Thermo Scientific Ion 520 User manual

For Research Use Only. Not for use in diagnostic procedures.
Ion 520™& Ion 530™ExT Kit – Chef
USER GUIDE
for use with:
Ion Chef™System
Ion S5™System
Ion S5™XL System
Ion GeneStudio™S5 System
Ion GeneStudio™S5 Plus System
Ion GeneStudio™S5 Prime System
Catalog Number A30670
Publication Number MAN0015805
Revision F.0

Life Technologies Corporation | 5781 Van Allen Way | Carlsbad, California 92008 USA
For descriptions of symbols on product labels or product documents, go to thermofisher.com/symbols-definition.
The information in this guide is subject to change without notice.
DISCLAIMER: TO THE EXTENT ALLOWED BY LAW, THERMO FISHER SCIENTIFIC INC. AND/OR ITS AFFILIATE(S) WILL NOT BE
LIABLE FOR SPECIAL, INCIDENTAL, INDIRECT, PUNITIVE, MULTIPLE, OR CONSEQUENTIAL DAMAGES IN CONNECTION WITH OR
ARISING FROM THIS DOCUMENT, INCLUDING YOUR USE OF IT.
Revision history: Pub. No. MAN0015805
Revision Date Description
F.0 24 November 2021 •Updated “Start the Ion Chef™run” to update the recommended concentration in
“Dilute the sample library” on page 25.
•Updated “Start the Ion Chef™run” to update the volume of library in “Dilute the
control library” on page 25.
E.0 25 June 2019 •Updated for Torrent Suite™Software 5.12.
•Separated "Prepare the libraries and consumables" into two topics: “Prepare the
consumables” and “Add the diluted libraries to the Ion Chef™Library Sample
Tube”.
•Updated “Start the Ion Chef™run” to include the Run Options screen for
step‑by‑step Ion Chef™run setup.
•Updated the guidance for storing loaded chips before sequencing in “Unload the
chip for sequencing”.
•Added a note in “Start the Ion Chef™run” and “Unload the chip for sequencing”
to NOT remove any residual liquid from chip wells after the Ion Chef™run.
•Removed Ion Chef™System maintenance procedures from “Maintain the
Ion Chef™System”.
•Updated “Maintain the Ion Chef™System” to add references to the Ion Chef™
System maintenance procedures in Ion Chef™Instrument User Guide.
•Updated “Related documentation” to include the Ion Chef™Instrument User
Guide.
D.0 12 December 2018 •Updated for Ion GeneStudio™S5 Series Systems.
•Updated for Torrent Suite™Software 5.10; Planned Run creation topics
reorganized for ease of use.
•Clarification made to dilute barcoded library pools, if used.
•Added a new troubleshooting topic: “Planned Run status does not advance to
"Planned"”.
•Removed reference to quality control of ISPs using the Attune™NxT Acoustic
Focusing Cytometer.
C.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 and
clean") procedure changed from 500 to 200, and recommended time to perform
the procedure after sequencer initialization changed from 27 hours to 24 hours
(see “Perform an instrument reset run with an initialized sequencer that is loaded
with an unused Reagents cartridge” on page 66)
•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 the terms and conditions of all applicable Limited Use Label Licenses.
Trademarks: All trademarks are the property of Thermo Fisher Scientific and its subsidiaries unless otherwise specified. Guava and
easyCyte are trademarks of Guava Technologies, Inc. Agilent and Bioanalyzer are trademarks of Agilent Technologies, Inc.
©2021 Thermo Fisher Scientific Inc. All rights reserved.

Contents
■CHAPTER 1 Product information .................................................. 6
Product description ............................................................. 6
Library compatibility ......................................................... 6
Ion GeneStudio™S5 Series instrument reference ............................... 6
Software compatibility ....................................................... 7
Kit contents and storage ......................................................... 7
Kit summary ............................................................... 7
Kit components ............................................................ 7
Related products ................................................................ 8
Compatible Ion Chip™kits ................................................... 8
Ion S5™ExT Calibration Standard ............................................. 8
Ion S5™Controls Kit Plus .................................................... 9
E. coli DH10B Control 600 Library ............................................. 9
Required materials not supplied ................................................... 9
About the Ion Chef™System ..................................................... 10
Ion Chef™Instrument components ........................................... 10
Interior hardware and consumables .......................................... 11
About the Ion Chef™System touchscreen interface ............................ 12
Workflow ..................................................................... 13
■CHAPTER 2 Before you begin .................................................... 14
Precautions ................................................................... 14
Avoid nucleic acid contamination ............................................ 14
Avoid chip damage ........................................................ 14
Protection by equipment .................................................... 14
Guidelines for using Ion Chef™consumables and reagents .......................... 15
Prepare the Ion Chef™Instrument for use ......................................... 15
■CHAPTER 3 Create a Planned Run .............................................. 17
About Planned Runs ........................................................... 17
Create a Planned Run .......................................................... 18
Planned Run workflow key fields ................................................. 22
Ion 520™& Ion 530™ExT Kit − Chef User Guide 3

■CHAPTER 4 Start the Ion Chef™run ............................................. 24
Materials required .............................................................. 24
Dilute the library ............................................................... 24
Guidelines for library dilution ................................................ 25
Dilute the sample library .................................................... 25
Dilute the control library .................................................... 25
Prepare the consumables ....................................................... 25
Add the diluted libraries to the Ion Chef™Library Sample Tube ....................... 26
Load the Ion Chef™System ..................................................... 27
Load the tip racks and PCR Plate ............................................ 28
Load the Reagents and Solutions cartridges .................................. 29
Load the Recovery Tubes and Enrichment Cartridge v2 ......................... 31
Load the Chip-loading centrifuge ............................................ 33
Confirm that consumables are correctly installed .............................. 35
Ion Chef™pre-run checklist ................................................. 36
Start the Ion Chef™run ......................................................... 37
Unload the chip for sequencing .................................................. 42
■CHAPTER 5 Initialize the sequencer ............................................ 45
Ion S5™/Ion GeneStudio™S5 Systems component positions ........................ 45
Before you begin .............................................................. 46
When a manual cleaning of the sequencer is required .............................. 46
Initialize the sequencer ......................................................... 47
■CHAPTER 6 Start the sequencing run ........................................... 49
Chip handling guidelines ........................................................ 49
Start the sequencing run ........................................................ 49
Maintain the sequencer ......................................................... 52
Materials required .......................................................... 52
Clean or decontaminate the sequencer ....................................... 52
Reagent consumables disposal .................................................. 52
CO2scrubber removal and disposal .......................................... 52
Recycle Ion S5™ExT Wash Solution and Ion S5™Cleaning Solution bottles ....... 53
■CHAPTER 7 Clean the Ion Chef™System ...................................... 54
About the cleaning protocol ..................................................... 54
Materials required .............................................................. 54
Clean the Ion Chef™Instrument .................................................. 54
Remove and dispose of used consumables ................................... 55
Inspect and clean the Recovery centrifuges and buckets ....................... 57
Start the cleaning ......................................................... 58
Contents
4Ion 520™& Ion 530™ExT Kit − Chef User Guide

■APPENDIX A Troubleshooting .................................................... 60
Ion Chef™System setup and operation ........................................... 60
Ion S5™/Ion GeneStudio™S5 Systems setup and operation ......................... 63
Instrument alarms and events ............................................... 63
Initialization—General errors ................................................ 64
■APPENDIX B Supplemental procedures ........................................ 65
Perform a manual cleaning of the sequencer ...................................... 65
Perform an instrument reset run with an initialized sequencer that is loaded with
an unused Reagents cartridge ................................................. 66
Quality control of Ion Sphere™Particles .......................................... 67
Quality control using the Guava™easyCyte 5 Benchtop Flow Cytometer .......... 67
Maintain the Ion Chef™System .................................................. 67
■APPENDIX C Safety ............................................................... 68
Symbols on this instrument ..................................................... 68
Conformity symbols ........................................................ 69
Location of safety labels on this instrument ........................................ 70
Safety information for instruments not manufactured by Thermo Fisher Scientific ...... 70
Instrument safety .............................................................. 70
General ................................................................... 70
Physical injury ............................................................. 70
Electrical safety ........................................................... 71
Cleaning and decontamination .............................................. 71
Ultraviolet (UV) Safety ...................................................... 71
Safety and electromagnetic compatibility (EMC) standards .......................... 71
Safety compliance ......................................................... 72
EMC ..................................................................... 72
Environmental design ...................................................... 72
Instrument safety — Ion S5™/Ion GeneStudio™S5 Systems ......................... 73
Chemical safety ................................................................ 73
Biological hazard safety ......................................................... 74
■Documentation and support ....................................................... 75
Related documentation ......................................................... 75
Customer and technical support ................................................. 76
Limited product warranty ........................................................ 76
Contents
Ion 520™& Ion 530™ExT Kit − Chef User Guide 5

Product information
■Product description .................................................................... 6
■Kit contents and storage ............................................................... 7
■Related products ...................................................................... 8
■Required materials not supplied ......................................................... 9
■About the Ion Chef™System ........................................................... 10
■Workflow ............................................................................ 13
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) contains all the reagents and materials that
are required to perform the following steps in the sequencing workflow.
•Prepare enriched, template-positive Ion Sphere™Particles (ISPs) and load them onto an Ion 520™
Chip or Ion 530™Chip using the Ion Chef™Instrument.
•Sequence the loaded chips on an Ion S5™Sequencer, Ion S5™XL Sequencer, or Ion GeneStudio™
S5 Series Sequencer.
The Ion Chef™Instrument uses automated isothermal amplification technology with the Ion 520™&
Ion 530™ExT Kit – Chef to prepare 400 to 600‑base‑read templates with greatly reduced run times
compared to emulsion PCR-based templating. Each kit contains all the supplies that are required
to load and sequence 4 chips. The kit supports 4 single-chip Ion Chef™runs and 4 sequencer
initializations (1 sequencing run/sequencer initialization).
Library compatibility
The Ion 520™& Ion 530™ExT Kit – Chef can be used with up to 400 to 600-base-read libraries of any
type prepared using any available Ion Torrent™-branded library kit.
Ion GeneStudio™S5 Series instrument reference
In this document, Ion GeneStudio™S5 Series Sequencer or Ion GeneStudio™S5 Series System refers
generically to the following systems, unless otherwise specified.
•Ion GeneStudio™S5 System (Cat. No. A38194)
•Ion GeneStudio™S5 Plus System (Cat. No. A38195)
•Ion GeneStudio™S5 Prime System (Cat. No. A38196)
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6Ion 520™& Ion 530™ExT Kit − Chef User Guide

Software compatibility
The Ion 520™& Ion 530™ExT Kit – Chef is compatible with Torrent Suite™Software v5.2.1 and later. Be
sure to update your Ion Torrent™Server to the latest available version of Torrent Suite™Software before
using this kit.
Kit contents and storage
IMPORTANT!
·Do not substitute components from any other Ion sequencing kits. We verified this protocol using
these specific materials. Substitution can adversely aect system performance.
·Store all consumables and cartridges under the recommended conditions and in an upright position.
Do NOT store the Ion S5™ExT Sequencing Reagents (Part No. A30522) on dry ice or in a closed
environment where dry ice is present.
Each Ion 520™& Ion 530™ExT Kit –
Chef contains all the supplies that are required to prepare and
sequence 4 Ion 520™Chips or Ion 530™Chips. Cat. No. A30670 supports 4 single-chip Ion Chef™
runs and 4 sequencer initializations (1 sequencing run/sequencer initialization) for 400 to 600‑base‑read
libraries.
On arrival, inspect all consumables and contact Technical Support if any of the products have been
damaged during shipping.
Kit summary
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
Kit components
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
Chapter 1 Product information
Kit contents and storage 1
Ion 520™& Ion 530™ExT Kit − Chef User Guide 7

Contents Amount / box Storage
Recovery Station Disposable Lid v2 15°C to 30°C2
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
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.
Related products
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 chips 15°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 Ion S5™ExT
Calibration Standard can be added to libraries to increase base-calling accuracy. The Ion S5™ExT
Calibration Standard (Cat. No. A30738) is ordered separately.
Contents Amount Storage
Ion S5™ExT Calibration Standard 16 µL –30°C to –10°C
Chapter 1 Product information
Related products
1
8Ion 520™& Ion 530™ExT Kit − Chef User Guide

Ion S5™Controls Kit Plus
The Ion S5™Controls Kit Plus (Cat. No. A30729) is ordered separately. See the Ion S5™Controls Kit Plus
Product Information Sheet (Pub. No. MAN0016206) for details.
Contents Amount Storage
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
E. coli DH10B Control 600 Library
The E. coli DH10B Control 600 Library includes sucient material for 4 templating runs on the
Ion Chef™System. The E. coli DH10B Control 600 Library (Cat. No. A32470) is ordered separately.
Contents Amount Storage
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" indicates that
the material is available from fisherscientific.com or another major laboratory supplier.
Item Source
Ion Chef™System 4484177
Microcentrifuge[1] MLS
2‑, 20‑, 200‑, and 1,000‑µL pipettes and appropriate filtered tips 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
(Optional) Uninterruptible Power Supply (UPS) [2] MLS
[1] 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.
[2] For laboratories that experience frequent power outages or line voltage fluctuations, we recommend that you use an uninterruptible
power supply that is compatible with 2500 W output or higher.
Chapter 1 Product information
Required materials not supplied 1
Ion 520™& Ion 530™ExT Kit − Chef User Guide 9

About the Ion Chef™System
The Ion Chef™System (Cat. No. 4484177) provides automated, high-throughput template preparation
and chip loading for use with an Ion S5™Sequencer, Ion S5™XL Sequencer, or Ion GeneStudio™S5
Series Sequencer. The system uses a set of cartridge-based consumables and reagents that enables
a user to load an Ion 520™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 Torrent Suite™Software to
enable sample and reagent traceability throughout the chip preparation workflow.
Ion Chef™Instrument components
The following figure illustrates the major external and internal features of the Ion Chef™Instrument.
4
1 2 3
5 6 7
8
1Door – Provides access to the interior of the
instrument. The door is locked in the closed position
during operation.
2Micropipettor – A mechanical positive-displacement
pipettor that performs all fluid transfers during sample
and chip preparation.
3Robotic arm – Enables fluid transfer by the
Micropipettor. The arm also contains an optical sensor
that reads the barcodes of instrument reagents and
consumables.
4Touchscreen – Provides access to all instrument
functions for operation, maintenance, and
troubleshooting.
5Power button – Power switch for the Ion Chef™
Instrument, where the states are on (illuminated) and
o.
6Power port – A 100–240 VAC port that provides power
to the Ion Chef™Instrument.
7Ethernet port – An RJ45 port that provides Ethernet
(100 Mbit) communication with the Ion Chef™
Instrument.
8USB port – Provides USB communication with the
Ion Chef™Instrument. Used to update the instrument
firmware and to transfer data during service or
maintenance.
Chapter 1 Product information
About the Ion Chef™System
1
10 Ion 520™& Ion 530™ExT Kit − Chef User Guide