Illumina Infinium HD Super Assay Bedienungsanleitung

Infinium HD Super Assay

Reference Guide

Document # 11322427 v03

August 2019

ILLUMINA PROPRIETARY

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

This document and its contents are proprietary to Illumina, Inc. and its affiliates ("Illumina"), and are intended solely for

the contractual use of its customer in connection with the use of the product(s) described herein and for no other

purpose. This document and its contents shall not be used or distributed for any other purpose and/or otherwise

communicated, disclosed, or reproduced in any way whatsoever without the prior written consent of Illumina. Illumina

does not convey any license under its patent, trademark, copyright, or common-law rights nor similar rights of any third

parties by this document.

The instructions in this document must be strictly and explicitly followed by qualified and properly trained personnel in

order to ensure the proper and safe use of the product(s) described herein. All of the contents of this document must be

fully read and understood prior to using such product(s).

FAILURE TO COMPLETELY READ AND EXPLICITLY FOLLOW ALL OF THE INSTRUCTIONS CONTAINED HEREIN MAY

RESULT IN DAMAGE TO THE PRODUCT(S), INJURY TO PERSONS, INCLUDING TO USERS OR OTHERS, AND DAMAGE

TO OTHER PROPERTY, AND WILL VOID ANY WARRANTY APPLICABLE TO THE PRODUCT(S).

ILLUMINA DOES NOT ASSUME ANY LIABILITY ARISING OUT OF THE IMPROPER USE OF THE PRODUCT(S)

DESCRIBED HEREIN (INCLUDING PARTS THEREOF OR SOFTWARE).

All trademarks are the property of Illumina, Inc. or their respective owners. For specific trademark information, see

www.illumina.com/company/legal.html.

Document # 11322427 v03

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

iii

Infinium HD Super Assay Reference Guide

Revision History

Document Date Description of Change

Document # 11322427 v03 August 2019 Removed all references to WG# in barcode

information in regards to DNA plates and MSA

plates.

Added Barcode Numbers section to Tips and

Techniques to explain the new barcode

numbering scheme.

Document # 11322427 v02 June 2019 Removed instructions for 4x1 and 4x3

BeadChips.

Changed document name to

Infinium HD Super

Assay Reference Guide

Document updated to reflect latest style and

formatting standards.

Added Preparation and Storage of User-Supplied

Reagent section to consolidate reagent

preparation and use instructions.

Corrected storage temperatures for MA1, PB1,

RA1, XC3, and XC4.

Corrected reagents listed in first and last bullets

of About Reagents section in the Extend and

Stain introduction.

Reference to Experienced User Cards replaced

with

Infinium HD Super Assay Checklist Manual

Protocol (document # 1000000080218)

and

Infinium HD Super Assay Checklist Automated

Protocol (document # 1000000080220)

which

provide current format of experienced user

instructions.

Added list of acronyms.

Consumables and equipment information moved

Infinium Assay Lab Setup and Procedures

Guide (document # 11322460)

Added Tip and Techniques section, includes:

• Tips, techniques, and general reagent storage

and preparation information previously

documented in protocol sections.

• Directions to seal plates before vortex and

centrifuge steps.

• BeadChip handling.

• Hybridization Chamber handling.

Document # 11322427 v01 November

2015

Corrected the hybridization oven temperature to

37°C when incubating the MSA1 plate and the

DNA.

Updated the total volume of PB1 required in

relation to the number of alignment fixtures in

use.

Document # 11322427 v03

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

Infinium HD Super Assay Reference Guide

Document Date Description of Change

Part # 11294809 Rev. D May 2012 Added that up to 24 BeadChips can be washed

with 550 ml of PB1.

Part # 11294809 Rev. C June 2010 Update.

Part # 11294809 Rev. B June 2009 Update.

Part # 11294809 Rev. A November

2008

Minor revisions.

Part # 11294809 Rev. A May 2008 Initial release.

Document # 11322427 v03

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

Infinium HD Super Assay Reference Guide

Table of Contents

Chapter 1 Overview 1

Introduction to Infinium HD Super Assay 1

Important Note 2

Additional Resources 2

DMAPFiles 2

Preparation and Storage of User-Supplied Reagents 2

Tips and Techniques 3

Acronyms 5

Illumina LIMS 5

No Illumina LIMS 6

Tracking Tools 6

Chapter 2 Manual Protocol 7

Introduction 7

Infinium HD Super Manual Workflow 7

Amplify DNA 9

Incubate DNA 10

Fragment DNA 10

Precipitate DNA 11

Resuspend DNA 13

Hybridize to BeadChip 14

Wash BeadChips 19

Extend and Stain BeadChips 25

Image BeadChip 34

Illumina GenomeStudio 35

Chapter 3 Automated Protocol 37

Introduction 37

Infinium HD Super Automated Workflow 37

Amplify DNA 39

Incubate DNA 41

Fragment DNA 41

Precipitate DNA 42

Resuspend DNA 44

Hybridize to BeadChip 46

Wash BeadChips 53

Extend and Stain BeadChips 58

Image BeadChip 68

Illumina GenomeStudio 69

Technical Assistance 70

Chapter 1 Overview

Overview

Introduction to Infinium HD Super Assay 1

Important Note 2

Additional Resources 2

DMAPFiles 2

Preparation and Storage of User-Supplied Reagents 2

Tips and Techniques 3

Acronyms 5

Illumina LIMS 5

No Illumina LIMS 6

Tracking Tools 6

Introduction to Infinium HD Super Assay

The Illumina Infinium HD Super Assay is an evolution for DNA analysis, by streamlining sample preparation and

enabling high multiplexing. Using Infinium I and Infinium II probe designs and a dual-color channel approach,

the Infinium HD Super Assay enables the DNA analysis of up to several million of SNPs and CNV markers per

sample.

The Infinium HD Super Assay accomplishes this unlimited multiplexing by combining whole-genome

amplification (WGA) sample preparation with direct, array-based capture, and enzymatic scoring of the SNP

loci. Locus discrimination or copy number variation (CNV) determination comes from a combination of high

bead type representation per feature, sequence-specific hybridization capture and array-based, single-base

primer extension. In the case of the Infinium II probe design, the 3' end of the primer is positioned directly

adjacent to the SNP site, or if a nonpolymorphic probe, directly adjacent to the nonpolymorphic site. With the

Infinium I probe design, the 3' end of the primer overlaps with the SNP site. If there is a perfect match,

extension occurs and signal is generated. If there is a mismatch, extension does not occur and no signal is

generated.

Allele-specific single base extension of the primer incorporates a biotin nucleotide or a dinitrophenyl labeled

nucleotide. C and G nucleotides are biotin labeled; A and T nucleotides are dinitrophenyl labeled. Signal

amplification of the incorporated label further improves the overall signal-to-noise ratio of the assay.

The Infinium HD Super Assay offers:

uHigh multiplexing

uHigh call rate and accuracy

uUnlimited genomewide marker selection

uSingle tube amplification, single chip, no PCR

uMinimal risk of carryover contamination

uLow DNA input—200 ng per sample

uWalk-away automation using Tecan Genesis or Freedom EVO Robots and Tecan GenePaint system

uCompatibility with Illumina iScan and HiScanSystem

uMultiple-sample BeadChip format

Document # 11322427 v03

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

Important Note

Before using the procedures in this guide, read the

Infinium Lab Setup and Procedures Guide

. The

Setup

and Procedures

guide explains how to equip and run an Infinium HD Super Assay laboratory, including

information on the following topics:

uPrevention of amplification product contamination

uSafety precautions

uConsumables and equipment to purchase in advance

uStandard lab procedures

uRobot use for automation

uPreparation for BeadChip imaging

uSystem Controls

uSystem maintenance

uTroubleshooting

The instructions apply equally to all Infinium BeadChips provided by Illumina™. All Infinium HD Super

documentation assumes that you have already set up the laboratory space and that you are familiar with the

standard procedures and safety precautions.

Additional Resources

Visit the Infinium HD Super Assay support page on the Illumina website for documentation, software

downloads, training resources, and information about compatible Illumina products.

The following documentation is available for download from the Illumina website.

Resource Description

Infinium HD Super Assay Checklist Automated Protocol

(document # 1000000080220)

Provides a checklist for experienced users of the Infinium

HD Super Assay when performing the automated protocol.

Infinium HD Super Assay Checklist Manual Protocol

(document # 1000000080218)

Provides a checklist for experienced users of the Infinium

HD Super Assay when performing the manual protocol.

Infinium Lab Setup and Procedures Guide (document

#11322460)

Describes how to set up an Infinium HD Super lab

including reagents, consumables, and equipment to

purchase in advance, and best practices for lab operation.

Infinium Consumables and Equipment List (document #

1000000084294)

Provides an interactive checklist of Illumina-provided and

user-provided consumables and equipment.

DMAPFiles

Before run setup, download the DMAP files for the arrays, and prepare a sample sheet. Use the Decode File

Client to download the DMAP files.

Preparation and Storage of User-Supplied Reagents

uMaintain a first in, first out (FIFO) system for reagents. Rotate the stock of the remaining reagents to avoid

using expired reagents.

uInfinium HD Super kits contain reagents in exact quantities needed for the assay. Measure reagents

carefully to avoid shortages.

Document # 11322427 v03

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

Infinium HD Super Assay Reference Guide

uUse fresh reagents for each batch of plates, and empty reservoirs between batches.

uTo minimize errors when preparing user-supplied reagents, prepare large batches of 0.1 N NaOH and

95% Formamide/1 mM EDTA using the following guidance.

Preparing Batches of 0.1 N NaOH

Prepare fresh 0.1 N NaOH in large batches. Divide batches into 15 ml or 50ml sealed tubes.

Store the sealed tubes for 6 months at 2°C to 8°C and use the stored 0.1 NNaOHas needed. Use the 0.1

NNaOHthe same day you open the tube, and discard any unused amounts.

Preparing Batches of 95% Formamide/1 mM EDTA

Prepare the 95%formamide/1mM EDTA mixture in large batches. Divide batches into 15ml or 50 ml sealed

tubes.

Store the sealed tubes for up to 6 months at -25°C to -15°C and use the stored mixture as needed. Use the

mixture the same day you open the tube, and discard any unused amounts.

Use Fresh RA1 Reagent for Each Step

Use fresh RA1 for each step in the assay where it is required. RA1 that has been stored properly and has not

been dispensed for use in either the XStain or Resuspension step is considered fresh RA1. After RA1 has

been exposed to room temperature air for extended periods of time, it is no longer fresh.

To make best use of RA1, only pour the amount needed for the current step. If you plan to perform additional

assay steps requiring RA1 the same day, leave the remaining thawed reagent in the original, closed bottle at

room temperature until it is needed. Follow the standard RA1 storage procedures described in this guide for

next-day processing and prolonged storage conditions.

Store PB1

For Infinium HD Super workflows do the following.

uStore PB1 at room temperature.

uKeep PB1 for up to 3 months.

Tips and Techniques

Unless a safe stopping point is specified, proceed immediately to the next step.

Avoid Cross-Contamination

When adding or transferring samples, change tips between

each sample

Measure Volumes Carefully

When measuring volumes of reagent to put in reservoirs, make sure that the measurements are exactly as

specified in the instructions. Do not estimate volumes.

HD Glass Back Plates

uFor optimal performance, use HD glass back plates that are free of chips and cracks along the beveled

edge and the long edges. Chipped or cracked HD glass back plates risk volume leakage and affect

overall performance.

Document # 11322427 v03

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

Infinium HD Super Assay Reference Guide

uClean HD glass back plates according to the procedures in the

Infinium Assay Lab Setup and Procedures

Guide (Document # 11322460)

uA 10% bleach soak is required weekly, or every seven uses, to maintain glass performance.

Barcode Numbers

uBarcode serial numbers used for workflow enforcement and positive sample tracking on MSA plates,

DNA plates, and reagents use a numbering pattern of two alphanumeric characters followed by seven

random numbers.

Sealing the Plate

uAlways seal plates before the vortex and centrifuge steps in the protocol.

uOrient sealing mats so that the A1 on a cap matches the A1 on the plate.

uMake sure that all 96 caps are securely seated in the wells to prevent evaporation and spills, which

introduce variability and cross-contamination.

uRemove sealing mats slowly and carefully to prevent splashing, and then set aside upside-down in a

safe location.

uWhen returning a sealing mat to a plate, make sure that the orientation is correct.

Heat Sealer

uIf you are using the ALPS 50 V model heat sealer, set it to 165°C and 2.5 seconds.

Pipetting

uMake sure that pipettes are properly calibrated, cleaned, and decontaminated.

uDispense slowly and carefully to prevent turbulence in the plate wells and flow-through chambers.

uUse a multichannel pipette whenever possible.

Centrifugation

uWhen centrifuging plates or BeadChips, place a balance plate or rack with BeadChips opposite each

plate or rack being centrifuged. Make sure that the weights are as similar as possible.

Washing and Coating BeadChips

Perform the following steps before starting the wash and coat process:

uPlace wash dish covers on wash dishes when not in use.

uClean wash dishes with low-pressure air to remove particulates before use.

uWash tube racks and wash dishes thoroughly before and after use. Rinse with DI H2O. Place them

upside down on wash rack to dry.

uPrepare an additional clean tube rack that fits the internal dimensions of the vacuum desiccator. Allow

one rack per eight BeadChips.

Handling Hybridization Chambers

uKeep the chamber lids and bases together. Adopt a labeling convention that pairs each chamber base

with the original lid.

Document # 11322427 v03

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

Infinium HD Super Assay Reference Guide

Inhaltsverzeichnis

Andere illumina Laborausrüstung Handbücher

illumina

illumina MiSeq System Bedienungsanleitung

illumina

illumina NextSeq 550Dx Bedienungsanleitung

illumina

illumina NextSeq 550Dx Bedienungsanleitung

illumina

illumina NextSeq 500 Stücklistenhandbuch

illumina

illumina MiniSeq Stücklistenhandbuch

illumina

illumina MiSeq System Stücklistenhandbuch

illumina

illumina MiSeqDx Instrument Installations- und Betriebsanleitung

illumina

illumina NeoPrep Library Prep System Installations- und Betriebsanleitung

illumina

illumina NovaSeq 6000 Bedienungsanleitung

illumina

illumina cBot 2 Stücklistenhandbuch

illumina

illumina HiSeq X Bedienungsanleitung

illumina

illumina Infinium LiHa Bedienungsanleitung

illumina

illumina MiSeqDx Bedienungsanleitung

Beliebte Laborausrüstung Handbücher anderer Marken

Qiagen

Qiagen DML 3000 Bedienungsanleitung

Queensgate

Queensgate NANOSCAN OP400 Programmierhandbuch

Conviron

Conviron GEN1000-GE Bedienungsanleitung

Parr Instrument

Parr Instrument 4560 Bedienungsanleitung

IKA

IKA IC control Bedienungsanleitung

Integra

Integra DOSE IT Bedienungsanleitung

Agilent Technologies

Agilent Technologies 5800 ICP-OES Bedienungsanleitung

Endress+Hauser

Endress+Hauser Cleanfit CPA875 Bedienungsanleitung

NI PXI-5422 Handbuch

Collomix

Collomix Aqix Bedienungsanleitung

SPEX SamplePrep

SPEX SamplePrep 6875 Freezer/Mill Series Bedienungsanleitung

Ocean Insight

Ocean Insight FLAME-NIR+ Bedienungsanleitung

Parker

Parker ALIGN-MG-NA Bedienungsanleitung

BD 644787 Bedienungsanleitung

DENTAURUM

DENTAURUM Compact Megaplus Bedienungsanleitung

Biuged Laboratory Instruments

Biuged Laboratory Instruments BGD 626 Bedienungsanleitung

VWR

VWR SAS Super IAQ Bedienungsanleitung

Yamato

Yamato BM510 Bedienungsanleitung

illumina Infinium HD Super Assay Bedienungsanleitung

Beliebte Laborausrüstung Handbücher anderer Marken