chip assembly is correctly

seated in the Chip-loading centrifuge bucket loaded in Position 1, with the adaptor clips inserted into the bucket slots.2) Centrifuge buckets containing the adapter/chip assembly and Chip Balance, loaded in Position 2, are securely seated in the centrifuge rotor, and freely pivot 90º outwards.

1) Solutions cartridge is pressed into place at the Solutions station so that the cartridge is firmly seated and level with the deck.

1) Reagents cartridge contents are thawed (leave at room temperature for 45 minutes).2) Reagents cartridge tubes are uncapped and pressed into place at the Reagents station so that the tubes are firmly seated and level with the deck.3) Library Sample tube is firmly seated and in the correct orientation with barcode facing the PCR Plate and cap removed.

Ion Chef™ pre-run checklist



Start the Ion Chef™ run

1. Ensure that you have loaded the instrument with all kits and consumables.

2. On the Ion Chef™ Instrument home touchscreen, touch Set up run.

3. Touch Step by step to have the instrument lead you through the instrumentsetup, or touch Quick Start to skip the instrument setup screens.

Note: If you selected Step by step, touch Template on the next screen to start anIon Chef™ templating workflow.

Chapter 3 Run the Ion Chef™ SystemStart the Ion Chef™ run 3


4. Follow the on-screen instructions. When prompted, close the instrument door byfirst lifting it slightly to disengage the locking mechanism, then push down onthe door until the locks engage. After the door closes, the instrument visionsystem activates.

IMPORTANT! Do not close the door by pulling it straight down from the openposition. You must lift the door slightly before you can close it. Ensure that bothsides of the door are locked after closing it.

12

3

1 Lift door first2 Lower3 Press down to lock

5. When prompted, touch Start check to begin Deck Scan. The instrument scans thebarcodes of all consumables and reagents to verify their presence andcompatibility.

Note: During Deck Scan, the touchscreen can display warnings if the Ion Chef™

Instrument detects missing or incompatible consumables. You must address allwarnings before the run can start. After you address each condition, touch Yes tocontinue.

IMPORTANT! The Deck Scan function is not a substitute for manual inspectionof the reagents and consumables on the Ion Chef™ Instrument before starting arun. To ensure proper and safe instrument operation, ensure that all consumablesare installed correctly before you continue.

6. When the Deck Scan is complete, touch Next to display the Data Destinationscreen.

Chapter 3 Run the Ion Chef™ SystemStart the Ion Chef™ run3


7. Ensure that the instrument displays the correct kit type, chip type, chip barcode,and Planned Run. If the correct Planned Run does not display, touch thedropdown menu to select the appropriate Planned Run, then touch Next.

IMPORTANT! If the kit and chip type are not correct, confirm that you are usingthe correct kit and chip. If you are using the correct kit and chip, and an incorrectkit or chip type appears on the screen, contact Technical Support.

8. On the Run Options screen, touch the appropriate option to complete the run,then, if needed, enter the time that you want the run to end after touching thebox on the right where the time at run completion displays. A dialog opens forentering a run completion time.

The Ion Chef™ Instrument provides two options for obtaining a quality control(QC) sample that can be used to evaluate templating efficiency. Depending onyour selection, the QC sample will be made available either during or after therun. In either case, you can obtain unenriched sample from the Library SampleTubes at Position A on the Reagents cartridge, or enriched sample from PositionE on the Enrichment Cartridge v2.

By selecting You can obtain the QC samples Approximate time afterrun start

Time immediately after the run ends, at thetime you specify:

6.75 hours

Pause when the instrument pauses operationbefore the chip loading step:

5.75 hours



Note: Select Pause if you are uncertain of library quality and want to evaluatetemplating efficiency before the chip is loaded. If you do not pause the run, youcan collect QC samples after the run. Save the samples until sequence analysis iscomplete to have them available for troubleshooting.

9. On the Run Options screen, touch Start run to begin the run.

Note: If you must stop the run for any reason, touch Cancel, then touch Yes toconfirm the cancellation.If the Ion Chef™ Instrument encounters a problem during the run, it aborts therun, then displays the error on the instrument touchscreen. If a run fails:

1. Remove the consumables from the deck, then clean the instrument. Ifpossible, retain the consumables for troubleshooting.

2. Reset, then re-attempt the run. If the run fails again, contact TechnicalSupport to troubleshoot the problem. To help troubleshooting, record theerror message by taking a photo of the touchscreen.

10. Initialize the Ion S5™ or Ion S5™ XL Sequencer at least 40 minutes before theIon Chef™ System finishes chip loading.

11. If you chose to pause the run to analyze the templating efficiency, remove thesample for testing when prompted to do so by the Ion Chef™ Instrument(approximately 5.75 hours into the run).

a. When prompted to remove the QC sample, open the instrument door.

Chapter 3 Run the Ion Chef™ SystemStart the Ion Chef™ run3


b. Transfer the QC sample (entire volume) from Position A of the Ion S5™ ExTChef Reagents cartridge on the instrument deck to a new labeledmicrocentrifuge tube.

IMPORTANT! Do not remove the Library Sample Tubes from the Ion S5™

ExT Chef Reagents cartridge.

IMPORTANT! If you unintentionally close the instrument door before youobtain the QC sample, you must wait until the end of the run before you cancollect it. You cannot pause the run or open the door after it has been closed.

D

A

B

C

000122161

1 Position A (QC sample)

c. If you are performing quality assessment of an enriched sample, transfer theQC sample from Position E of the Enrichment Cartridge v2 to a new labeledmicrocentrifuge tube.

d. Analyze the QC sample. See “Quality control using the Guava™ easyCyte™ 5Flow Cytometer“ or “Quality control using the Attune™ Acoustic FocusingCytometer“ on page 62.

e. Close the instrument door, then touch Continue to complete the run.

12. When the run is complete, unload the Ion Chef™ Instrument and sequence thechip immediately. You can collect QC samples from the Reagents and/orEnrichment cartridges if you have not done so already.

Note: If you cannot sequence a loaded chip immediately, place the chip into aseparate chip storage container, then store at 4°C until you are ready to sequenceit (up to 6–8 hours maximum).



Unload the chip for sequencing

1. Open the instrument door:a. In the instrument touchscreen, touch (Open Door), then wait for the latch

to open.

b. Lift the instrument door to the top of the travel until the latch mechanismengages.

1


2. Open the lid of the Chip-loading centrifuge, then unload the adapter/chip/bucketassembly from the Ion Chef™ Instrument.

A

B

C

D

E

F

G

H

1 2 3 4 5 6 7 8 9 10 11 12

530

DABD02226


Chip Balance

530DABD02226


Chip Balance

1

1 Remove the adaptor/chip/bucket assemblyTo unload the chip:

a. Remove the adapter/chip/bucket assembly from the Chip-loadingcentrifuge.

b. Apply pressure to both ends of the Chip Adapter, then remove and discardit.

Chapter 3 Run the Ion Chef™ SystemUnload the chip for sequencing3


c. Grasp the chip by its edges, carefully lift the chip out of the bucket and set itaside on a clean, static-free surface, then return the bucket to the Chip-loading centrifuge.

530

CBDD02226

1

2

3

5

4

1 Chip Adapter2 Ion chip3 Bucket4 Press and remove adapter5 Remove carefully

3. Close the instrument door by first lifting it slightly to disengage the lockingmechanism, then push down on the door until the locks engage.

IMPORTANT! Do not close the door by pulling it straight down from the openposition. You must lift the door slightly before you can close it. Confirm that bothsides of the door are locked after closing it.

12

3

1 Lift door first2 Lower3 Press down to lock

4. Load the chip into a Ion S5™ Sequencer and promptly start the sequencing run.If you cannot sequence a loaded chip immediately or plan to sequence two chipsper initialization, place the chip into a separate chip storage container and storeat 4°C until you are ready to sequence it (up to 6–8 hours maximum).

IMPORTANT! If you chose to store a loaded chip, remove the chip from thecontainer and place it on a clean surface in the dark to warm to roomtemperature at least 20 minutes before you intend to run it.

Chapter 3 Run the Ion Chef™ SystemUnload the chip for sequencing 3



■ Ion S5™ System component positions . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 42

■ Before you begin . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 43

■ When a manual cleaning of the sequencer is required . . . . . . . . . . . . . . . . . . . . . 43

■ Initialize the sequencer . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 44

■ Reagent consumables disposal . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 45

Initialization takes ~40 minutes.

Ion S5™ System component positions

7

2

3

5

6

4

1

1 Touchscreen2 Power button3 Ion S5™ ExT Sequencing Reagents

cartridge4 Chip clamp

5 Ion S5™ ExT Wash Solution bottle. Wastereservoir located behind the WashSolution bottle (shown on the right).

6 Ion S5™ Cleaning Solution bottle7 Waste reservoir

Note: The system uses RFID technology to verify that the proper reagents are loadedin positions 3, 5, and 6. Reagents that exceed their expiration date or usage countgenerate an error message prompting the user to replace the reagent beforeperforming the run.

Note: RFID regulatory information can be found on the main screen underOptions4Regulatory info.

4


Before you begin

The Ion S5™ and Ion S5™ XL Sequencers are equipped to verify the compatibility ofeach chip and consumable loaded during initialization and sequencing, and that thesecomponents do not exceed their expiration date. To avoid exceptions duringinitialization, inspect this information for each consumable before installing onto theinstrument.

• Unbox the Ion S5™ ExT Sequencing Reagents cartridge 45 minutes before use,then allow it to equilibrate to room temperature.Do not remove the Ion S5™ ExT Sequencing Reagents cartridge from its foilpackaging until immediately before loading, so that you can return the unusedcartridge to storage if your sequencing run is delayed.

• Unbox the Ion S5™ ExT Wash Solution bottle. Invert the bottle 5 times, then swirlat an angle to mix thoroughly.

• Remove the red cap from the Ion S5™ ExT Wash Solution and Ion S5™ CleaningSolution bottles immediately before installing on the instrument.

When a manual cleaning of the sequencer is required

The Ion S5™ and Ion S5™ XL Sequencers require that a cleaning be performed beforeinitialization. This is normally performed automatically at the completion of theprevious sequencing run. When two sequencing runs are performed on a singleinitialization, the post-run cleaning is performed after the second sequencing run.However, if the "Enable post-run clean" checkbox is unchecked to allow a second run,and a second run is not performed, the instrument will not allow the subsequentinitialization to proceed until a manual cleaning has been performed. See “Perform amanual cleaning of the sequencer“ on page 63 for more information on how toperform a manual cleaning.

If an Ion S5™ Sequencer or an Ion S5™ XL Sequencer is initialized and a sequencingrun is not started within 24 hours, or a run is not started or completed due to a powerfailure or an abort, do not perform a manual cleaning. An instrument reset run isrequired before reinitialization. See “Perform an instrument reset run with aninitialized, unused Sequencing Reagents cartridge“ on page 64 for moreinformation.

Chapter 4 Initialize the sequencerBefore you begin 4



1. In the instrument touchscreen main menu, touch Initialize. The door, chip, andReagent cartridge clamps unlock.

2. When prompted, remove the Ion S5™ ExT Wash Solution bottle to access thewaste reservoir, then remove and empty the waste reservoir.

3. Reinstall the empty waste reservoir.

4. Replace the expended Ion S5™ ExT Sequencing Reagents cartridge with a newcartridge equilibrated to room temperature.

5. Ensure the new Ion S5™ ExT Wash Solution bottle is thoroughly mixed. Thenremove the red cap and install.

1

2

3

Chapter 4 Initialize the sequencerInitialize the sequencer4


6. Ensure that the used sequencing chip from the previous run is properly seated inthe chip clamp and the chip clamp is pushed in all the way.

7. If necessary, install a new Ion S5™ Cleaning Solution bottle.

Note: The Ion S5™ Cleaning Solution bottle contains sufficient reagent tocomplete 4 cleanings.

8. Close the door, then touch Next.The instrument confirms that the consumables and chip are properly installedand that the Ion S5™ Cleaning Solution contains sufficient reagent to perform thepost-run clean. Follow all on-screen recommendations to ensure properinstallation of required consumables.

IMPORTANT! If the allowed number of post-run cleans has been met, theinstrument prompts the user to replace the Ion S5™ Cleaning Solution bottle.

Note: See “Bottle leak check fails“ on page 61 if a Checking Reagent: Failedwarning appears.

9. When initialization is complete (~30−40 minutes), touch Home.

The instrument is now ready for a sequencing run.

Reagent consumables disposal

IMPORTANT! Follow all applicable local, state/provincial, and/or national regulationswhen recycling or disposing of Ion S5™ reagent consumables.

To properly dispose of the CO2 scrubber, you must first remove it from the Ion S5™

ExT Sequencing Reagents cartridge.

1. Invert the Ion S5™ ExT Sequencing Reagentscartridge over an appropriate receptacle to drainall residual liquid.

2. Wearing gloves, insert the Ion S5™ Cartridge Toolfirmly into the CO2 scrubber until the flangestops on the top of the cartridge.

CO2 scrubberremoval anddisposal

Ion S5™ Cartridge Tool

Chapter 4 Initialize the sequencerReagent consumables disposal 4


3. Pull straight up on the tool while holding the nucleotide reagent cartridge down.

4. Remove the scrubber from the cartridge tool, then dispose of the scrubberaccording to applicable hazardous waste regulations.

The Ion S5™ ExT Wash Solution and Ion S5™ Cleaning Solution bottles are made ofrecyclable plastic.

1. Open the expended bottle by unscrewing the cap.

2. Remove the cap, sipper, and filter, then pour any residual liquid into anappropriate receptacle.

3. Rinse the empty bottle with water. Pour out the rinse water into the same liquidwaste receptacle.

4. Recycle or dispose of the clean bottle, cap, sipper, and filter according toapplicable regulations.

Recycle Ion S5™

Wash andCleaning solutionbottles

Chapter 4 Initialize the sequencerReagent consumables disposal4



■ Chip handling guidelines . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 47

■ Start the sequencing run . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 48

■ Maintain the sequencer . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 50

Chip handling guidelines

To place a chip in the chip clamp:

1 2 3 4

1 Slide the chip clamp out.2 Remove the chip currently in the clamp.3 Place the appropriate loaded chip in the

chip clamp with the chip notch in thebottom ‑front corner.

Note: Do not force the chip into theclamp. If the chip does not fit easily in theclamp, confirm that the notch is orientedas shown in the drawing.

4 Slide the metal tab in fully to engage theclamp, then close the instrument door.

5



We recommend that you sequence loaded chips on the Ion S5™ or Ion S5™ XLSequencer as soon as possible after chip loading and instrument initialization arecomplete. However, successful sequencing runs can be started up to 24 hours afterinstrument initialization.

IMPORTANT! Do not press the power button during a run. Interrupting power to theinstrument during a run can result in sequencing run failure and loss of sample.

1. After completion of initialization, press Run in the instrument touchscreen. Thedoor and chip clamp unlock.

2. Remove the used sequencing chip, then secure a chip loaded with template-positive Ion Sphere™ Particles in the chip clamp.

3. Push the chip clamp all the way in to engage, close the instrument door, thenpress Next.

4. Confirm that the correct Planned Run has auto-populated, then press Review.

Note: Ensure the Enable post-run clean checkbox is selected so that the post-runcleaning is performed automatically.

5. Confirm the remaining pre-populated settings are correct, or edit using thebuttons and dropdown lists if needed.

Chapter 5 Start the sequencing runStart the sequencing run5


6. Ensure that the instrument door is closed, then press Start run to start thesequencing run.

IMPORTANT! During a run, do not open the instrument door, and avoidtouching the instrument. Touching the instrument during the sequencing run canreduce the quality of the measurements.

When the sequencing run is complete, the instrument automatically performs thecleaning procedure unless the Post-Run/Clean checkbox was deselected. Aftercleaning, the touchscreen returns to the main menu. Use the Torrent Browser toreview the results.

Chapter 5 Start the sequencing runStart the sequencing run 5


Maintain the sequencer

• Lint-free wipes• 70% isopropanol• (optional) 10% bleach solution

In the event of a spill or leak on or inside the instrument, perform the following steps.

Note: Dispose of all waste in appropriate liquid or solid waste containers.

1. Remove the Ion S5™ ExT Wash Solution bottle, then remove and empty the wastereservoir.

2. Remove the Ion S5™ ExT Sequencing Reagents cartridge.

3. Inspect the waste and nucleotide reagent bays for liquid.

4. Using absorbent paper soak up as much liquid as possible, then wash theaffected area with 10% bleach solution.

5. Wipe the affected surfaces with 70% isopropanol, then allow to air-dry.

Requiredmaterials

Clean ordecontaminate thesequencer

Chapter 5 Start the sequencing runMaintain the sequencer5


Clean the Ion Chef™ System

■ About the cleaning protocol . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 51

■ Materials required . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 51

■ Clean the Ion Chef™ Instrument . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 52

About the cleaning protocol

The Ion Chef™ System includes an automated cleaning function that must beperformed following every run. The cleaning routine is initiated from the Ion Chef™

Instrument touchscreen and is designed to minimize potential contamination. Duringthe routine, the instrument irradiates the deck with ultraviolet light for 1 minute afterall consumables have been removed from the instrument.

IMPORTANT! Although the Ion Chef™ Instrument cleaning routine provides someprotection against contamination, it is not a substitute for good laboratory techniqueor precautions. When preparing DNA libraries for use or when preparing theIon Chef™ Instrument, make certain to observe sterile laboratory procedures at alltimes to ensure minimal contamination.

Materials required

• Gloves, powder-free nitrile• Isopropanol, 70% solution• Wipes, lint-free

6


Clean the Ion Chef™ Instrument

IMPORTANT! Clean the Ion Chef™ Instrument as described in the following pagesafter every run. To prevent contamination, do not operate the instrument unless it hasbeen recently cleaned.

4

5

3

2 1

8 7

6

Ion Chef™ Instrument stations1 Waste pipette tip position2 Empty Ion S5™ ExT Tip Cartridge: move to

waste pipette tip station3 Thermal cycler sample block4 Reagents station

5 Solutions station6 Recovery centrifuges7 Enrichment station8 Chip-loading centrifuge

Chapter 6 Clean the Ion Chef™ SystemClean the Ion Chef™ Instrument6


IMPORTANT!· Do not discard the empty Ion S5™ ExT Tip Cartridge.· Make sure to transfer the QC samples before you remove and discard the Reagents

cartridge.

1. Touch (Open Door) in the instrument touchscreen, then wait for the latch toopen.

2. Lift the instrument door to the top of the travel until the latch mechanismengages.

1


3. Remove, then discard the PCR plate from the thermal cycler sample block.

4. Remove, then discard the box of used pipette tips from the waste tip position.

IMPORTANT! Handle the disposable reservoir in the waste tip position withcare. During the run, liquid waste collects in the reservoir. Dispose of the liquidwaste by tipping the reservoir on one corner and pouring the waste into anappropriate waste container:

IMPORTANT! Do not reuse the waste pipette tip rack. Always move the emptyIon S5™ ExT Tip Cartridge from the new tip position to the waste tip position.

Remove anddispose of usedconsumables

Chapter 6 Clean the Ion Chef™ SystemClean the Ion Chef™ Instrument 6


5. Move the empty Ion S5™ ExT Tip Cartridge to the waste tip position.

6. Remove, then discard the• Ion S5™ ExT Chef Reagents cartridge• Ion S5™ Chef Solutions cartridge• Enrichment Cartridge v2

7. Remove, then discard the consumables from the Recovery centrifuges, includingthe:

• Recovery Station Disposable Lids v2• Recovery Tubes v2

8. Close the Chip-loading centrifuge lid.

1. Inspect the Recovery centrifuges, then clean the components if excess liquid ispresent.

Is liquidpresent? Action

No Go to “Start the cleaning“ on page 55.

Yes Clean the centrifuge bowl and buckets as described below.

IMPORTANT! Clean the Recovery centrifuges occasionally, onlywhen excess liquid is noticeable in the bowl and/or buckets. Youdo not need to clean the centrifuge after every run.

IMPORTANT! Wear powder-free, nitrile gloves when cleaning the Recoverycentrifuge.

2. Remove the buckets from the Recovery centrifuges. Clean the inside and outsideof each bucket using a lint-free wipe, then place the buckets on a clean, drysurface while you clean the centrifuge.

1

2

1 Bucket2 Lint-free wipe

Inspect and cleanthe Recoverycentrifuges andbuckets



3. Use lint-free wipes to remove all fluid from inside the centrifuge bowl.

1

2

1 Inside rim of the centrifuge2 Bottom of the centrifuge bowls

4. Use lint-free wipes treated with 70% isopropanol to clean the:• Inside rim of the centrifuge.• Bottom of the centrifuge bowl.• Outside and inside of the centrifuge buckets.

5. Dry the centrifuge and buckets with lint-free wipes.

6. Install the centrifuge buckets, then close the Recovery centrifuge cover.

1

1 Buckets (cleaned and installed)

1. Close the instrument door by first lifting it up slightly to disengage the lockingmechanism, then pushing down on the door until the locks engage.

IMPORTANT! Before closing the door, ensure the covers of the centrifuges areclosed.

Start the cleaning

Chapter 6 Clean the Ion Chef™ SystemClean the Ion Chef™ Instrument 6


2. To start the cleaning, touch Next on the Ion Chef™ Instrument touchscreen thatappears after run completion.

Note: You can also clean the instrument at any time starting from the hometouchscreen. Touch Settings4Clean Ion Chef.

3. Confirm that you have removed all consumables from the Ion Chef™ Instrument,except the empty pipette tip rack in the waste tip position, then touch Next.

4. With the door closed, touch Start. The instrument performs a Deck Scan beforestarting the cleaning routine. The Ion Chef™ Instrument stops ventilation, thenilluminates the ultraviolet (UV) light in the instrument for ~1 minute.

CAUTION! The Ion Chef™ Instrument emits UV light at 254 nm. Wearappropriate eye wear, protective clothing, and gloves when working nearthe instrument. Do not look directly at the UV light while it is illuminatedduring the cleaning routine.



Troubleshooting

■ Ion Chef™ System setup and operation . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 57

■ Ion S5™ System setup and operation . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 60

Ion Chef™ System setup and operation

Observation Possible cause Recommended action

Instrument does not display thehome screen when powered on

Multiple causes are possible. 1. Power off the instrument, wait30 seconds, then power on theinstrument.

2. If the instrument fails again, contactTechnical Support.

Cannot open the instrumentdoor

An obstruction is present on oraround the door mechanism.

Remove the obstruction blocking the door, thenoperate the instrument normally.

A hardware or software errorhas occurred.

Contact Technical Support to report theproblem and for further assistance.

Instrument stops during a run The instrument hasencountered an internal error.

1. Record the error displayed on theinstrument display, then touch OK.

2. Contact Technical Support to report theproblem and for further assistance.

A consumable was improperlyloaded.

Ensure that all consumables are loadedaccording to the instructions provided.

Liquid residue is present in theRecovery centrifuge following arun

During normal instrumentoperation, a noticeable coatingof liquid can collect on the bowland buckets of the Recoverycentrifuge following repeatedruns.

Remove the residue as instructed in “Clean theIon Chef™ Instrument“ on page 52.

A



Instrument will not begin a run The instrument hasencountered a Deck Scan error(one or more consumables areabsent or loaded improperly).

1. Confirm that the touchscreen does notdisplay any Deck Scan warnings. If alarmsare present, note the error(s) displayed,replace the missing consumable asdirected, press No when prompted thenpress Next to cancel the run. Afterreturning to the home screen, restart therun.

2. If the error persists, confirm that:

• All buckets are seated correctly inthe rotors of the Recovery and Chip-Loading Centrifuges.

• All cartridges are loaded correctlyand are level on the instrument deck.

• The barcodes of the Ion Chef™

Library Sample Tubes are visible andpositioned correctly.

• All tubes are both present anduncapped on the Ion S5™ ExT ChefReagents (Library Sample Tubess,NaOH tube, and the empty tube).

3. If the error persists after you check theconsumables on the instrument deck, doone of the following:

• If you are confident that the IonChef™ Instrument is set up correctlyand you are comfortable disregardingthe warnings, touch YES followingDeck Scan to proceed with the run.

• If the instrument cannot begin therun, contact Technical Support forfurther assistance.

The instrument hasencountered an internal error.

1. Record the error displayed on theinstrument display, then touch OK.

2. Contact Technical Support to report theproblem and for further assistance.

Appendix A TroubleshootingIon Chef™ System setup and operationA



Instrument displays one ormore alerts during a run

The instrument has detectedone or more problems duringthe run.

After the instrument completes the run,contact Technical Support.

IMPORTANT! The detected problem mightimpact the performance of the sequencing run.

• Network connection to theserver has beeninterrupted.

• User name or password isincorrect.

1. Touch the Instrument status button toview the alert(s).

2. In the Instrument status screen, confirmthat the name of the Torrent Serverconnection is red.

3. Contact your network administrator toconfirm that:

• The Torrent Server can be accessedfrom the network port used by theIon Chef™ Instrument. If not,troubleshoot the network connection.

• The user name and password usedby the Ion Chef™ Instrument arevalid. If not, contact the serveradministrator to renew thecredentials.

4. If the alert persists, contact TechnicalSupport for further assistance.

Appendix A TroubleshootingIon Chef™ System setup and operation A


Ion S5™ System setup and operation


Red "Alarms" and/or "Events" message in MainMenu

• Softwareupdatesavailable

• Connectivityissues

• Instrument notdetectingrequired filesor hardware

Click on the red pop-up to see detailedmessages.

• If a message states "NewerSoftware Available":

IMPORTANT! After updates areinstalled, the instrument must berestarted.

a. On the home screen, selectSettings4Check for Updates.

b. Select the Released Updatescheckbox, then press Update.

c. When installation is complete,follow the onscreen promptsto restart the instrument.

Note: In some cases, theinstrument restartsautomatically after softwareinstallation.

• If a message states "NoConnectivity to Torrent Server", "NoConnectivity to ftp server", or"Network Manager not connected",disconnect and re-connect theethernet cable, confirm that therouter is operational, and verifythat the network is up and running.

• For any other messages:a. Power off the instrument: On

the home screen, selectSettings4System Tools4ShutDown4Shut Down.

b. Wait 30 seconds, then pressthe button on the front ofinstrument to power on theinstrument.

• If the red "Alarms" and/or "Events"message still appears in the mainmenu, contact Technical Support.

Ion S5™

Sequencer alarmsand events

Appendix A TroubleshootingIon S5™ System setup and operationA



Bottle leak check fails • Bottle seal is not tight.

• Bottle may be damaged /defective.

1. Finger-tighten the bottles.2. If the bottle continues to leak, replace the

bottle.3. If leak check continues to fail, contact

Technical Support.

Initialization—General errors

Appendix A TroubleshootingIon S5™ System setup and operation A


Supplemental procedures

■ Quality control of Ion Sphere™ Particles . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 62

■ Perform a manual cleaning of the sequencer . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 63

■ Perform an instrument reset run with an initialized, unused SequencingReagents cartridge . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 64

■ Maintain the Ion Chef™ System . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 65

Quality control of Ion Sphere™ Particles

The Guava™ easyCyte™ 5 Flow Cytometer can be used for quality assessment ofunenriched and enriched Ion Sphere™ Particles generated for up to 400 to 600 base-read sequencing on the Ion S5™ Sequencer or Ion S5™ XL Sequencer.

• Unenriched sample – Obtain the QC sample from the Library Sample Tube onthe Ion S5™ ExT Chef Reagents cartridge at position A.

• Enriched sample – Obtain enriched sample from the tube on the EnrichmentCartridge v2 at position E.

For details, see the Ion PI™ Ion Sphere™ Particles Quality Assessment Using the Guava™

easyCyte™ 5 Flow Cytometer User Bulletin (Pub. No. MAN0007496), available at thermofisher.com.

The Applied Biosystems™ Attune™ Acoustic Focusing Cytometer can be used forquality assessment of unenriched and enriched Ion Sphere™ Particles generated for upto 400 to 600 base-read sequencing on the Ion S5™ Sequencer or Ion S5™ XL Sequencer.

• Unenriched sample – Obtain the QC sample from the Library Sample Tube onthe Ion S5™ ExT Chef Reagents cartridge at position A.

• Enriched sample – Obtain enriched sample from the tube on the EnrichmentCartridge v2 at position E.

For details, refer to the Demonstrated Protocol: Ion Sphere™ Particles (ISPs) QualityAssessment using the Applied Biosystems™ Attune™ Acoustic Focusing Cytometer UserBulletin (Pub. No. 4477181), available at thermofisher.com.

B

Quality controlusing the Guava™

easyCyte™ 5 FlowCytometer

Quality controlusing the Attune™

Acoustic FocusingCytometer


http://www.thermofisher.com/

http://www.thermofisher.com/

Perform a manual cleaning of the sequencer

A cleaning protocol is normally performed automatically at the completion of eachsequencing run. In the event that a cleaning is necessary, follow the listed steps:

1. On the home screen, select Settings4Clean Instrument.The instrument door unlocks allowing access to the consumables.

2. Remove the Ion S5™ ExT Wash Solution bottle to access the waste reservoir, thenremove and empty the waste reservoir.

1

2

3

3. Reinstall the empty waste reservoir and a used Ion S5™ ExT Wash Solution bottle.

4. Ensure the Ion S5™ ExT Sequencing Reagents cartridge and Ion S5™ ExT WashSolution bottle are properly installed.

IMPORTANT! Perform the cleaning with a used reagent cartridge and washsolution bottle installed. The cleaning procedure pumps cleaning solution intothe wash solution bottle and reagent cartridge making them unsuitable forsequencing.

5. Place a used sequencing chip in the chip clamp, then push the chip clamp in allthe way to engage.

6. Close the instrument door, then press Next.Cleaning takes ~35 minutes to complete. Upon completion the instrument doorautomatically unlocks and the chip and cartridge clamps disengage.

7. Proceed to Chapter 4, “Initialize the sequencer“.

Appendix B Supplemental proceduresPerform a manual cleaning of the sequencer B


Perform an instrument reset run with an initialized, unusedSequencing Reagents cartridge

Cleaning is normally performed at completion of a sequencing run automatically. Ifan Ion S5™ Sequencer or an Ion S5™ XL Sequencer is initialized and

• a sequencing run is not started within 24 hours after initialization, or• a sequencing run is not completed due to a power failure or an abort, and <200flows occurred before the stoppage

an instrument reset run is required to ensure proper cleaning before reinitialization.Do NOT perform a manual cleaning with an unused, initialized Ion S5™ ExTSequencing Reagents cartridge.

Note:· If a power failure or abort occurs during the second of two runs started after a

single initialization, a manual cleaning (page 63) is sufficient.· If the number of flows that occurred before a power failure or abort is unknown,

perform an instrument reset run.

To perform an instrument reset run, use the following procedure beforereinitialization:

1. In the instrument touchscreen main menu, press Run.The instrument door and chip clamp unlocks.

2. Ensure that a used sequencing chip is in the chip clamp, then push the chipclamp in all the way to engage.

3. Close the instrument door, then press Next.

4. When prompted, select Planned Run (none). Ensure that the Enable post-runclean checkbox is selected, then press Review.

5. In the Select Run screen, press Edit, then in the Detail screen set the number offlows to 200 manually. Ensure that the Post-Run/Clean checkbox is selected, thenpress Close.

6. Press Start run, then press Accept to confirm that Post-Run Clean is enabled, andto start the run.

When the instrument reset run completes, the instrument automatically performs thecleaning procedure. After cleaning, the touchscreen returns to the main menu.

Appendix B Supplemental proceduresPerform an instrument reset run with an initialized, unused Sequencing Reagents cartridgeB


Maintain the Ion Chef™ System

The Ion Chef™ System supports the user-performed maintenance procedures found inthis section.

To ensure proper operation of the Ion Chef™ Instrument, we recommend periodicallyconfirming that your instrument is running the most current firmware. Weoccasionally release updates to the instrument firmware, which can include importantchanges to the Ion Chef™ System operation. To ensure that your instrument is runningthe most current firmware, check the firmware version as explained below.

Check the Ion Chef™ Instrument firmware

1. On the Ion Chef™ Instrument touchscreen, touch Settings.

2. On the Settings screen, touch Check for updates.

3. On the Software Update screen, touch Release to search the server for updates tothe Ion Chef™ Instrument firmware.

Note: Users are also notified of the availability of firmware updates on theNotifications screen.

4. Select the available component update, touch Update, then wait for theinstrument to complete the update.

Note: If the Ion Chef™ Instrument is operating on an isolated network andcannot connect to our website, you must transfer the firmware update manually.To perform a manual update, transfer the firmware files to a USB drive and insertthe drive into the USB port at the rear of the instrument. Touch USB on theSoftware Update screen, select from the list of available software components,then touch Update.

When finished, the Ion Chef™ Instrument displays the update status.

5. Power off and on the Ion Chef™ Instrument to complete the update.

IMPORTANT! If you are updating the firmware manually, remove the USB drivebefore powering on the Ion Chef™ Instrument.

Install a firmwareupdate

Appendix B Supplemental proceduresMaintain the Ion Chef™ System B


The following procedure describes how to change the name used to identify the IonChef™ Instrument both on the network and in the data that it generates.

1. In the Ion Chef™ Instrument home screen, touch Settings.

2. In the Settings screen, touch Instrument settings.

3. In the Instrument settings screen, touch Set Instrument Name. Touch the namefield and enter a new instrument name using the keypad that appears, thentouch Save.

Note: Use only alphanumeric characters. Do not use special characters or spacesin the name.

4. In the next screen, touch Save again. Power off and on the Ion Chef™ Instrumentto effect the name change.

Change theinstrument name

Appendix B Supplemental proceduresMaintain the Ion Chef™ SystemB


Ion Chef™ and Torrent Servernetwork setup

■ Networking overview . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 67

■ Ion Chef™ Networking Configuration Options . . . . . . . . . . . . . . . . . . . . . . . . . . . 71

■ Editing the Torrent Server firewall tables to allow the LAN port IP . . . . . . . . . 75

■ Enable Planned Run sharing among Torrent Servers . . . . . . . . . . . . . . . . . . . . . . 77

■ Transfer a Planned Run . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 79

■ Undo a Planned Run transfer (administrator) . . . . . . . . . . . . . . . . . . . . . . . . . . . . 81

■ Undo a Planned Run transfer (user) . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 81

Networking overview

If you are preparing a laboratory for use with new Ion instruments, refer to therespective site preparation and safety guides. If you currently have an Ion Chef™

instrument configured in your laboratory to communicate to more than one TorrentServer, Torrent Suite™ Software v4.4 or later requires the configuration of theIon Chef™ instrument to communicate with one primary Torrent Server. Afterupgrading to Torrent Suite™ Software v4.4 or later, follow the guidelines in thisdocument to configure the instrument to communicate with a primary Torrent Server.From that Torrent Server, the new Planned Run sharing option can be used to targetadditional Torrent Servers if needed.

Networking with the Torrent Server running Torrent Suite™ Assay DevelopmentSoftware and the Ion PGM™ Dx System requires v5.0 or later. The Ion Chef™ will notconnect to the Ion PGM™ Dx in IVD mode.

If you need to connect a Torrent Server to the network for the first time, see theTorrent Server networking requirements in the site prep and safety guide for therespective sequencer.

This guide describes how to:• Set up the Ion Chef™ instrument to talk to the Torrent Server on a local area

network (LAN).• Enable Planned Run sharing.

C


The instrument is factory-configured for IPv4 TCP/IP communication and includes afast Ethernet adapter (1000 Mbps) with a RJ45-type connector for integrating thedevice into a local area network (LAN). If the instrument will be connected to a LAN,an active network port must be available/assigned in place before the scheduledinstallation date.

If you are activating a newly installed LAN port for your Ion Chef™ instrument,please ensure the LAN has been configured to permit HTTP-443, SSH-22, andFTP-20/21 traffic. For additional information, please refer to the NetworkingRequirements section of the Ion Chef™ Site Preparation Guide (Pub. no. MAN0007956).

The Torrent Server is an integral part of the system and includes a quad-port gigabitNIC for direct communication with the network or instruments such as Ion Chef™,PGM™, Ion Proton™, or Ion S5™ instruments. When the Torrent Server is connected tothe network, Torrent Servers that are on the same software versions and subnet cantransfer planned runs from one Torrent Server to another.

We recommend that you consult a network administrator before connecting theIon Chef™ instrument to your laboratory network. Please refer to the networkingrequirements in the Ion Chef™ System Site Preparation Guide.

Collect the required network information.

q

Connect the Ion Chef™ instrument to the configured LAN port.q

Configure the Ion Chef™ instrument with the required information.q

Monitor the Ion Chef™ instrument to test the network connection.

This guide provides a basic description of how to integrate the Torrent Server and theIon Chef™ instruments into several possible network architectures. Because yournetwork may contain advanced features, such as a firewall or network domains, werecommend that you consult a network administrator before connectingIon Chef™instrument to your laboratory network.

Network examples include:• Direct connection• Static IP (indirect connection)• DHCP IP (indirect connection)

Obtain the following information from your network administrator:• Network policy for obtaining IP addresses (DHCP IP or static IP).• If the network requires static IP addresses, obtain the static IP address, subnet

mask, gateway address, and DNS server name for the LAN port.• For the Planned Run sharing and FTP functionality to be successful, the subnet

must be the same on all Torrent Servers and Ion Chef™ instruments on the LAN.

About theIon Chef™

instrument

About the TorrentServer

Networkingguidelines andbest practices

Network setupworkflow

Network setupexamples

Collect therequired networkinformation

Appendix C Ion Chef™ and Torrent Server network setupNetworking overviewC


An ethernet cable with RJ45 connectors (CAT6 Ethernet cable for a 1000Mbit/snetwork connection or a CAT5 for 100Mbit/s connection).

See Ion Chef™ System Site Preparation Guide (Pub. no. MAN0007956) for additionalinformation.

See Ion Chef™ System Site Preparation Guide (Pub. no. MAN0007956).

DHCP IP

Ion Chef

Network

DHCP Server

Torrent Server

If using DHCP for the LAN port, please make sure the port is set to auto-populatewhen the Ion Chef™ instrument is connected and ensure that all the belowrequirements are met.

Static IP

Ion Chef

Network

Torrent Server

If the network requires static IP addresses, obtain the static IP address, subnet mask,gateway address, and DNS server name for the LAN port.

For planned run sharing and FTP functionality to be successful, the subnet must bethe same on all Torrent Servers and Ion Chef™ instruments on the LAN.

Materials required

Generalrequirements

Appendix C Ion Chef™ and Torrent Server network setupNetworking overview C


Direct connection

Ion Chef

Network

Torrent Server

To configure the direct connection between an Ion Chef™ instrument and a TorrentServer, you will need a Category 6 cable of sufficient length.

Another direct connection configuration option consists of one Ion Chef™ instrument,one Torrent Server and two Ion S5™ XL Sequencers. However, this configuration is notrecommended due to increased analysis times.

WARNING! The direct configuration shown below is known to result inlengthy analysis times.

Ion Chef

Network

Torrent Server

S5 XL 1 S5 XL 2

Appendix C Ion Chef™ and Torrent Server network setupNetworking overviewC


Ion Chef™ Networking Configuration Options

With the Torrent Suite™ software v4.4 release, the Ion Chef™ software will onlyconnect to one Torrent Server. From that server, Planned Run sharing can beconfigured to access additional servers. When the Ion Chef™ is connected to a TorrentServer, an icon on the lower left side of the screen appears.

Connect the Ion Chef™ Instrument to the Torrent Server in one of three ways: Direct,Static IP (non-direct) or DHCP IP (non-direct). See respective instructions below.

To target the Torrent Server IP address from the Ion Chef™user interface, you will firstlook up the Torrent Server IP address and then enter it into the Ion Chef™userinterface as the Torrent Browser target and the Update Target.

1. Log into Torrent Browser as the admin.

2. Click the Gear button and select Configure.

Target the TorrentServer IP Address

Appendix C Ion Chef™ and Torrent Server network setupIon Chef™ Networking Configuration Options C


3. Scroll down to the Database Administration section and click the AdminInterface link.

Appendix C Ion Chef™ and Torrent Server network setupIon Chef™ Networking Configuration OptionsC


4. The Site Administration screen appears. Scroll to the bottom and click ViewNetwork Settings.

Appendix C Ion Chef™ and Torrent Server network setupIon Chef™ Networking Configuration Options C


5. Note (copy or write down) the Torrent Server IP address displayed on theNetwork Settings screen.

6. Go back to the Ion Chef™ user interface and tap on the Torrent Browser IPaddress. This opens the Manage Connections window.

Appendix C Ion Chef™ and Torrent Server network setupIon Chef™ Networking Configuration OptionsC


7. Enter the server IP address under Server and repeat for Update Site. Click Save.

Editing the Torrent Server firewall tables to allow the LAN port IP

In addition, when connecting through a LAN, the Torrent Server firewall settings willneed to be modified for both DHCP and Static LAN connections.

Note: While directly connected, instruments are on the 192.168.20x.x subnet fromwhich all traffic is enabled, so this is not required.

Any exception would mean that editing the local firewall tables to include the LAN IPis required. Also, if the Ion Chef™ instrument is getting a DHCP address from yournetwork, you will need to create an exception for the entire DHCP subnet.

1. On the Torrent Server, edit the /etc/iptables.custom file and add a firewall rule toallow all traffic from Ion Chef™ instrument's IP address as shown below. (This isa blank file initially and is intended for custom firewall configuration.)

IMPORTANT! Only edit the /etc/iptables.custom file and not /etc/iptables.rules,since /etc/iptables.rules is overwritten when software is updated.

2. Enter "sudo^nano^/etc/iptables.custom" and add the three lines below:• *filter• -A INPUT -s xxx.xxx.xxx.xxx -j ACCEPT• COMMIT

Create anexception

Appendix C Ion Chef™ and Torrent Server network setupEditing the Torrent Server firewall tables to allow the LAN port IP C


Note: -A INPUT -s xxx.xxx.xxx.xxx -j ACCEPT adds the rule to theformat. Enter the IP address of the Ion Chef™ instrument in place of"xxx.xxx.xxx.xxx". When finished editing, save changes by pressing CTRL +O and CTRL + X.

3. Follow this command by manually applying the change:sudoîptables-restore^--noflush^<^/etc/iptables.customNote: You can also list the firewall rules with: sudoîptables-L.

If this command returns with no output, it means it succeeded.

4. If the Ion Chef™ instrument is getting a DHCP address from your network,please create the exception for the entire DHCP subnet. The example belowassumes that the DHCP address for the Ion Chef™ instrument is 172.16.48.0 andthe DHCP subnet range is 172.16.48.1 to 172.16.48.254. Add these 3 lines:

• *filter• -A INPUT -s 172.16.48.0/24 -j ACCEPT• COMMIT

5. Reboot the server.

6. If you want to repeat this process and completely drop all rules and start fresh,enter:

• sudoîptables-F• sudoîptables-restore^<^/etc/iptables.rules• sudoîptables-restore--noflush^</êtc/iptables.custom

Removing a rule from either of these files would remove it from the activefirewall rules, because the iptables-restore command does not overwriterules; it appends them to all active rules.

Appendix C Ion Chef™ and Torrent Server network setupEditing the Torrent Server firewall tables to allow the LAN port IPC


Enable Planned Run sharing among Torrent Servers

New in Torrent Suite™ v4.4, Planned Runs created on one Torrent Server can now betransferred to another Torrent Server if the sequencer connected directly to the serveris offline or busy. Possible configurations include:

Ion Chef™ instrument 1 → Torrent Server 1 ↔ Torrent Server 2 → Sequencer 2

Ion Chef™ instrument 1 → Torrent Server 1 ↔ Torrent Server 3 → Sequencer 3

Network

TS 1(origin)

IC 2IC 1 S5 XL 1 S5 XL 2 IC 3

TS 2(destination)

TS 3(destination)

S5 XL 3

Requirements include:• All Torrent Servers must be running the same software version.• All Torrent Servers must have the same genomic reference, barcode set, BED files,

Variant Caller config files, etc. All Torrent Servers must be on the same subnet.

Appendix C Ion Chef™ and Torrent Server network setupEnable Planned Run sharing among Torrent Servers C


1. On the origin server (e.g., TS1) Site administration page, scroll down and selectShared servers. The Select shared server to change window appears.

2. If you are adding your destination server for the first time, click Add sharedserver.

3. Define your destination server.a. Enter the name, address (can be IP address), user name and password for

the destination server.

b. Click Active if you want this server enabled for sharing.

c. (Optional) Add a comment.

d. Click one of the Save options.

4. (Optional) If you want to configure the origin Torrent Server to also be adestination server, you must go to another server and repeat these steps to set theorigin server as a destination server. Once the Torrent Servers are configured, youor a user can now transfer Planned Runs between Torrent Servers.

Set up ServerNetwork (adminaction)

Appendix C Ion Chef™ and Torrent Server network setupEnable Planned Run sharing among Torrent ServersC


Transfer a Planned Run

You can execute a Planned Run on the Ion Chef™ instrument connected to the originserver, and then, when the run is complete, transfer the Planned Run to a destinationserver for sequencing.

Alternatively, you can create the Planned Run on an origin server and then transfer itto a destination server for both the Ion Chef™ run and the sequencing run.

1. Using the Torrent Browser on the origin Torrent Server, go to Plan4Planned RunList.

2. Open the Gear menu of the Planned Run you want to transfer, and selectTransfer. Then select the destination Torrent Server.

3. A confirmation window appears. Check the information, then click Transfer.

Note: You can no longer access this Planned Run on the origin server after it hastransferred.

Appendix C Ion Chef™ and Torrent Server network setupTransfer a Planned Run C


4. A status window displays the results of the transfer:• The green box lists the samples successfully processed and the required

target BED files found on the destination server.• The red box lists any required BED files or plugins that are not present on

the destination server. To successfully perform the run, you will need to editthe transferred Planned Run on the destination server and manually add themissing BED files or plugins.

5. To edit the transferred Planned Run and add missing files:a. Download required files using the References tab of the Torrent browser of

the destination server

b. Go to the Edit Plan wizard of the transferred Planned Run by selecting Editon the gear pull-down menu to the right of the Planned Run.

c. Select the files or plugins as needed, then click Update Plan.

Note: You can also navigate to the Edit Plan wizard by clicking the Edit testplan link in the status page above.

Appendix C Ion Chef™ and Torrent Server network setupTransfer a Planned RunC


Undo a Planned Run transfer (administrator)

1. On the destination server, delete the transferred Planned Run from either thePlanned Run page or the admin page.

2. On the origin server, locate the plan onthe /admin/rundb/plannedexperiment/page, uncheck PlanExecuted and changePlanStatus to Planned.

Undo a Planned Run transfer (user)

If you transferred a Planned Run in error, you can transfer it back to the origin serveror to another server.

1. On the destination Torrent Server, navigate to Plan4Planned Run List andlocate the transferred Planned Run.

2. From the Gear menu of the Planned Run, select Transfer, then select theTorrent Server to which you wish to transfer the run.

Appendix C Ion Chef™ and Torrent Server network setupUndo a Planned Run transfer (administrator) C


Safety

WARNING! GENERAL SAFETY. Using this product in a manner not specifiedin the user documentation may result in personal injury or damage to theinstrument or device. Ensure that anyone using this product has receivedinstructions in general safety practices for laboratories and the safetyinformation provided in this document.

· Before using an instrument or device, read and understand the safetyinformation provided in the user documentation provided by themanufacturer of the instrument or device.

· Before handling chemicals, read and understand all applicable Safety DataSheets (SDSs) and use appropriate personal protective equipment (gloves,gowns, eye protection, etc). To obtain SDSs, see the “Documentation andSupport” section in this document.

Symbols on this instrument

Symbols may be found on the instrument to warn against potential hazards or conveyimportant safety information. In this document, the hazard symbol is used along withone of the following user attention words:

• CAUTION! – Indicates a potentially hazardous situation that, if not avoided,may result in minor or moderate injury. It may also be used to alert againstunsafe practices.

• WARNING! – Indicates a potentially hazardous situation that, if not avoided,could result in death or serious injury.

• DANGER! – Indicates an imminently hazardous situation that, if not avoided,will result in death or serious injury.

Symbol English Français

Caution, risk of danger

Consult the manual for further safetyinformation.

Attention, risque de danger

Consulter le manuel pour d’autresrenseignements de sécurité.

Protective conductor terminal (mainground)

Borne de conducteur de protection(mise à la terre principale)

D


Symbol English Français

Do not dispose of this product inunsorted municipal waste

CAUTION! To minimizenegative environmentalimpact from disposal ofelectronic waste, do notdispose of electronic waste inunsorted municipal waste.Follow local municipal wasteordinances for properdisposal provision andcontact customer service forinformation about responsibledisposal options.

Ne pas éliminer ce produit avec lesdéchets usuels non soumis au trisélectif.

CAUTION! Pour minimiserles conséquences négativessur l’environnement à la suitede l’élimination de déchetsélectroniques, ne paséliminer ce déchetélectronique avec les déchetsusuels non soumis au trisélectif. Se conformer auxordonnances locales sur lesdéchets municipaux pour lesdispositions d’élimination etcommuniquer avec le serviceà la clientèle pour desrenseignements sur lesoptions d’éliminationresponsable.

The following table applies only to the Ion Chef™ Instrument.

Conformitymark Description

Indicates conformity with safety requirements for Canada and U.S.A.

Indicates conformity with European Union requirements for safety andelectromagnetic compatibility.

Indicates conformity with Australian standards for electromagneticcompatibility.

Conformitysymbols

Appendix D SafetySymbols on this instrument D


Location of safety labels on this instrument

Safety information for instruments not manufactured by ThermoFisher Scientific

Some of the accessories provided as part of the instrument system are not designed orbuilt by Thermo Fisher Scientific. Consult the manufacturer's documentation for theinformation needed for the safe use of these products.

Instrument safety

CAUTION! Do not remove instrument protective covers. If you remove theprotective instrument panels or disable interlock devices, you may be exposedto serious hazards including, but not limited to, severe electrical shock, laserexposure, crushing, or chemical exposure.

CAUTION! Moving Parts. Moving parts can crush, pinch and cut. Keep handsclear of moving parts while operating the instrument. Disconnect power beforeservicing.

General

Physical injury

Appendix D SafetyLocation of safety labels on this instrumentD


WARNING! Ensure appropriate electrical supply. For safe operation of theinstrument:

· Plug the system into a properly grounded receptacle with adequate currentcapacity.

· Ensure the electrical supply is of suitable voltage.· Never operate the instrument with the ground disconnected. Grounding

continuity is required for safe operation of the instrument.

WARNING! Power Supply Line Cords. Use properly configured and approvedline cords for the power supply in your facility.

WARNING! Disconnecting Power. To fully disconnect power either detach orunplug the power cord, positioning the instrument such that the power cord isaccessible.

CAUTION! Cleaning and Decontamination. Use only the cleaning anddecontamination methods specified in the manufacturer's user documentation.It is the responsibility of the operator (or other responsible person) to ensurethe following requirements are met:

· No decontamination or cleaning agents are used that could cause aHAZARD as a result of a reaction with parts of the equipment or withmaterial contained in the equipment.

· The instrument is properly decontaminated a) if hazardous material isspilled onto or into the equipment, and/or b) prior to having the instrumentserviced at your facility or sending the instrument for repair, maintenance,trade-in, disposal, or termination of a loan (decontamination forms may berequested from customer service).

· Before using any cleaning or decontamination methods (except thoserecommended by the manufacturer), users should confirm with themanufacturer that the proposed method will not damage the equipment.

The Ion Chef™ System uses a UV lamp which emits light at 254 nm. Under normaloperating conditions, the UV lamp is powered on when performing the cleaningprotocol. Safety interlocks are used to ensure that the UV lamp is not powered whenthe door is open.

Safety and electromagnetic compatibility (EMC) standards

The instrument design and manufacture complies with the standards andrequirements for safety and electromagnetic compatibility as noted in the followingtable:

Electrical

Cleaning anddecontamination

Ultraviolet (UV)Safety

Appendix D SafetySafety and electromagnetic compatibility (EMC) standards D


Reference Description

EU Directive2006/95/EC

European Union “Low Voltage Directive”

IEC 61010-1

EN 61010-1

UL 61010-1

CSA C22.2 No.61010-1

Safety requirements for electrical equipment for measurement,control, and laboratory use – Part 1: General requirements

IEC 61010-2-010

EN 61010-2-010

Safety requirements for electrical equipment for measurement,control and laboratory use – Part 2-010: Particular requirementsfor laboratory equipment for the heating of materials

IEC/EN 61010-2-020 Safety requirements for electrical equipment for measurement,control and laboratory use – Part 2-020: Particular requirementsfor laboratory centrifuges

IEC 61010-2-081

EN 61010-2-081

Safety requirements for electrical equipment for measurement,control and laboratory use – Part 2-081: Particular requirementsfor automatic and semi-automatic laboratory equipment foranalysis and other purposes


Directive 2004/108/EC European Union “EMC Directive”

FCC Part 15 U.S. Standard “Industrial, Scientific, and Medical Equipment”

AS/NZS 2064 Limits and Methods of Measurement of ElectromagneticDisturbance Characteristics of Industrial, Scientific, andMedical (ISM) Radiofrequency Equipment

ICES-001, Issue 3 Industrial, Scientific and Medical (ISM) Radio FrequencyGenerators


Directive 2012/19/EU European Union “WEEE Directive” – Waste electrical andelectronic equipment

Directive 2011/65/EU European Union “RoHS Directive” – Restriction of hazardoussubstances in electrical and electronic equipment

Instrument safety — Ion S5™/Ion S5™ XL System

For detailed information on Ion S5™ and Ion S5™ XL instrument safety symbols andalerts, safety and electromagnetic compatibility standards, and general instrumentsafety, see the Safety appendix of the Ion S5™ and Ion S5™ XL Instrument User Guide(Pub. No. MAN0010811), available at thermofisher.com.

Safety compliance

EMC

Environmentaldesign

Appendix D SafetyInstrument safety — Ion S5™/Ion S5™ XL SystemD


https://www.thermofisher.com

Chemical safety

WARNING! GENERAL CHEMICAL HANDLING. To minimize hazards,ensure laboratory personnel read and practice the general safety guidelines forchemical usage, storage, and waste provided below. Consult the relevant SDSfor specific precautions and instructions:

· Read and understand the Safety Data Sheets (SDSs) provided by thechemical manufacturer before you store, handle, or work with any chemicalsor hazardous materials. To obtain SDSs, see the “Documentation andSupport” section in this document.

· Minimize contact with chemicals. Wear appropriate personal protectiveequipment when handling chemicals (for example, safety glasses, gloves, orprotective clothing).

· Minimize the inhalation of chemicals. Do not leave chemical containers open.Use only with adequate ventilation (for example, fume hood).

· Check regularly for chemical leaks or spills. If a leak or spill occurs, followthe manufacturer's cleanup procedures as recommended in the SDS.

· Handle chemical wastes in a fume hood.· Ensure use of primary and secondary waste containers. (A primary waste

container holds the immediate waste. A secondary container contains spillsor leaks from the primary container. Both containers must be compatiblewith the waste material and meet federal, state, and local requirements forcontainer storage.)

· After emptying a waste container, seal it with the cap provided.· Characterize (by analysis if necessary) the waste generated by the particular

applications, reagents, and substrates used in your laboratory.· Ensure that the waste is stored, transferred, transported, and disposed of

according to all local, state/provincial, and/or national regulations.· IMPORTANT! Radioactive or biohazardous materials may require special

handling, and disposal limitations may apply.

Appendix D SafetyChemical safety D


Biological hazard safety

WARNING! BIOHAZARD. Biological samples such as tissues, body fluids,infectious agents, and blood of humans and other animals have the potential totransmit infectious diseases. Conduct all work in properly equipped facilitieswith the appropriate safety equipment (for example, physical containmentdevices). Safety equipment can also include items for personal protection, suchas gloves, coats, gowns, shoe covers, boots, respirators, face shields, safetyglasses, or goggles. Individuals should be trained according to applicableregulatory and company/ institution requirements before working withpotentially biohazardous materials. Follow all applicable local, state/provincial,and/or national regulations. The following references provide generalguidelines when handling biological samples in laboratory environment.

· U.S. Department of Health and Human Services, Biosafety in Microbiologicaland Biomedical Laboratories (BMBL), 5th Edition, HHS Publication No. (CDC)21-1112, Revised December 2009; found at:www.cdc.gov/biosafety/publications/bmbl5/BMBL.pdf

· World Health Organization, Laboratory Biosafety Manual, 3rd Edition,WHO/CDS/CSR/LYO/2004.11; found at:www.who.int/csr/resources/publications/biosafety/Biosafety7.pdf

Appendix D SafetyBiological hazard safetyD


http://www.cdc.gov/biosafety/publications/bmbl5/BMBL.pdf

http://www.who.int/csr/resources/publications/biosafety/Biosafety7.pdf

Documentation and support

Customer and technical support

Visit thermofisher.com/support for the latest in services and support, including:• Worldwide contact telephone numbers• Product support, including:

– Product FAQs– Software, patches, and updates– Training for many applications and instruments

• Order and web support• Product documentation, including:

– User guides, manuals, and protocols– Certificates of Analysis– Safety Data Sheets (SDSs; also known as MSDSs)

Note: For SDSs for reagents and chemicals from other manufacturers,contact the manufacturer.

Limited product warranty

Life Technologies Corporation and/or its affiliate(s) warrant their products as set forthin the Life Technologies' General Terms and Conditions of Sale found on LifeTechnologies' website at www.thermofisher.com/us/en/home/global/terms-and-conditions.html. If you have any questions, please contact LifeTechnologies at www.thermofisher.com/support.


http://thermofisher.com/support

http://www.thermofisher.com/us/en/home/global/terms-and-conditions.html

http://www.thermofisher.com/us/en/home/global/terms-and-conditions.html

http://www.thermofisher.com/support

thermofisher.com/support | thermofisher.com/askaquestion

thermofisher.com

22 February 2017

http://thermofisher.com/support

mailto:thermofisher.com/askaquestion

http://thermofisher.com

ion 520 & ion 530 ext kit −chef user guide (pub. no...

Documents