illumina MiSeqDx User manual
FOR IN VITRO DIAGNOSTIC USE
ILLUMINA PROPRIETARY
Document # 15038353 v04
March 2018
MiSeqDx®Instrument
Reference Guide
ii Document # 15038353 v04
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Revision History
Document# Date Description of Change
Document # 15038353
v04
March
2018
Added information about the Illumina Proactive monitoring
service in the Run Options Screen section.
Document # 15038353
v03
September
2017
Updated regulatory markings.
Document # 15038353
v02
March
2017
Corrected the Intended Use Statement. Removed, "The
MiSeqDx is intended to be used with Illumina supplied
IVDassays and reagent kits."
Document # 15038353
v01
December
2016
Changed the name of the guide from the MiSeqDx Reference
Guide to the MiSeqDx Instrument Reference Guide.
Added a section on the Illumina Worklist Manager application.
Changed the formamide warning to a general warning for
potentially hazardous chemicals, with a recommendation to
wear protective equipment.
Changed the type of water required for washing the
instrument from DNase-free, Rnase-free water to laboratory-
grade water. Listed acceptable examples of laboratory-grade
water, including Illumina PW1.
Removed the Measuring Expected Wash Volumes section.
Changed the Welcome screen to the Home screen.
Changed the Intended Use Statement to make it the same as
the statement in the MiSeqDx Instrument Package Insert
(document # 15050260).
Added storage and handling information.
Marking and formatting changes.
Part # 15038353 Rev. B February
2015
Updated the formamide warning with a web address for
access to safety data sheet (SDS) information.
Added inspecting reagent cartridge positions 2 and 4 to the
instructions in the Inspect the Reagent Cartridge section
because the cartridge positions are the same size as position 1.
Updated the Resolving RFID Read Failure directions to include
obtaining a temporary RFID bypass code from Illumina
Technical Support.
Marking and formatting changes.
Part # 15038353 Rev. A February
2014
Initial release.
MiSeqDx Instrument Reference Guide iii
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Table of Contents
Revision History iii
Table of Contents v
Chapter 1 Getting Started 1
Intended Use 2
Components 3
Start the MiSeqDx 6
Required Consumables 7
Storage and Handling 7
Anti-Virus Software 8
Chapter 2 MiSeqDx Software 9
Introduction 10
Illumina User Management Software 11
Illumina Worklist Manager Software 15
MiSeq Operating Software (MOS) 16
Real-Time Analysis (RTA) 34
MiSeq Reporter 35
Chapter 3 Performing a Run 37
Introduction 38
MiSeqDx Workflow 39
Prepare the Reagent Cartridge 40
Log In and Select Run Type 42
Clean the Flow Cell 43
Load the Flow Cell 45
Loading Reagents 47
Starting the Run 49
Monitoring the Run 50
Chapter 4 Maintenance Procedures 51
Introduction 52
Perform a Post-Run Wash 53
Perform a Maintenance Wash 55
Perform a Standby Wash 58
Shut Down the Instrument 60
Chapter 5 Troubleshooting 61
Introduction 62
Resolve Run Setup Errors 63
Resolve RFID Read Failure 64
Prevent Reboots During a Run 66
Troubleshoot Flow Rate Error 67
Perform a Volume Test 68
Resolve MiSeq Reporter Analysis Errors 70
Index 71
Technical Assistance 73
MiSeqDx Instrument Reference Guide v
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Getting Started
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Intended Use
The Illumina MiSeqDx is a sequencing instrument that measures fluorescence signals of
labeled nucleotides through the use of instrument specific reagents and flow cells (MiSeqDx
Universal Kit 1.0), imaging hardware, and data analysis software. The MiSeqDx Platform
is intended for targeted sequencing of human genomic DNA from peripheral whole blood
samples. The MiSeqDx Platform is not intended for whole genome or de novo sequencing.
Components
MiSeqDx Instrument Reference Guide 3
Components
The MiSeqDx has the following exterior components:
AFlow cell compartment—Contains the flow cell stage that houses the flow cell
throughout the run. Flow cell stage motors move the stage out of the enclosed optical
module for flow cell loading and returns the stage when the run begins.
BEnclosed optics module—Contains optical components that enable imaging of the flow
cell.
CStatus bar—Uses 3 colors to indicate instrument status. Blue indicates that the
instrument is processing, orange indicates the instrument needs attention, and green
indicates that the instrument is ready to begin the next run.
DTouch screen monitor—Enables on-instrument configuration and run setup using the
software interface.
EExternal USB port—Facilitates the transfer of files and data to the instrument computer
from the touch screen monitor.
FReagent compartment—Holds reagents at proper temperatures, wash solutions, and
the waste bottle. A magnetic latch secures the reagent compartment door.
The MiSeqDx interface guides users through the run setup steps using the touch screen
monitor.
Getting Started
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Flow Cell Compartment
AFlow Cell Stage
BFlow Cell Compartment Door
CFlow Cell Latch
DFlow Cell
EFlow Cell Latch Release Button
The flow cell compartment houses the flow cell stage, thermal station, and fluidics
connections to the flow cell. The flow cell stage holds the flow cell and the flow cell latch
secures and positions the flow cell. When the flow cell latch closes, 2 pins near the latch
hinge auto-position the flow cell.
The thermal station, located beneath the flow cell stage, controls changes in flow cell
temperature required for cluster generation and sequencing.
Flow Cell
AOutlet Port
BImaging Area
CInlet Port
The MiSeqDx flow cell is a single-use glass-based substrate on which clusters are generated
and the sequencing reaction is performed.
Reagents enter the flow cell through the inlet port, pass through the single-lane imaging
area, and then exit the flow cell through the outlet port. Waste exiting the flow cell is
delivered to the waste bottle.
During the sequencing run, the single lane is imaged in small imaging areas called tiles.
Components
MiSeqDx Instrument Reference Guide 5
Reagent Compartment
AReagent Chiller
BSipper Handle (shown in raised position)
CMiSeqDx SBS Solution (PR2) Bottle
DWaste Bottle
EReagent Cartridge
The reagent compartment contains the reagent chiller, and positions for the MiSeqDx SBS
Solution (PR2) bottle and the waste bottle.
During the run, the reagent chiller holds a single-use reagent cartridge. During the
instrument wash, the reagent chiller holds the wash tray. The software automatically
lowers sippers into each reservoir of the reagent cartridge at the appropriate time during a
run depending on the process being performed.
To the right of the reagent chiller are 2 form-fitted slots, 1 for the MiSeqDx SBS Solution
(PR2) bottle and 1 for the waste bottle. The sipper handle locks the bottles in place and
lowers the appropriate sipper into each bottle.
Reagents are pumped through the sippers and fluidics lines, and then to the flow cell.
Reagent waste is delivered to the waste bottle throughout the process.
Getting Started
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Start the MiSeqDx
NOTE
Illumina recommends that you leave the instrument on continuously. However, if the
instrument must be turned off follow the shutdown procedure described in Shut Down the
Instrument on page 60. Wait a minimum of 60seconds before turning the power switch back
to the ON position.
1If the MiSeqDx is not already on, reach around the right side of the instrument to locate
the power switch on the back panel. It is in the lower corner directly above the power
cord.
Figure 1 Power Switch Location
2Turn the power switch to the ON position. The integrated instrument computer starts.
3Log in to the operating system.
Wait until the operating system has finished loading. The MiSeq Operating Software
(MOS) launches and initializes the instrument automatically.
After the initialization step is complete, the Home screen appears.
Figure 2 Home Screen
Required Consumables
MiSeqDx Instrument Reference Guide 7
Required Consumables
Sequencing Consumables
The sequencing consumables required to run the MiSeqDx are provided separately as part
of an in vitro diagnostic kit.
User-Supplied Consumables
Make sure that the following user-supplied consumables are available before beginning a
run.
Consumable Purpose
Alcohol wipes, 70% Isopropyl
or
Ethanol, 70%
Cleaning the flow cell holder
Lab tissue, low-lint Cleaning the flow cell stage
Lens paper, 4x6in. Cleaning the flow cell
Tween 20 Washing the instrument
Tweezers, square-tip plastic (optional) Removing flow cell from flow cell shipping
container
Water, laboratory-grade Washing the instrument
Guidelines for Laboratory-Grade Water
Always use laboratory-grade water to perform instrument procedures. Never use tap water.
Any of the following are acceptable examples:
}Illumina PW1
}18Megaohm (MΩ) water
}Milli-Q water
}Super-Q water
}Molecular biology-grade water
Storage and Handling
Element Specification
Temperature Transportation and Storage: -10°C to 40°C (14°F to 104°F)
Operating Conditions: 19°C to 25°C (66°F to 77°F)
Humidity Transportation and Storage: Non-condensing humidity
Operating Conditions: 30–75% relative humidity (non-condensing)
Getting Started
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Anti-Virus Software
Illumina strongly recommends installation of user-supplied anti-virus software to protect
the computer against viruses.
To avoid interfering with MiSeqDx operation or losing data, configure the anti-virus
software updates as follows:
}Set for manual scans, not automatic scans.
}Perform scans only when the instrument is not in use.
}Set updates to download but not install without user authorization.
}Do not automatically reboot the computer upon update.
}Exclude the data drive and application directory from any real-time file system
protection.
MiSeqDx Software
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Introduction
The software described in this chapter is used to configure, run, and analyze data from the
MiSeqDx.
Figure 3 Interaction between MiSeqDx Software Components
}Illumina Worklist Manager (IWM)—Is used to create a sample sheet during library
preparation. The sample sheet ensures positive sample tracking throughout the
process. This software runs off-instrument and the sample sheet is associated with the
run during run setup on the MiSeqDx. See the kit-specific reference guide for
information on generating a sample sheet. See Illumina Worklist Manager Software on
page 15.
}Illumina User Management Software—Is used to grant various access level
permissions to users, as appropriate. This software runs off-instrument and the
permissions are saved in a database file, which the MiSeqDx is configured to reference.
See Illumina User Management Software on page 11.
}MiSeq Operating Software (MOS)—Controls the instrument operation, including
various configuration settings. This software is installed and runs on the instrument.
See MiSeq Operating Software (MOS) on page 16.
}Real Time Analysis (RTA)—Performs image processing and base calling (hereafter
referred to as primary analysis). This software is installed and runs on the instrument.
See Real-Time Analysis (RTA) on page 34.
}MiSeq Reporter—Performs secondary analysis such as demultiplexing, alignment,
variant calling, and report generation. The specific functions that are supported vary by
kit. This software can be installed on- or off-instrument. The output is viewed through
a web browser. See workflow-specific information in the MiSeq Reporter Software
Reference Guide (document#15038356) for analyzing data.
Illumina User Management Software
MiSeqDx Instrument Reference Guide 11
Illumina User Management Software
User names, passwords, and access levels are managed using the Illumina User
Management Software, which creates a database file that enables login permissions for
authorized users. The User Management Software is not installed on the instrument
computer and is intended for use off-instrument.
A user database file is required prior to operating the MiSeq Operating Software (MOS). For
information on linking the MiSeqDx and the user database file, see Run Options Screen on
page 20.
Starting the User Management Software
The following instructions describe how to download, install, and start the Illumina User
Management Software, and how to create a user database file.
}Downloading the Software—The Illumina User Management Software can be
downloaded from the Illumina website at http://www.illumina.com. Go to the MiSeqDx
support page and click Downloads. A MyIllumina account is required.
}Installing the Software—The Illumina User Management Software can be run on any
Windows platform. To start the installation, double-click the Setup.exe file. The
installation wizard opens.
}Starting the Software—To start the software, go to Start | All Programs | Illumina |
Illumina User Management Software, or click the desktop icon.
The Illumina User Management Software is used off-instrument. Upon opening the
application for the first time, create an Admin account and assign a password.
Figure 4 Create Admin Account
Only users assigned to an Admin access level can add new users, change user types,
change user passwords, or create a new database.
After logging in, the Illumina User Management Software interface opens.
Using the Illumina User Management Software
Use the Illumina User Management Software to open an existing database file, create a new
database file, add or remove users, and assign passwords and user types for authorized
users.
}Open an existing database file—Select Browse and browse to the location where the
database file is stored. The database file location is defined on the Folders Settings tab
of the Run Options screen. For more information, see Run Options Screen on page 20.
When the database file opens, the user names contained in the file appear in the
Database Contents and Settings section.
MiSeqDx Software
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}Create a new database file—Select Create New and assign a name to the database
file. The file format defaults to *.db.
Figure 5 Illumina User Management Software Interface
Changes made to the database file are saved automatically with the exception of changes to
attributes in the User Information section. After making changes to user information, select
Save before closing the Illumina User Management Software.
To associate the database file with the MiSeqDx, go to the Run Options screen, Folder
Settings tab in the MiSeq Operating Software (MOS).
Add or Remove Users
Each authorized user has four attributes that are assigned in the database file: password,
user type, duration of password, and number of days the user is reminded prior to
password expiration. For information about the specific functionality permitted with each
user type, see User Access Levels on page 13.
}Add a new user—Select Add User. In the User Information section, add a user name,
assign a password, and select a user type from the User Type drop-down list: Admin,
SuperUser, or User. Click Save to create the new user. The name and user type of the
new user appear in the Database Contents and Settings section.
}Edit user information—In the Database Contents and Settings section, select the user to
edit. Select Edit. The User Information section becomes active.
}Remove a user—In the Database Contents and Settings section, select the user to
remove. Select Delete. The user is removed from the user authentication database file.
Illumina User Management Software
MiSeqDx Instrument Reference Guide 13
Figure 6 Adding a New User
Set User Information
In the Database Contents and Settings section, select a user, and then select Edit. Perform
any of the following steps, and then click Save.
}Change a password— In the User Information section, enter a new password in the
Password field, then re-enter the new password in the second password field. Any user
level can change their own password.
}Change a user type—In the User Information section, select a user type from the User
Type drop-down list: Admin,SuperUser, or User. Only an Admin level user can
change a user type.
}Set password duration—In the Database Contents and Settings section, use the drop-
down list to specify the number of days that the current password will remain active.
}Set password reset reminder—In the Database Contents and Settings section, use the
drop-down list to specify the number of days that the user will be reminded to reset the
password prior to password expiration.
Show Logs
Click Show Logs to open a history log of each change made to a database file, and the date
and time that the change was made. Logs are unique to and part of the database file, and
are not stored as separate files. The log that appears is specific to the open database file
listed in the Select a Database section.
User Access Levels
User permissions are grouped into the following user levels and assigned to each user in
the user authentication database file. An Admin level user type is required to change user
type assignments.
MiSeqDx Software
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Level Permissions
Admin Allows access to all software features, the Windows operating system on the
instrument computer, user management activities, and data management
functionality.
Super User Allows mid-level access to software features, the Windows operating system on
the instrument computer, and other functions.
User Allows access to start, monitor, pause, and stop sequencing runs for in vitro
diagnostic kits.
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