Neumodx™ Laboratory Developed Test (LDT) Supplement: Implementation of Ldts On The Neumodx™ 96 and 288 Molecular Systems

NeuMoDx™ Laboratory Developed
Test (LDT) Supplement

Implementation of LDTs on the
NeuMoDx™ 96 and 288 Molecular Systems
© 2020, NeuMoDx Molecular, Inc. All rights reserved. PN 40600250 Rev D

Copyrights
© 2020, NeuMoDx Molecular, Inc. All rights reserved. No part of this publication may be reproduced, transmitted,
transcribed, stored in retrieval systems, or translated into any language or computer language, in any form or by any
means: electronic, mechanical, magnetic, optical, chemical, manual, or otherwise, without prior written permission from
NeuMoDx Molecular, Inc.
The information in this guide is subject to change without notice. NeuMoDx Molecular, Inc reserves the right to change its
products and services at any time to incorporate the latest technological developments. Although this guide has been
prepared with every precaution to ensure accuracy, NeuMoDx Molecular, Inc assumes no liability for any errors or
omissions, nor for any damages resulting from the application or use of this information. NeuMoDx Molecular, Inc
welcomes customer input on corrections and suggestions for improvement.
Trademarks
NeuMoDx, the NeuMoDx logo, and all other trademarks are property of NeuMoDx Molecular, Incorporated.
Patents
www.neumodx.com/patents
Regulatory Information
For Laboratory Developed Tests Only.
PN: 40600250 Rev D

Table of Contents
About This Supplement . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 5

Contents . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 5
Conventions . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 6
Chapter 1: Introduction . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .7
Definitions . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 7
Acronyms . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 7
Additional Documentation . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 8
Technical Support . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 8
Chapter 2: Overview . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 9
LDT Workflow . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 9
Sample Processing Overview . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .10
Liquid Handling Process A (LHPA) . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 10
Lysis/Binding . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 11
Liquid Handling Process B (LHPB) . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 11
XPCR Extraction . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 11
Liquid Handling Process C (LHPC) . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 11
Real-Time PCR . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 12
Chapter 3: Defining an Assay Definition File (ADF) . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 13

Defining an Assay Definition File (ADF) . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 13
Assay Editor General Settings . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 13
Assay Editor Specimen Settings . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .17
Assay Editor PCR Steps . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 20
Assay Editor PCR Targets . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 23
Assay Editor PCR Target Settings . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 24
Result Codes Settings . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 29
Assay Editor Qualitative External Control Settings . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 34
Standard Curve Setting (Quantitative and Qualitative/Quantitative Assays) . . . . . . . . . . . . . 35
Assay Editor Standard Settings (Quantitative and Qualitative/Quantitative Assays) . . . . . . . 37
Assay Editor Calibrator Settings (Quantitative and Qualitative/Quantitative Assays) . . . . . . 38
Assay Editor Quantitative External Control Settings (Quantitative and
Qualitative/Quantitative Assays) . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 40
Assay Editor Reflex Settings . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 42
Assay Editor Localization Settings . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 43
Assay Editor Summary and Export . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 46
Creating an LDT Assay from an Existing Template . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 47
PN: 40600250 Rev D iii

Chapter 4: Creating a Test Order . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 49
Manually Creating a Test Order from the Test Status Display . . . . . . . . . . . . . . . . . . . . . . . 49
Manually Creating a Test Order from the Loaded Specimen Carrier Display . . . . . . . . . . . . 52
Creating a Test Order by Importing a Test Order File in Excel Format . . . . . . . . . . . . . . . . . 57
Creating the Test Order Excel File . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 58
Importing the Test Order File . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 58
Downloading a Test Order File from a Laboratory Information System (LIS) . . . . . . . . . . . . 59
Chapter 5: Running an LDT . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 61

Loading LDT Master Mix Test Strips . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 61
Populating the LDT Primer/Probe Strip with Primers/Probe(s) . . . . . . . . . . . . . . . . . . . . . . 62
Adding Primer/Probe Mix to the LDT Primer/Probe Strip . . . . . . . . . . . . . . . . . . . . . . . . . . . 62
Loading and Unloading LDT Primer/Probe Strips . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 63
Specimen Loading . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 69
Loading Specimen Tube Carriers Initiates the Test(s) . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 69
Test Status . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 71
Completed Tests . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .71
Chapter 6: Extraction Only LDT Testing . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 81

Extraction Only LDT Testing . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 81
Extraction Only Workflow . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 81
Extraction Only LDT Implementation Sample Processing Overview . . . . . . . . . . . . . . . . . . . . 82
Setting Up an ADF for Extraction Only Testing . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 82
Configuring an Extraction Only Test Strip . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 84
Initiating and Completing an Extraction Only Test . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 86
Completed Extraction Only Tests . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 88
Exporting the Extraction Only Mapping . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 89
Chapter 7: Settings . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 93
Report Settings . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 93
Assay Settings . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 94
Standard Curves . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 94
Controls Settings . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 99
Creating User-Defined Controls . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 99
Control Mapping . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .101
Chapter 8: Troubleshooting . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .107

Flags . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .107
Software Flags . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 107
Results Processing Flags . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 107
Index . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 109
iv NeuMoDx Laboratory Developed Test (LDT) Supplement PN: 40600250 Rev D

About This Supplement
This supplement is intended for users of the NeuMoDx 96 Molecular System and the NeuMoDx 288 Molecular
System, referred to hereafter as the NeuMoDx Molecular Systems. It provides information on how to perform
Laboratory Developed Tests (LDTs) on the NeuMoDx Molecular Systems.
This supplement must be used in conjunction with the NeuMoDx 96 Molecular System Operator’s Manual or the
NeuMoDx 288 Molecular System Operator’s Manual for information on running LDTs and system functionality
related to LDT implementation.
IMPORTANT: Read the NeuMoDx Molecular System Operator’s Manual to find detailed information on
instrument operation and maintenance, system accessories, and consumables, as well as all precautions,
limitations, and safety information related to the system. The main operator’s manual also includes information
on software features not necessarily related to a test, such as settings and tools.
Read this supplement carefully prior to use. All instructions in this document must be followed accordingly.
Reliability of the instrument cannot be guaranteed if there are deviations made from the instructions outlined in
this supplement.
For reagent-specific information, refer to the instructions for use (IFU) shipped with each reagent.
CAUTION: Using specimen types other than those validated may not yield expected results. Please
consult with a NeuMoDx Application Specialist before attempting the use of alternate specimen types
in LDT mode.
Contents
This supplement contains the following information:
• Chapter 1, “Introduction” provides a brief description of Laboratory Developed Tests (LDTs) and lists contact
information for NeuMoDx Molecular, Inc. technical support.
• Chapter 2, “Overview” provides general information about Laboratory Developed Tests, including the
workflow and sample processing overviews.
• Chapter 3, “Creating a Test Order” covers how to create a test order.
• Chapter 4, “Defining an Assay Definition File (ADF)” provides information on how to create an assay
definition file (ADF).
• Chapter 5, “Running an LDT” describes how to prepare and load test strips and specimens.
• Chapter 6, “Extraction Only LDT Testing” provides information on running an Extraction Only test using an
RUO ADF.
• Chapter 7, “Settings” covers LDT-specific information in the Settings tab, specifically standard curve
functionality that applies only to LDTs, and control mapping features for LDTs only.
• Chapter 8, “Troubleshooting” covers LDT-specific errors and flags reported by the system.
PN: 40600250 Rev D 5

Conventions
The following conventions are used in this manual. Warnings and cautions appear in boxes with their respective
symbols. Text and keyboard conventions are also provided.
WARNING: The use or misuse of the system could result in damage to the system, minor or severe
injury, or death.
CAUTION: Unsafe practice that could result in damage to the device or other property, data loss,
failure in a procedure, or possible injury.
CAUTION: Contact with biological specimens and materials can transmit a potentially fatal
infection. Use universal precautions when handling biological specimens or instruments that contact
the specimen.
 NOTE: Notes provide helpful information that supplements the topic material.
Boldface type is used to present buttons and options that appear on the screen.
Italics are used to highlight book titles and to emphasize certain terms.
6 NeuMoDx Laboratory Developed Test (LDT) Supplement PN: 40600250 Rev D

Chapter 1
Introduction
In addition to IVD assays, the NeuMoDx™ Molecular Systems can also be used as an open system to process
Laboratory Developed Tests (LDTs) that have been created and validated by your lab. The operation of the system
for LDTs is similar to IVD assays, except:
1 Rather than a single IVD test strip, the system requires the operator to load either an LDT Master Mix RNA (LDT
MM RNA) strip or an LDT Master Mix DNA (LDT MM DNA) strip and an LDT Primer/Probe Strip. The liquid probe(s)
and primers for the LDT are loaded manually by the lab into the wells of the empty LDT Primer/Probe Strip.
2 Your laboratory is responsible for developing and validating the LDT, including designing the test,
determining performance, and completing required verification and validation.
Definitions
The following definitions are provided to distinguish between the following terms used in the rest of the
document:
Specimen refers to clinical material collected from a patient which has been appropriately preserved and
transported to the laboratory for testing on the NeuMoDx Molecular System. Specimens are introduced to the
system in a specimen tube.
Sample refers to an aliquot of a specimen aspirated from the specimen tube during processing by the NeuMoDx
Molecular System.
 NOTE: The terms system and instrument are used interchangeably.
Acronyms
Acronym Term
ADF Assay Definition File

IFU Instructions for Use
LDT Laboratory Developed Test
LHPA, LHPB, LHPC Liquid Handling Process A, B, C
LHR Liquid Handling Robots
LIS Laboratory Information System
PCR Polymerase Chain Reaction
XPCR The combination of sample extraction and PCR
PN: 40600250 Rev D Introduction 7

Additional Documentation
The following table lists the available documents required for proper instrument operation.
Document Description
Instructions for Use (IFUs) These documents contain important information on the use, storage,
performance, and limitations of the reagents.
Safety Data Sheets (SDS) Provide chemical hazard information for reagents and consumables used on
the NeuMoDx Molecular Systems.
Technical Support
For technical questions regarding the product:
• Read the section of the appropriate NeuMoDx Molecular System Operator’s Manual specific to the operation
you are performing. Use the table of contents and index to locate the information.
• See the Troubleshooting section in the operator’s manual.
Contacting NeuMoDx
If additional assistance is required or a question arises, which is not answered in this supplement or the
operator’s manual, contact NeuMoDx technical support:
1250 Eisenhower Place
Ann Arbor, MI 48108
Phone: 1.734.477.0111 or 1.888.301.6639
When contacting NeuMoDx, have the following information available:
• Product name, part number, and serial number
• Any error messages you encounter
• Details of recent instrument performance

Chapter 2
Overview
LDT Workflow
Qualitative LDT workflow Quantitative LDT workflow
QUALITATIVE LDT QUANTITATIVE LDT

IMPLEMENTATION IMPLEMENTATION
Define ADF in ƐƐĂǇ Editor Define Initial ADF [Qualitative]
LOAD GENERIC CONSUMABLES LOAD GENERIC CONSUMABLES LOAD TEST SPECIFIC

x NeuMoDx Cartridges REAGENTS
x NeuMoDx Cartridges LOAD TEST SPECIFIC
x NeuMoDx Extraction Plates x NeuMoDx Lysis Buffer
x NeuMoDx Extraction Plates REAGENTS
x 100uL Tips x LDT Primer/Probe Strips
x 100uL Tips x NeuMoDx Lysis Buffer
x 300uL Tips x LDT Quantitative
x 300uL Tips x LDT WƌŝŵĞƌͬWƌŽďĞ Strips x LDT MM, DNA or LDT MM, RNA Standards
x LDT MM, DNA or LDT MM, RNA
Process YES
Process External/ Define Standard
Quantitative Is SC Valid?
User-Defined Curve (SC)
Standards
Controls
STANDARD CURVE
YES
FINALIZE QUANTITATIVE ADF
Are Controls
Valid? Repeat as prompted
by NeuMoDx
Software
YES
Are YES Process External/
Process External Are Controls
Calibrators User-Defined
PROCESS ROUTINE SAMPLES Calibrators Valid?
Valid? Controls
Controls and Calibrators YES
PROCESS ROUTINE SAMPLES
PN: 40600250 Rev D Overview 9

Sample Processing Overview
As specimens are loaded into the system, sample processing is initiated. Sample processing involves the
following steps:
No. Step Description
1 Liquid Handling Process A Samples are mixed with lysis buffer in the extraction plate.
(LHPA)
2 Lysis/Binding Cell lysis and nucleic acid binding takes place in the extraction plate.
3 Liquid Handling Process B The lysate and magnetic bead mixture is aspirated from the extraction
(LHPB) plate and loaded into the cartridge.
4 XPCR Extraction Further purification and release of bound nucleic acid occurs within the
cartridge.
5 Liquid Handling Process C Eluted nucleic acid is mixed with user-supplied primers and probe(s) in the
(LHPC) LDT Primer/Probe Strip, transferred to and mixed with dried general-
purpose PCR or RT-PCR reagents (LDT MM, RNA or DNA) in the test strip and
then delivered into the PCR regions of the cartridge.
6 PCR/Real-Time PCR Thermal cycling of the PCR mix occurs in the PCR regions of the cartridge
lane to amplify and detect the desired target(s) and sample process control.
Liquid Handling Process A (LHPA)

LHPA refers to the process of the Liquid Handling Robot (LHR) adding appropriate amounts of Lysis/Binding
buffer and specimen to the extraction plate prior to initiating lysis. In this process, the LHR:
• Picks up 1000 μL tips for samples
• Moves to the lysis buffer and aspirates the desired volume of buffer
• Moves to the appropriate wells in the extraction plate, pierces the foil covering each well and dispenses the
buffer into the extraction plate wells from all tips
• Returns the tips to the tip rack for later use
• Picks up additional 1000 μL tips
• Moves to specimen tubes and aspirates test-specific sample volume
• Moves to corresponding extraction plate wells
• Simultaneously dispenses all samples into the extraction plate wells
• The instrument turns on the required extraction plate heaters to the test-specific temperature
• Mixes the entire volume contained in each extraction plate well
• Disposes of the 1000 μL tips into the biohazard waste
 NOTE: Once tips have been used for specimen aspiration, the Liquid Handling Robot does not allow the used
tips to travel directly above other consumables.

Lysis/Binding
At the end of the LHPA process, the extraction plate heaters in the extraction plate module are turned on to their
specified, matrix-specific lysis temperature. The extraction plate heaters perform the following steps:
• Heat the prepared specimen with the reconstituted extraction plate reagents for a predefined time
(10 minutes or less) to lyse samples, bind nucleic acids to affinity coated paramagnetic microspheres, and
prepare samples for further purification within the cartridge.
• Lysis temperatures are determined by supported sample type (DNA/RNA) and matrix.
Liquid Handling Process B (LHPB)

LHPB refers to the process of aspirating the mixture from the extraction plate and delivering it to one or more
lanes of the microfluidic cartridge in the XPCR Module. In this process, the Liquid Handling Robot (LHR):
• Picks up the 1000 μL tips associated with each sample position for LHPA. These tips were previously used to
dispense buffer and did not come into contact with the specimen.
• Thoroughly mixes test-specific volume contained in the extraction plate well
• Aspirates the entire volume for all the wells simultaneously
• Moves to the sample injection port (referred to as S-port) of the first unused lane of a cartridge in an XPCR
Module
• Using independent z-axis motion and TADM pressure sensing, ensures each tip is properly mated with the
S-ports in the cartridge lane
• Dispenses the entire volume into the cartridge lane
• Disposes of the 1000 μL tips into the biohazard waste
 NOTE: Once tips have been used for sample delivery into the cartridge, the Liquid Handling Robot does not
allow the used tips to travel directly above other consumables.
XPCR Extraction
XPCR Extraction refers to the steps performed in a cartridge loaded into an XPCR Module to further purify nucleic
acid with Wash Reagent and release bound nucleic acid by a combination of Release Reagent and heat.
Liquid Handling Process C (LHPC)

For Open System/LDT implementation, LHPC refers to the process of aspirating the eluted nucleic acid from the
cartridge, mixing it with the user-supplied primers/probes, using the mixture to reconstitute the dried LDT
Master Mix reagents in the test strip, and then loading the PCR/RT-PCR ready solution into the appropriate lane
on the cartridge housed in the XPCR Module. In this process, the LHR:
• Picks up new 300 μL tips for each discrete eluted nucleic acid sample
• Punctures appropriate foil-covered LDT MM (RNA or DNA) test strip wells
• Moves to the appropriate PCR injection port (referred to as the P-port) on the cartridge, and using
independent z-axis control, aspirates all of the eluate (combined with the XPCR Module syringe pump
pushing air)
PN: 40600250 Rev D Overview 11

• Moves to the LDT Primer/Probe Strip and dispenses the entire volume of the eluate into the well containing
the user-supplied Primers/Probes
• Thoroughly mixes the volume contained in the LDT Primer/Probe Strip well
• Using independent z-control, finds the bottom of each well and aspirates all of the eluate plus primers/
probes
• Moves to previously punctured wells in the LDT Master Mix test strip and dispenses the entire volume of eluate
• Thoroughly mixes the volume contained in the LDT Master Mix test strip well
• Using independent z-control, finds the bottom of each well and aspirates nearly all of the eluate
• Moves back to the same P-ports in the cartridge
• Dispenses the entire volume into the cartridge
• Disposes of the 300 μL tips into the Biohazard Tip Waste Bin or Biohazard Waste Container (for the NeuMoDx
96 Molecular System or NeuMoDx 288 Molecular System, respectively)
 NOTE: Once tips have been used for aspiration of the eluted material, the Liquid Handling Robot does not
Real-Time PCR
The process of thermal cycling the PCR mix in the amplification channel of the cartridge lane to amplify the
desired target(s) and sample process control. This process:
• Confirms all cartridge valves for active lanes are locked or closed
• Thermally cycles the amplification chamber as per specified heating profile
• Detects fluorescence in desired optical channels from the specified amplification channels of the specific
cartridge lane

Chapter 3
Defining an Assay Definition File (ADF)
Defining an Assay Definition File (ADF)

Three types of LDTs are available—qualitative, quantitative, and qualitative/quantitative. LDTs are defined
using the Assay Editor Wizard in the Assay tab under Tools. The defined test method is called the Assay
Definition File (ADF). You must create an ADF for qualitative, quantitative, and qualitative/quantitative LDTs.
 NOTE: Only Supervisor or Biomed-level users can define an ADF.
Assay Editor General Settings

Use the Assay Editor Wizard in the Assay tab to create an ADF for LDTs.
1 To access the Assay Editor Wizard, select Tools, then Assay.
The wizard guides you through the steps to set up an LDT ADF.
2 Select Create New, then select Next.
PN: 40600250 Rev D Defining an Assay Definition File (ADF) 13

The Assay Editor General Settings screen is displayed. LDT is automatically selected for Assay Type. This
cannot be modified.
3 Select the Result Type (Qualitative, Quantitative, or Qualitative/Quantitative).
4 Select a Chemistry Type (RNA or DNA).

Once a Chemistry Type is selected, more options are displayed on the touchscreen.
 NOTE: Fields are colored teal if any changes are made.
5 Select the Master Mix Test Strip that will be used for the LDT.
6 Enter an Assay Display Name (1 to 30 characters).
7 Enter an Assay Name (1 to 15 characters).

8 The Assay Version defaults to 1.0.0.
The version may be changed, but must be in the format X.X.X. Each time the assay is updated, the Assay
Version will automatically increment, but you may change it.
9 The Release Date is displayed.

The default is the current date, but you may change it using the calendar.
10 Select a Classification.
• Select RUO (Research Use Only) for assay optimization and development.
• Select LDT for the final optimized assay.
11 (Optional) Enter a barcode.

Entering a barcode will allow you to use a CSR test strip.
 NOTE: Do not enter a barcode when developing your own LDT.
12 The number of Extractions is set to 1. This cannot be modified.
13 Scroll down to see more General Settings
14 The number of PCR Mixes is set to 1. This cannot be modified.
15 RP (Results Processing) Version is set to V2. This value cannot be changed.
16 Select a Result Processing Algorithm.

• Ct and EP (default): Uses a combination of the calculated Ct value and End Point (EP) fluorescence to
determine pass/fail criteria. This is the recommended setting.
• Ct: Uses only the calculated Ct value to determine pass/fail criteria.
17 Choose a desired Ct Calling Algorithm.

• Second Derivative (default): Uses a NeuMoDx-developed algorithm to determine the Ct value. This is the
recommended setting.

• Threshold: Uses a specified threshold line to determine the Ct value.
18 Select Starting Fluorescence Start and End Cycles.

Together, the Starting Fluorescence Start Cycle and Starting Fluorescence End Cycle define the cycles used
for background correction in the calculation of a Ct value.
• The Start Cycle must be greater than 1 and less that the Starting Fluorescence End Cycle.
• The End Cycle must be less than the total number of cycles and greater than the Starting Fluorescence
Start Cycle.
General guidelines for Starting Fluorescence Start and End Cycles:

• This window between start and end cycles (End Cycle – Start Cycle) needs to have a minimum of 1 cycle.
Typically, it spans ~3 cycles.
• Typically, this value should be ~3-8 cycles before the earliest anticipated Ct value for the LDT.
CAUTION: These guidelines are not meant as specific recommendations. All analysis parameters for
LDT use must be developed and validated by the lab.
19 Select End Point Fluorescence Start and End Cycles.

Together, the End Point Fluorescence Start Cycle and End Point Fluorescence End Cycle define the cycles
used for calculation of the overall End Point Fluorescence. A valid Ct value must be called before the End
Point Fluorescence Start Cycle.
• The End Point Fluorescence Start Cycle must be greater than the Starting Fluorescence End Cycle and less
than the End Point Fluorescence End Cycle.
• The End Point Fluorescence End Cycle must be greater than or equal to End Point Fluorescence Start Cycle
and less than the total number of cycles.
General guidelines for End Point Fluorescence Start and End Cycles:
• This window between start and end cycles (End Cycle – Start Cycle) can have a minimum of 0 cycles.
Typically, it spans ~3 cycles.
• Typically, this should include cycles close to or at the last cycle value for the LDT.
CAUTION: These guidelines are not meant as specific recommendations. All analysis parameters for
LDT use must be developed and validated by the lab.
20 Enter a time (in hours) for the Open-Life.

Open-Life refers to the LDT Primer/Probe Strip which holds the user-supplied primers/probes for an LDT. It
enables the system to track the onboard age of the LDT Primer/Probe Strip and invalidate it for use for any
test started beyond the set Open-Life.
 NOTE: While this parameter is also required for ADFs being made for extraction only testing, it does not
affect the Open-Life of test strips mapped for extraction only testing. See Chapter 6, “Extraction Only LDT
Testing” for information on extraction only testing.

21 Select a Fill Check Reporter.
Together, the Fill Check Reporter and Fill Check Threshold (below) are used to monitor and ensure that the
PCR mix reaches and fills the PCR portion of the cartridge lane. If the Fill Check Threshold is not met for the
specified Fill Check Reporter, an Indeterminate (IND) result is reported, representing a system error.
• Yellow (530/550), Red (625/660), Green (470/510), Orange (585/610), or Far Red (680/715) may be
selected as the Fill Check Reporter. Selecting a channel used for an assay target as the Fill Check Reporter
is recommended for LDT as it will also require the presence of a probe for that reporter in the LDT Primer/
Probe Strip to pass the Fill Check.
22 Enter a Fill Check Threshold.

The Fill Check Threshold must be between 0 and 10,000.
23 Select Next to proceed to the Assay Editor Specimen Settings.
Assay Editor Specimen Settings

The Assay Editor Specimen Settings is used to define the specimen type, specimen stability, specimen aspirate
volume, and lysis and binding duration in the extraction plate.
• At least one specimen type must be defined.
• If more than one specimen type is defined, one must be designated at the default specimen type.

To enter Specimen Settings
1 Select Add to add a specimen type.
2 Select a Specimen Type.

Only supported specimen types are displayed.
3 Check the box to treat the first rerun as a direct repeat with no changes to processing settings. If checked,
the given assay/specimen type will use repeat parameters when an Unresolved result is reported for that
specimen, assuming both Repeat and Rerun are checked in the Assay tab under Settings (see “Assay
Settings” on page 94).
• If a sample has an Indeterminate result (IND), it will repeat (if Repeat is checked in the Assay tab under
Settings).
• If a sample has an Unresolved result (UNR), it will use the rerun parameters set in the ADF, unless the user
would rather directly repeat those UNR samples first, instead of using the different settings.
• If the “Treat first rerun as repeat” box is checked, and Repeat and Rerun are enabled in the Assay tab
under settings, then the system will perform a Repeat, then a Rerun (using Rerun settings) if it is still
UNR.
4 Enter the onboard Specimen Stability time (1 to 48 hours).

The specimen stability enables the system to track the onboard age of the specimen and invalidate it for use
after a certain time on the system worktable.
5 If applicable, check the Enable Calibration Factor box to add a calibration factor for the specimen type
(quantitative and qualitative/quantitative assays only).
If checked, you must enter a calibration factor. This calibration factor (in Log) will be applied to quantitative
results for samples defined with that specimen type. This factor can be used to adjust the concentration for
specimen volume differences between two matrices when using a shared standard curve.
6 Select the Specimen Aspirate Volume.

Only supported specimen aspirate volumes are displayed.

7 Enter the Specimen Mix Volume (5 to 900 μL).
8 Enter the Rerun Specimen Mix Volume (5 to 900 μL).
9 Enter the Specimen Mix Count (0 to 10).
10 Enter the Rerun Specimen Mix Count (0 to 10).
11 Scroll down to see more Specimen Settings.
12 Select the Rerun Specimen Aspirate Volume.

Only supported specimen aspirate volumes are displayed.
13 If applicable, check the Enable Rerun Calibration Factor box to add a calibration factor only for samples that
are rerun/repeated (quantitative and qualitative/quantitative assays only).
If checked, you must enter a calibration factor. This calibration factor (in Log) will be applied to quantitative
results for samples defined with that specimen type that are rerun/repeated by the system.
14 Select the Buffer type.

The buffer type is set based on specimen and chemistry type.
The buffer type can only be changed if Use Override Buffer is checked. Once checked, a new Override Buffer
drop-down menu appears, in which a different buffer type can be selected. Checking the box allows you to
select a re-run buffer, not the buffer. The buffer is greyed out when you check the box.
15 Select the Rerun Buffer type.

The re-run buffer type is set based on specimen and chemistry type.
The re-run buffer type can only be changed if Rerun Use Override Buffer is checked. Once checked, a new Re-
Run Override Rerun Buffer drop-down menu appears, in which a different buffer type can be selected.
16 Select the Extraction Plate Heater Temperature (°C) [Low, Medium, or High].
This is the temperature at which lysis and binding of the nucleic acid to the magnetic particles occurs in the
extraction plate.
• High: ~ 60°C

• Medium: ~45°C
• Low: ~37°C
17 Enter a Lysis Duration (1 to 1,200 seconds).

The recommended value is between 420 and 600 seconds.
18 If desired, select Add to add a second specimen type to the assay. Then populate all the fields as in the steps
above.
• To change the default specimen type to the newly created type, select Set Default.
• To remove a specimen type, select it, then select Remove.
19 Select Next to proceed to Assay Editor PCR Steps.
Assay Editor PCR Steps

The Assay Editor PCR Steps screen is used to define the PCR thermal cycling profile for the assay. There are two
types of PCR Stages—Hold Stage and Cycle Stage. Each Cycle Stage is comprised of a number of steps, typically
denature and anneal setpoints. Detection can be set to occur on the 2nd or 3rd step of a cycling stage.
Defining the Number of PCR Stages

The PCR Stages field is pre-populated with two stages—one Hold Stage and one Cycling Stage.
• The maximum number of PCR Hold Stages is five.
• The maximum number of PCR Cycle Stages is five.
• The total number of PCR cycle counts across stages is 60.

To define the number of PCR stages
1 Select Add to add another PCR Stage.
2 Select Remove to remove a PCR Stage. The highlighted stage is removed.
3 Use the Move Up and Move Down buttons to change the order of each stage in the PCR Stages list.
Defining a Hold Step

Hold Steps are typically used for polymerase activation/inactivation, reverse transcription, etc.
To define a Hold Step

1 (Optional) Enter a Label (up to 20 characters).
2 Enter a duration (2 to 1,200 seconds).
3 Enter a temperature (45°C to 99°C).

Defining a Cycling Step
By default, two Cycle Steps are defined—Denature and Anneal. These steps may be deleted, or the labels may be
changed.
Only one step in each PCR Cycle step can perform fluorescence detection via the optics module.
For each PCR cycling stage

1 Select the PCR Cycle Stage in the PCR Stages box.
2 (Optional) Enter a Label (up to 20 characters) for the highlighted PCR Cycle Stage.
3 Enter a Cycle Count (1 to 60 cycles).

The default Cycle Count is 45.
To define a Cycle Step

1 Select Add to add PCR Steps in a Cycling Stage.
To remove the highlighted PCR Step in a Cycling Stage, select Remove.
2 Select a Cycle Step in the PCR Steps box to edit the settings for that Cycle Step.
3 (Optional) Add or change the Label (up to 20 characters) for the highlighted Cycle Step.
4 Enter a Duration (2 to 63 seconds).
5 Enter a Temperature (45°C to 99°C).
6 Repeat the above steps for all Cycle Steps in the PCR Stage.
7 Check the box next to Detect? if fluorescence detection is required on that Cycle Step.
Fluorescence detection is required for Anneal steps.
8 Repeat for all Cycle Steps across all Cycle Stages.

9 When the cycling profile is defined for all PCR Steps and Stages, select Next to proceed to the Assay Editor
PCR Targets.
Assay Editor PCR Targets

The Assay Editor for PCR Targets screen is used to define and match the desired targets with a fluorescent
detection color/channel, as well as set up detection settings and parameters for each target.
• For quantitative assays, you must have exactly one target defined as a sample process control and one target
that is not defined as a sample process control.
• For qualitative assays, you must have exactly one target defined as a sample process control and at least one
(and up to five) target that is not defined as a sample process control.
 NOTE: The NeuMoDx Sample Process Control (SPC1 for DNA, SPC2 for RNA) is present in every extracted
sample. The channel on which the SPC is detected is specific for the selected Master Mix Test Strip type. Refer
to the Master Mix Test Strip IFU.
To add PCR Targets

1 Select PP1 in the Primer/Probes box.
The SPC1 or SPC2 target is automatically populated in the detection list. This channel is used to detect the
sample processing control which is co-extracted and amplified with every specimen.
2 Select Add to add additional targets to the detection list. You can change the reporter in step 5 below.
3 Select a target in the detection list to display the parameters to set for that target and fluorescence detection
channel.
4 Enter a target Name (1 to 20 characters). Each target must have a unique name.
5 Select a Reporter or Detection Channel.

• Red (625/660)
• Green (470/510)

• Orange (585/610)
• Far Red (680/715)
• Yellow (530/555)
 NOTE: The appropriate channel for the Master Mix Test Strip type must be selected as the Sample Process
Control.
6 Select Next to proceed to Assay Editor PCR Target Settings.
Assay Editor PCR Target Settings

The Assay Editor PCR Target Settings are used to set up detection settings and parameters for each PCR target
added in the previous section.
To adjust PCR Target Settings

1 Select PP1 in the Primer/Probes box.
2 Select Add to add the target to the detection list.

3 Select a target in the detection list to set the parameters for that target and fluorescence detection channel.
4 Select the Specimen Type from the drop-down list.
5 Enter a Minimum Peak Height.

Typically start with a peak height of 10 and gradually increase or decrease this value depending on observed
LDT sensitivity and specificity.
6 Enter Peak Minimum and Maximum Cycles.

These are the earliest and latest cycles that a Ct would be generated for a specific target allowing for a
positive call.
7 Enter a Minimum End Point Fluorescence.

This is the minimum end point fluorescence value, determined from the last programmed cycle, that would
enable a positive call. This value is only used if Ct+EP is chosen as the Results Processing Algorithm on the
Assay Editor General Settings screen.
8 Enter the Max Outliers.

NeuMoDx software automatically executes an outlier removal algorithm to enable efficient result processing
and analysis. This parameter allows you to define the maximum number of outliers that may be removed
automatically during the results processing step. An Indeterminate (IND) result is returned if the number of
outliers exceeds this value.
9 Enter a Baseline Start Cycle.

This is the first cycle to be used in the baseline calculation for each target for use in normalization.

10 Scroll down for more PCR Target Settings. The list of parameters will vary depending on whether you are
creating an ADF for a quantitative assay or qualitative assay.
11 Depending on what you selected for the Ct calling algorithm on the Assay Editor General Settings screen (see
step 17 on page 15), complete the following options.
• If the Ct calling algorithm is set to Second Derivative and the Second Derivative Ct calculation does not
yield a valid baseline:
– Enter the Fixed Baseline Start and Fixed Baseline End cycles.
– Enter the number of Baseline Lookback Cycles.
This is the number of cycles that are subtracted from the calculated Ct value to determine the last cycle
to be used in the baseline calculation for each target.
– Enter the Baseline Minimum Window Cycles.
The minimum number of cycles to be used in the baseline calculation for the algorithm to be valid for
each target.
NeuMoDx software automatically executes a linear regression fit on the baseline (flat section of the
PCR curve) to normalize the fluorescence curves obtained during PCR. The ability to define thresholds
on this parameter allows you to define the upper and lower bounds of an acceptable baseline slope. An
Indeterminate (IND) result is returned if the baseline slope is out of this range.
– Enter the Baseline Slope Lower Threshold.
This is the threshold value for the lower bounds of an acceptable baseline slope.
– Enter the Baseline Slope Upper Threshold.
This is the threshold value for the upper bounds of an acceptable baseline slope.
• If the Ct calling algorithm is set to Threshold, enter the Baseline End Cycle.
12 Enter a Maximum Starting Fluorescence.

The highest starting raw fluorescence value that may be used for the algorithm to be valid
13 Enter an Inhibition Check cycle.

This check is intended to help detect inhibition that is not attributed to competition between two targets.

When multiple targets are assigned, if a positive target amplifies with a Ct > Inhibition Check Cycle AND the
internal control is not detected (or Ct is outside of acceptable range), any other non-positive targets will be
called Unresolved.
14 If the Ct calling algorithm is set to Threshold, enter the Ct Threshold.
15 Enter an EPR Threshold.

End Point Ratio (EPR) – The EPR is used to provide a ratio of the fluorescence increase. This value is
calculated as End Point Fluorescence [average]/Starting Fluorescence [average], and can be used as an
additional parameter to improve the confidence of identifying successfully amplified PCR curves using the
NeuMoDx software results processing algorithm.
When the EPR value is used to determine amplification status of a PCR curve, the EPR Threshold value
represents the minimum EPR value required to demonstrate successful amplification. In the event other
parameters (Ct and EP) supporting successful amplification are obtained, the EPR Threshold provides
another layer of stringency and helps mitigate false amplification detection.
16 Enter an EPR Check Ct Threshold.

The EPR Check Ct Threshold parameter is used to set the Ct value below which the EPR Threshold will be used
to determine the amplification status (in addition to Ct or Ct and EP).
If Ct < Ct (EPR check), then EPR must be > EPR min.
An Indeterminate (IND) result is returned if the Ct value is ≤ EPR Check Ct Threshold AND the EPR obtained is
< EPR Threshold. The EPR Check Ct Threshold value should be set to 0 to disable the EPR check and set to a
value equal to the last cycle, if desired to be implemented for all tests.
17 Choose the EPR Check Failed Result.

The drop-down will show two options: Indeterminate and Negative.
• An Indeterminate result is preferred when samples with low-end fluorescence could potentially be
positive or false-positive. In this case repeat testing is desired.
• A Negative result is preferred when samples with low-end fluorescence are to be considered negative.
18 Proceed to the appropriate section:

• For quantitative and qualitative/quantitative assays, see additional parameters in “PCR Target Settings
for Quantitative and Qualitative/Quantitative LDT” in the following section.
• For qualitative assays, proceed to “Result Codes Settings” on page 29.

PCR Target Settings for Quantitative and Qualitative/Quantitative LDT
If you are performing a quantitative assay and defining the settings for a target other than the sample process
control (SPC1 for DNA or SPC2 for RNA), continue with the following parameters.
19 Enter the Lower and Upper Limit of Quantification (log Conc.).

Used in Log Concentration quantitation determination.
• If the calculated log concentration is less than the Target Lower Limit, the software applies the Result
Below Target Lower Limit flag.
• If the calculated log concentration is greater than the Target Upper Limit, the software applies the Result
Above Target Upper Limit flag.
20 Select the units of concentration to display on the Sample Results Report.

Units include:
• IU/mL
• IU/μL
• c/mL
• c/μL
• ng/mL
• ng/μL
• None
21 (Optional) Check the box next to “Enable Conversion Factor” to enter a Concentration Conversion Factor.
This will convert the reported value to a different unit using a conversion factor, if desired.
Select the Concentration Conversion Factor Display Unit from the drop-down, the options are identical to the
Concentration Display Unit. The selected option cannot be the same as the Concentration Display Unit.
22 (Optional) Enter the Quant Check Ct Threshold.

Used to define the cycle after which the sample process control shift could be an indicator of a system or
sample issue and not due to target competition. If the calculated Ct is greater than the Quant Check Ct

threshold and the sample process control is not amplified (past IC peak maximum cycle), then the
Quantitation Error flag will be present.
 NOTE: The default is the last cycle defined for the assay.
23 (Optional) Check the box next to “Enable Quant Adjustment.”
If Enable Quant Adjustment is selected, the actual quantitation is adjusted based on how much the sample
process control is delayed past the peak maximum cycle of the sample process control.
 NOTE: If Quant Check Ct Threshold is set, but Enable Quant Adjustment is disabled, then an adjusted
quant is not calculated and the quantitation error informational flag is applied.
24 Repeat these steps to define the PCR Target Settings for each Target added to the Target Detection List.
25 Select Next to proceed to Result Code Settings in the following section.
Result Codes Settings

The Result Code identifies the assay run for a particular specimen throughout the system options, in raw data
and in reports. Result Codes are also used in Test Orders to tell the system which assay is to be run for those
samples. Controls that are run for the assay will be associated with the Assay Name defined on the Assay Editor
General Settings screen, and the results will apply to all Result Codes defined under that Assay Name.
The Result Name is displayed in the Test Status tabs when samples are loaded or a Test Order is imported into the
system. The Result Name and Result Code are both used in the ADF Summary to identify the Target(s) to be
reported. The Assay Name defined on the Assay Editor General Settings screen is the default Result Name, but
this can be modified, if desired.
A quantitative assay only allows for one target, thus only allowing for one Result Code to be defined. Additional
optional steps for defining Result Codes for qualitative assays are explained in the section “Result Codes Settings
(Qualitative Assays)” on page 31.

To define a Result Code for Quantitative assays and Qualitative assays with only one target
1 Select Add to add a new Result Code. A Result Code must be defined for every assay.
2 Select the assay name in the box at the top of the screen.
3 Enter a Result Code (2 to 4 characters and must begin with the letter “L”). A Result Code is populated by
default and can be changed.
4 Enter a Result Name (1 to 15 characters). The Assay Name defined on the Assay Editor General Settings screen
is populated by default.
The Result Name cannot contain the special characters / \ : * ? " < > |.
5 This Result Type defaults to the Result Type defined on the Assay Editor General Settings screen. This cannot
be changed.
6 Check the box next to the Target Name to define the target. The target is automatically selected by default.

7 Select Next to move to the next Assay Editor screen, which is the Standard Curve Setting screen for
Quantitative and Qualitative/Quantitative assays or the Assay Editor Qualitative External Control Settings for
Qualitative assays.
Result Codes Settings (Qualitative Assays)

A qualitative LDT assay can have multiple targets in addition to an internal control. A Result Code must be
defined for all targets. Additionally, you have the option to define Result Codes for the targets separately. This
will allow you to run specimens using the same test strip but detect and report on only one of the defined
targets, if desired.
Each Result Code must be unique. It is recommended, but not mandatory, to use different Result Names to aid in
the identification of the targets being tested under each Result Code.
To define a Result Code for assays with multiple targets

1 Select Add to add a new Result Code for a target or set of targets.
Select Remove to remove the selected Result Code and accompanying inputs. At least one Result Code must
be defined for all targets.

2 Enter a Result Code (2 to 4 characters and must begin with the letter “L”) that is different from the other
Result Codes defined for the assay.
3 Enter a Result Name (1 to 15 characters). The default Result Name is the Assay Name defined on the Assay
Editor General Settings screen.
4 Check the box next to the target or target set you want to be associated with this Result Code.
5 Follow these steps again for each Result Code you would like to define.
6 Select Next once you have defined all applicable Result Codes.

Result Codes Settings (Qualitative/Quantitative Assays)
A qualitative/quantitative LDT assay is used to detect the target of an assay either qualitatively or quantitatively
using the same test strip. A Result Code must be defined for both the qualitative and quantitative Result Types.
Each Result Code must be unique. It is recommended, but not mandatory, to use different Result Names to aid in
the identification of the Result Type selected for each Result Code.
To define a Result Code for Qualitative/Quantitative assays

1 Select Add to add a new Result Code for the Result Type. Select Remove to remove the selected Result Code
and accompanying inputs. A Result Code must be defined for each Result Type.
2 Enter a Result Code (2 to 4 characters and must begin with the letter “L”) and a Result Name (1 to 15
characters) for each Result Type. The default Result Name is the Assay Name defined on the Assay Editor
General Settings screen.
3 Select Next once you have defined all Result Codes.

Assay Editor Qualitative External Control Settings
The Assay Editor Qualitative External Control Settings screen is an optional way to set up and define external
controls for qualitative assays. If external controls are set up in this screen, the system treats them as required
and prompts for them at the defined frequency. User-defined controls can also be set up by assay in the Controls
tab under Settings (see “Controls Settings” on page 99).
For qualitative assays, you can only define controls. For quantitative assays, you must define either a standard
curve or standards that can be used to generate a standard curve. Additionally, calibrators and external controls
may be defined. For information on external controls for quantitative assays, see “Assay Editor Quantitative
External Control Settings (Quantitative and Qualitative/Quantitative Assays)” on page 40.
To define external controls for qualitative assays

The Assay Editor Qualitative External Control Settings screen is displayed after selecting Next from the Result
Codes Settings screen.
1 Enter the desired External Controls Run Frequency (1 to 190 days).
2 Select Add to add an External Control Set.

A default External Control is automatically populated based on the specimen type.
3 If applicable, select the Specimen Type associated with the external control set.
4 Enter a Name (1 to 20 characters).
5 Select the Specimen Type from the drop-down menu.
6 (Optional) Check the box next to Assign Mapping.

Check this box if you want an optional specimen ID (barcode) to be linked to the standard. If a specimen ID
(barcode) is specified, the Specimen Type must also be selected.
7 Select the Expected Amplification State for the target(s) defined in your assay (Target Not Amplified or
Target Amplified).
8 To add additional external controls, select Add or Copy and repeat the steps above.
To remove an external control, select Remove.
9 Select Next to proceed to “Assay Editor Reflex Settings” on page 42.
Standard Curve Setting (Quantitative and Qualitative/Quantitative Assays)

The Standard Curve Setting screen is an optional way to define a standard curve within the ADF. If a standard
curve is set up in this screen, the system will treat it as a “Master Standard Curve,” meaning it will be used for the
calculation of concentrations for all samples run using that ADF. Calibrators, if defined, will be applied to the
defined standard curve and used to generate a calibration coefficient. For more information, see “Assay Editor
Calibrator Settings (Quantitative and Qualitative/Quantitative Assays)” on page 38.
Standard curves can also be defined in the Assay tab under Settings. See “Standard Curves” on page 94. In this
case, the standard curve can be either manually entered or generated using standards defined here in the ADF.
You may wish to define the standard curve in the ADF if you plan to use the same standard curve with every lot of
test strips. If you plan to use a new standard curve with each lot of test strips, consider either running standards
and generating a standard curve in the Assay tab, or manually entering a standard curve in the Assay tab.

To define a standard curve
1 Select the check box next to Define Standard Curve in ADF. Or select Next to continue without defining a
standard curve and proceed to defining standards.
2 Enter the following information:

• Name (5 to 20 characters)
• Slope (value between –4 and –2, with a maximum of 4 decimal places)
• Y intercept (acceptable range is dependent on the number of cycles for the test with a maximum of
4 decimal places)
• R2 (between 0.9 and 1 with a maximum of 4 decimal places)
3 Select Next to proceed to Assay Editor Calibrator Settings.

Assay Editor Standard Settings (Quantitative and Qualitative/Quantitative
Assays)
Use the Standard Settings if you wish to optionally run standards for defining a standard curve outside of the
ADF. See “Standard Curves” on page 94 for information on generating the standard curve.
To define standards for use in generating a standard curve

1 Some standards are already populated. Select Next to continue without changing the concentrations of the
populated standards and proceed to defining calibrators. Or complete the following steps to change the
populated standards.
2 Select a standard to set its parameters.
3 Enter the following information for the selected standard:

• Enter a name for the standard (1 to 20 characters).

• Enter the Concentration in Log10 (–5 to 15).
• Enter the number of Replicates (1 to 6).
This is the number of replicate samples processed from the same specimen tube.
• Assign Mapping
Check this box if you want an optional specimen ID (barcode) to be linked to the standard. If a specimen
ID (barcode) is specified, the Specimen Type must also be selected. Only Specimen Types supported by the
assay are displayed.
4 To add additional standards, select Add or Copy and repeat the steps above.
To remove a standard, select Remove.
5 Select Next to proceed to Assay Editor Calibrator Settings.
Assay Editor Calibrator Settings (Quantitative and Qualitative/Quantitative

Assays)
Calibrator Settings allow you to optionally define and run calibrators that the software will use to generate a
calibration coefficient. The calibration coefficient is used to automatically adjust the standard curve for slight
variations between different lots of Primer/Probe Mix, Master Mix Test Strips, or Systems.

To define calibrators
1 Select Add to add a Calibrator Set. Or select Next to continue without defining calibrators, and proceed to
external controls.
A calibrator will be populated with the specimen type selected based on the previously defined specimen type
in the ADF.
2 Enter a name for the calibrator (1 to 20 characters).
3 Enter the Concentration in Log10 (–5 to 15).
4 Enter the Concentration Lower and Upper Limit in Log10 (–5 to 15).
5 Enter the number of Replicates (1 to 6).

This is the number of replicate samples processed from the same specimen tube.
6 Enter the Validity Criteria

The number of replicate samples that must have a valid result. Only valid results are used for the generation
of the calibration coefficient used to adjust the standard curve.
7 Scroll down to see more Calibrator Settings options.
8 Select the Specimen Type.
9 Enter or scan the specimen ID (barcode) for the calibrator.
10 To add additional calibrators, select Add or Copy and repeat the steps above.
To remove a calibrator, select Remove.
11 Select Next to proceed to the Assay Editor Quantitative External Control Settings.
Assay Editor Quantitative External Control Settings (Quantitative and

Qualitative/Quantitative Assays)
The Assay Editor Quantitative External Control Settings screen allows you to optionally set up and define
external controls for quantitative and qualitative/quantitative assays. If external controls are set up in this
screen, the system treats them as required and prompts for them at the defined run frequency. User-defined
controls can also be set up by test in the Controls tab under Settings (see “Controls Settings” on page 99).

To define external controls
1 Enter the desired External Controls Run Frequency (1 to 190 days).
2 Select Add to add an external control set. Each external control set defines the specimen type it applies to
and the controls required for those specimen types.
An external control will be populated with the specimen type selected based on the previously defined
specimen type in the ADF.
3 Enter a name for the control (1 to 20 characters).
4 Select the Expected Amplification State for the target defined in your assay (Target Amplified or Target Not
Amplified).
5 Check the box next to Use specific concentration values? to allow for a quantitative external control with
a defined expected concentration for that target. If not selected, the external control will be treated as a
qualitative control (Amplified or Not Amplified, no specific quantitation required).

If this option is selected, the following options must be defined (scroll to see all options):
• Enter the Concentration in Log10 (–5 to 15).
• Enter the Concentration Lower and Upper Limits in Log10 (–5 to 15).
6 Select the Specimen Type.
7 Assign Mapping
Check this box if you want an optional specimen ID (barcode) to be linked to the standard. If a specimen ID
(barcode) is specified, the Specimen Type must also be selected.
8 To add an additional external control, select Add or Copy and repeat the steps above.
To remove an external control, select Remove.
9 Select Next to proceed to Assay Editor Reflex Settings in the following section.
Assay Editor Reflex Settings

The Assay Editor Reflex Settings allow you to automatically process a new test order based on the qualitative
results of a first test (provided sufficient sample volume is available). This is an optional setting and can be
defined in the ADF or can be user defined in the Assay tab under Settings (see the appropriate NeuMoDx
Molecular System Operator’s Manual).
The Assay Editor Reflex Settings screen is displayed after selecting Next from the Assay Editor Quantitative
External Control Settings (above) for quantitative and qualitative/quantitative assays, or from the Assay Editor
Qualitative External Control Settings screen for qualitative assays.
1 Select Add to define a reflex test process. Or select Next to continue without selecting reflex test settings.
2 Check the box under Target(s) to select which target’s result will prompt the reflex test.

3 Check the box under Specimen(s) to select the specimen type for the reflex test.
 NOTE: The reflex test must support the specimen type used in the initial test.
4 The following information can be specified for the reflex test.

• Result (Positive or Negative)
• Reflex Assay (List of active assays)
• Reflex Result Name (List of Result Names associated with the selected assay)
5 To add additional reflex settings, select Add and repeat the steps above.
To remove a reflex test, select Remove.
6 Select Next to proceed to Assay Editor Localization Settings.
Assay Editor Localization Settings

The Assay Editor Localization Settings allow you to enter localized text strings for the entire assay or for
individual targets (either the Sample Process Control or the specified Target) by assigned result and locale. This
is an optional setting.

To set a localized string for an individual target
1 Select the Target tab. Or select Next to continue without defining localization settings.
2 Select Add to add a localized string.
3 For each localized string:

• Choose a Target (the Sample Process Control or a target defined in the ADF), or select <Assay> to enter a
localized string for the entire assay.
• Choose an Assigned Result.
– For a target set to the Sample Process Control or a target defined in the ADF, select Target Unresolved,
Target Indeterminate, Target Amplified, Target Not Amplified, Target Aborted, N/A, or No Result.
– For a target set to <Assay>, select Unresolved, Indeterminate, Negative, Positive, Valid, Invalid,
Aborted, N/A, or No Result.
• Enter a Locale (1 to 5 characters).

• Enter the desired Result String (1 to 15 characters).
4 If desired, select Add to add another localized string. Populate the fields as in the steps above.
5 Select Next to proceed to the Assay Editor Summary.
To set a localized string for the entire assay

1 Select the Assay tab.
2 Enter a localized string for the default description.
3 Enter a localized string for the default disclaimer.
4 Select Add to add specific localized strings based on locale.

5 For each localized string:
• Enter a Locale (1 to 5 characters).
• Enter a Description.
• Enter a Disclaimer.
6 Select Next to proceed to the Assay Editor Summary screen.
Assay Editor Summary and Export

The Assay Editor Summary screen displays a summary of the newly created assay. From this screen, the assay can
be exported, making it available for use.
1 Select:
• Export to export the assay as an ADF and make it available for use within NeuMoDx software.
The software will prompt you for the desired network or USB path.
• Select Export Only to export the assay.
The exported assay will not be available for use within the software. To make this assay available for use at
a later time, import the assay from the Assay tab in the Settings tab (see “Assay Settings” on page 94).

Creating an LDT Assay from an Existing Template
You can use an existing LDT as a starting point to create a new LDT.
1 To access Assay settings, select Tools, then Assay.
The Assay Editor Wizard is displayed.
2 Select Create From Template.
3 Select the assay you wish to use as a template from the list of available assays on the left.
• Active to see only active assays.
• Current to see only current assays; assays that have not been archived (both active and inactive).
• Archive to see only archived assays.
4 Select Next to continue.
5 Using Back and Next, navigate through the settings menus described in the previous section, “Defining an
Assay Definition File (ADF)” on page 13.
6 Change the desired parameters and Export the assay as described in “Assay Editor Summary and Export” on
page 46. Fields containing parameters that differ from the previous settings are highlighted in teal.
If created from an active assay, the newly created assay will become the active assay.

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Chapter 4
Creating a Test Order
Creating a Test Order

There are four ways to create a test order:
• Manually create a test order for individual samples from the Test Status Display (see “Manually Creating a Test
Order from the Test Status Display” below)
• Manually create a test order for individual samples from the Loaded Specimen Carrier Display (see “Manually
Creating a Test Order from the Loaded Specimen Carrier Display” on page 52)
• Create a test order by importing a test order file in Excel format (see “Creating a Test Order by Importing a
Test Order File in Excel Format” on page 57)
• Download a test order from a connected Laboratory Information System (LIS) [see “Downloading a Test Order
File from a Laboratory Information System (LIS)” on page 59]
Manually Creating a Test Order from the Test Status Display

Follow the steps below to manually create a test order for individual specimens.
1 Select Test Status, then Pending.
2 Select Create.
PN: 40600250 Rev D Creating a Test Order 49

The Enter Test Orders window opens.
3 Fill in the following fields:

• Specimen ID
• Patient ID (Optional)
• Sample Specimen Type
• Specimen Tube Type
• Specimen Tube Size
• Assay
• Result Name
• Test Specimen Type
For the Specimen ID and Patient ID, use the touchscreen, an optional keyboard, the handheld barcode
scanner, or any combination of the three methods.
The defined Sample Specimen Type determines which assay(s) can be selected. The defined Assay determines
which Result Name and Test Specimen Type can be selected.
The default Specimen Tube Type is the 13 x 75 mm secondary tube, following the guidelines for tube
dimensions and minimum fill volumes for the 32-position, 24-position, and low-volume Specimen Tube
Carriers. If a Specimen Tube Type is selected, you must select a Specimen Tube Size.

The Specimen Tube Type can also be defined when manually creating a test order from the Specimen Tube
Carrier details screen (by selecting Set Tubes), when importing a test order file in Excel format, or when
downloading a test order from the LIS. See the following sections for more information.
4 Select Add to add the test order to the specified test.

Check the box next to STAT if you want the system to prioritize the test.
• To remove a defined test order, select the test order and select Remove.
• To add another test order, select another assay from the Available Assays and Available Result Names and
select Add.
5 Select:
• Save & New to save the current test order and clear the fields to enter an additional test order.
• Save & Close to save the current test order and close the window.

The created test order is displayed in the Pending Tab and used to instruct the system to process the test
once a loaded specimen tube has the matching specimen ID barcode.
Manually Creating a Test Order from the Loaded Specimen

Carrier Display
The Specimen Tubes screen can also be used to define a test order for specimen tubes.
1 From the home screen, select the Specimen Tubes bar.
2 Alternatively, from any other screen in the Home tab, select the Specimen Tubes tab.
3 Load the specimen tube carrier by tapping the up arrow under the desired carrier.

4 If a test is not already assigned by a pending test order or Default Test (see “Assay Settings” on page 94), the
carrier will show a red error state and the Specimen Tube Details Screen is displayed.
If the Details screen is not displayed automatically, select the loaded specimen tube carrier to access the
Details screen.

 NOTE: You can select Set Tubes from the Carrier Details screen to define the tube types for all the
specimens in that carrier.
5 Selecting the red exclamation point icon on the right side of the touchscreen displays more information. In
this case, it displays “No test order assigned,” meaning the specimen ID doesn't have an associated pending
test order.
6 Select a specimen ID in the list to enter a test order for that specimen.

The Edit Specimen screen is displayed.
7 (Optional) Enter a Patient ID.
8 Select the Sample Specimen Type, Specimen Tube Type, and Specimen Tube Size.
9 Select a desired assay and result name from the Assay and Result Name menus.
The available assays and result names will vary based on the selected sample specimen type. The defined
Assay determines which Result Name and Test Specimen Type can be selected.
10 Select Add Test Order.

The new test order is displayed in the Test Order(s) box below.
11 Once the test order is created, the test can be checked STAT, if desired.
If STAT is checked, the specimen will be prioritized for processing over other loaded specimen tubes on the
worktable.
12 Enter Specimen Comments, if desired.
13 To delete the test order, tap the X under the Cancel column.
 NOTE: A test order can be deleted when a test is running if it does not need to be completed. If the test is
in any step other than PCR, the step will finish before aborting and no more steps will be completed with the
sample. If the test is in the PCR step, the sample will abort immediately.
14 Select Apply to save the test order and exit to the Specimen Tubes Carrier Details screen. Or, select Cancel to
exit the Edit Specimen window without adding the test order.
15 Repeat until all specimen tubes in the carrier have an assigned test order.

Creating a Test Order by Importing a Test Order File in Excel Format
This feature imports the data from a test order file into the software as a pending test. The format of the file is
Excel(.xlsx).
 NOTE: It is not possible to create the test order on the NeuMoDx Molecular System; an external computer
must be used.
The test order consists of Specimen ID, Assay (or test to be performed), Specimen Type, and Specimen Tube Type
of the sample. If a specimen tube type is not defined, the secondary tube type will be used as default.
If “Allow Duplicate Test Orders in Import File” is not selected in the Workflow settings (General Settings tab) and
duplicate test orders are present in the test order file, then the first test order will be imported but all other
duplicate test orders will be ignored.
Column Name Description Data Type
Specimen ID The barcode for the specimen tube to be used Required (maximum of 20 characters)
for the test order
Result Code Result Code of the test to be run (e.g., GBS) Required (if default assay is selected)
Specimen Type Type of specimen (e.g., transport media) Optional
Patient ID Unique ID assigned to the patient Optional
Comment Any comments about the sample Optional
Specimen Tube Type Type of specimen tube* Optional
* The Specimen Tube Type Excel Code indicates the type and size. See the following table for Excel Codes for
available tube types and sizes.
The Excel codes for the Specimen Tube Types are defined in the following table.
Specimen Tube Type Excel Code
13 x 75 mm PPS13x75
Plasma/Serum Tube 13 x 100 mm PPS13x100
16 x 100 mm PPS16x100
13 x 75 mm PPTSST13x75
BD PPT™/SST™ Tube 13 x 100 mm PPTSST13x100
16 x 100 mm PPTSST16x100
13 x 75 mm WBT13x75
Whole Blood Tube 13 x 100 mm WBT13x100
16 x 100 mm WBT16x100

Specimen Tube Type Excel Code
13 x 75 mm SDT13x75
Secondary Tube 13 x 100 mm SDT13x100
16 x 100 mm SDT16x100
16x100 mm UTM3
Transport Medium
12x80 mm UTM1
16x100 mm SIT3
Swab in Transport Medium
12x80 mm SIT1
Low Volume Tube LVT1
Creating the Test Order Excel File

1 Using Excel, open the test order template (e.g., TestOrders.xlsx)
2 Fill in the template.
3 Save the file in Excel Worksheet file format by using the .xlsx file extension. The file should be saved to a
USB drive or network folder.
Importing the Test Order File

2 Select Import.
3 The Select Test Order Import File window opens. Navigate to the correct test order file (.xlsx format only),
and select OK.

4 A dialog box opens, showing the status of the import. Select OK.
The imported specimen IDs are displayed in the Pending tab.

 NOTE: The first row of the file is header information and is ignored during file import. If not populated,
optional fields will use default values.
Downloading a Test Order File from a Laboratory Information

System (LIS)
When configured for bi-directional communication, the software can be used to initiate requests to download
open test orders from the LIS host for currently active assays.
 NOTE: When loading a specimen tube, if no test order is found and if LIS is configured, the software will
automatically perform a host query for the given specimen ID.
The following instructions describe how to manually query the LIS host for test orders.
2 Select Download.
All available test orders will begin downloading from the LIS.
The imported specimen IDs are displayed in the Pending tab.


Chapter 5
Running an LDT
Loading LDT Master Mix Test Strips

The NeuMoDx LDT Master Mix test strips are filled with NeuMoDx primers and probes for the sample process
control (SPC1 for DNA Master Mix or SPC2 for RNA Master Mix).
 NOTE: Master Mix test strips are not needed for Extraction Only testing.
Loading and Unloading LDT Master Mix Test Strips

Follow the instructions below to load and unload the LDT Master Mix test strips on the system worktable.
1 From the Home screen, select the Test Strips/Buffers bar.
2 Alternatively, from any other screen in the Home tab, select the Test Strips/Buffers tab.
PN: 40600250 Rev D Running an LDT 61

3 Unload (eject) the carrier by tapping the down arrow on the touchscreen under the desired carrier.
4 Populate the carrier with new Master Mix test strips (if necessary).
5 To load the carrier:

• Set the carrier on the autoloader tray in line with the desired position.
• Gently push the carrier forward towards the system until it reaches a hard stop.
• Tap the up arrow under the desired carrier shown on the touchscreen.
6 Repeat for all carriers until all desired Master Mix test strips are loaded and there are no red error states.
Populating the LDT Primer/Probe Strip with Primers/Probe(s)
Adding Primer/Probe Mix to the LDT Primer/Probe Strip

For LDTs, you will need to prepare a concentrated solution of LDT-specific primers and probe(s). This solution is
referred to as the primer/probe mix.
Guidelines for preparing primer/probe mixes:
• Dilute primers and probes in deionized RNase/DNase-free water, 10 mM Tris pH 8.0, or 1X TE with low EDTA
(0.1 mM EDTA). The final concentration of the primer/probe mix should be 1X after mixing with 20 μL of
eluate in the LDT Primer/Probe Strip.
Example: Add 4 μL of 6X primer/probe mix to the desired wells in the LDT Primer/Probe Strip. Once eluate is
added to the wells, there will be 24 μL at 1X primer/probe mix.

• NeuMoDx recommends adding up to 10 μL of the prepared primer/probe mix per well to the LDT Primer/
Probe Strip.
To add Primer/Probe Mix to the LDT Primer/Probe Strip

1 Obtain LDT Primer/Probe Strips.
2 Using a clean pipette tip, add the desired volume of prepared primer/probe mix to the desired number of
wells in the LDT Primer/Prove Strips.
LDT Primer/Probe Strips are foil covered. You can peel back the foil or puncture it and carefully add the
primer/probe mix to the bottom of the well.
• Avoid introducing air bubbles to the primer/probe mix upon dispensing into the
bottom of the well.
• Only one assay (created in the Assay Editor) may be assigned to each LDT Primer/
Probe Strip.
• Wells must be consumed starting from the bottom left-most well. See the figure
for loading order, from 1 to 16.
• There is no need to fill all wells, but loading order must be from 1 to 16 to avoid
wasting wells on the LDT Primer/Probe Strip.
• These positions will be mapped through NeuMoDx software once the LDT Primer/
Probe Strip is loaded onto the system worktable.
Loading and Unloading LDT Primer/Probe Strips

Follow the instructions below to load and unload the LDT Primer/Probe Strips. Prior to
loading, ensure that the primer/probe mix has been added to the wells of the LDT
Primer/Probe strips and that the strips have been snapped into the test strip carrier.
Use the Test Strips/Buffers screen to load/unload LDT Primer/Probe Strips (along with the Master Mix test
strips).
1 From the Home screen, select the Test Strips/Buffers bar.
2 Alternatively, from any other screen in the Home tab, select the Test Strips/Buffers tab.

3 Unload (eject) the carrier by tapping the down arrow on the touchscreen under the desired carrier.
4 Populate carrier with the LDT Primer/Probe Strips containing the primer/probe mix at the bottom of the
wells.
5 To load the carrier:
• Set the carrier on the autoloader tray in line with desired position.
• Gently push the carrier forward towards the system until it reaches a hard stop.
• Tap the up arrow on the touchscreen under the desired carrier.
6 Repeat for all carriers until all desired LDT Primer/Probe Strips are loaded. The loaded LDT Primer/Probe
Strips will display a yellow status.
A message is displayed asking if you would like to configure the unconfigured test strips.
7 Select Configure to configure the test strips. Or select Cancel to navigate away from the pop-up and
configure the test strips at a later time. Check the box next to “Do Not Notify Again” to temporarily turn off
this notification when inserting LDT Primer/Probe Test Strips.

If you select Configure, you will be taken to the LDT Test Strip Configuration screen. The configuration
selected through this workflow will apply to all test strips listed in the LDT Test Strip Configuration dialog. If
you select Cancel, you will need to select the LDT Primer/Probe Strip carrier to access carrier details and
configure each test strip individually. The yellow status will show that the test strip is not yet configured.
8 Select the yellow [Not Configured] box that shows to configure the LDT Primer/Probe Strip.
Doing so will open the LDT Test Strip Configuration window.

9 Choose the Primer/Probe configuration type. (For information on configuring test strips for extraction only
testing, see Chapter 6, “Extraction Only LDT Testing”)
To configure the LDT Primer/Probe test strips for full-process testing

1 From the LDT Test Strip Configuration window, select Primer/Probe as the Type.

2 Select the appropriate assay from the active LDTs in the Assay menu.
3 Enter the Primer Probe Lot number (1 to 15 alphanumeric characters, including spaces). This is a user-
defined lot number.
4 Enter the Master Mix Lot number (must be exactly 6 alphanumeric characters). This is the lot number
specified on the NeuMoDx RNA Master Mix or DNA Master Mix reagent.
5 Define the well(s) containing the primer/probe mix.

• Select Fill to populate all 16 wells.
• Select + to populate one well at a time.

• Select – to remove a well.
6 Select OK when configuration is complete, or Cancel to go back to the Carrier Details screen without
making any changes.
 NOTE: Once changes are saved, they cannot be modified for that test strip.
7 Repeat for each loaded LDT Primer/Probe Strip.

Specimen Loading
 NOTE: In this section, the term “specimen” includes any of the following: external controls, user-defined
controls, calibrators, and/or patient specimens.
The main steps in specimen loading by an operator include:
• Affix a specimen barcode label to a specimen tube; wrap the barcode label around the tube approximately
1 inch from the top. See the appropriate NeuMoDx System Operator’s Manual for more information on
positioning the barcode.
• If using a secondary tube, transfer the specimen to the barcoded specimen tube. See the appropriate
NeuMoDx System Operator’s Manual for more information on the minimum volume of specimen.
• Load compatible primary specimen tubes or secondary tubes in the specimen tube carriers and remove the
caps if present.
• See the following section for information on loading the carriers.
 NOTE: The system will inform you if there are any issues or samples that need attention prior to test
initialization. If not, specimen processing begins, and you may walk away. If “Manually Confirm Specimen
Carrier Settings” is selected in the Workflow settings (General Settings tab), you must select Continue in the
Specimen Carrier Details tab to proceed with specimen processing (see the Carrier Details for Specimen Tubes
in the appropriate NeuMoDx System Operator’s Manual for more information).
Loading Specimen Tube Carriers Initiates the Test(s)

Follow the instructions below to load and unload specimen tubes from the system worktable.
1 From the home screen, select the Specimen Tubes bar.
2 Alternatively, from any other screen in the Home tab, select the Specimen Tubes tab.
3 Place the specimen tube carrier containing specimens to be processed in any open position on the
autoloader shelf that currently doesn't have a specimen tube carrier on the worktable. Gently slide the
carrier toward the system until it reaches a hard stop.

4 Load the specimen tube carrier by tapping the up arrow under the desired carrier position.
5 Repeat for all specimen tube carriers until all desired specimens are loaded.
6 If “Manually Confirm Specimen Carrier Settings” is selected in the Workflow settings (General Settings tab),
select the carrier, pending confirmation, to verify the test orders and set tubes, if necessary. Then, select
Continue in the Specimen Carrier Details tab to proceed with specimen processing.
 NOTE: Loading a specimen tube carrier will initiate the test unless “Manually Confirm Specimen Carrier
Settings” is selected in the Workflow settings. The system will begin processing the specimens without

additional input. If controls, calibrators, or additional consumables are required to process the test(s), a red
or yellow status bar will appear at the bottom of the screen.
At the Completion of the Run

If still loaded, remove the processed specimen tube(s) from the worktable. If fully consumed or the onboard use
time has expired, remove and dispose of the used lysis buffer containers and test strips according to the
guidelines stated in the respective NeuMoDx System Operator’s Manual. If prompted by the system, remove and
dispose of the Wash Reagent and/or Release Reagent and empty the Priming Waste bottle. Remove and replace
the used tip racks and refill the cartridges, as necessary. If prompted by the system, empty the Biohazard Waste
Container (NeuMoDx 288 System) or the Biohazard Waste Bin and Tip Waste Bin (NeuMoDx 96 System) following
the guidelines in the respective manuals.
Test Status
The Test Status screen allows you to view the status of all in-process (current), completed, and pending tests
(tests that were entered by a test order).
For information on current and pending tests, see the appropriate NeuMoDx System Operator’s Manual.
Completed Tests
The Completed tab allows you to view tests that have completed processing since the last database purge. You
can also export and print Sample Results Reports and Summary Reports from this tab, as well as export and
import raw data.
To access the Completed tab

1 Select Test Status, then Completed.

Filtering Completed Tests
1 Filter the displayed results by selecting Filter, then Filter By from the Test Status, Completed tab.
Options available depend on the user login privileges.
2 Within the Filter By window, select the date range by Date Range Type:
• Today
• Previous Day
• This Week
• Previous Week
• This Month
• Previous Month
• Custom
3 For the Custom Date Range Type, the following can be set:
• Start Filter Date
• Start Filter Time (in hours)
• End Filter Date
• End Filter Time (in hours)
4 In addition to Time/Date filtering, completed tests can be filtered by:

• Specimen ID
• Patient ID
• Assay Name

• Result Name
• Sample Type
• Result
• Standard Curve Name
• Operator
• Test Strip Lot Number
The following filtering options are available only to BioMed users.

• XPCR Module Serial Number
• XPCR Lane
• Extraction Plate Module Serial Number
• Heater Well
5 Enter all desired filter parameters and select OK to continue. Or select Cancel to exit the window without
applying any filters.
Viewing, Exporting, and Printing Sample Results Reports

Sample Results Reports are generated from the Completed tab under Test Status. The View Curves button allows
the viewing of PCR amplification curves. This option is enabled if the Include Graphs option is selected in the
Assay tab under Settings (see “Assay Settings” on page 94). The Export Raw Data button is enabled for LDTs.
The tests that have completed processing are displayed.
2 Check the box(es) next to the desired test(s).
3 Select Report.
4 Select View Report.
5 The Sample Results Report is displayed. If more than one test is selected, the report for the first selected test
in the list is displayed.

Example pages from an LDT Report
6 To navigate within the Sample Results Report screen:

• Use the up/down arrows to advance through the pages of the individual report.
• Use the left/right arrows to advance to the next or previous individual report (if more than one test was
selected from the Completed Tests screen).
• Use the +/– magnifying glass icons to zoom the report in or out.
• Use the Specimen drop-down menu at the top-right corner to select a different individual report (if more
than one test was selected from the Completed Tests screen).
7 Select Export to export the Sample Results Reports.

 NOTE: Export file type, destination, and access level can be configured in the Report tab under Settings
(see the Report Settings section in the appropriate NeuMoDx System Operator’s Manual).
• Sample Results Reports are automatically saved to the specified location if a Default Export Type and
Default Output Path for the Report Export was set in the Report tab under Settings.
• If a Default Export Type was not specified in the Report tab, the Sample Results Report will be saved as a
PDF file.
• If a Default Output Path was not specified in the Report tab, a pop-up window allows you to set a path
(e.g., USB drive).
8 Select Export As to Export the Sample Results Reports and select a File Type from the following list of options:
• CSV files (*.csv)
• PDF files (*.pdf) [Default]
9 Select Export Raw Data to export the Raw Data file.

Raw Data Exports are automatically saved to the specified location if a Default Output Path for the Raw Data
Exports was set in the Report tab under Settings. If a specified Default Output Path was not set in the Report
tab, a pop-up window allows you to set a path (e.g., USB drive).
10 Select Print to Print the displayed report.

Sample Results
The following Overall Results may be displayed on the Sample Results Report for qualitative assays:
• Positive
• Negative
• Indeterminate (IND)
• Unresolved (UNR)
• Aborted
• No Result
The following Overall Results may be displayed on the Sample Results Report for external controls and user-
defined controls used in conjunction with qualitative assays:
• Valid
• Invalid
The following table explains the decision criteria used in the Overall Result call for qualitative assays:
Sample Process Control

Result Target System Events
(SPC1, SPC2)
Positive Amplified N/A No relevant errors

Negative Not Amplified Amplified No relevant errors
Indeterminate Not Amplified Not Amplified Relevant errors
Unresolved Not Amplified Not Amplified No relevant errors
Aborted No Result No Result Sample was aborted by user
No Result No Result No Result Relevant errors
Indeterminate Results are accompanied by relevant flags and may be caused by:
• General system failures or system errors (see the Troubleshooting section of the appropriate NeuMoDx
System Operator’s Manual) that are identified once the sample has completed processing.
• Failure of the PCR region of the cartridge to fill with PCR mix
Unresolved Results may be caused by:

• Test inhibition
• Other processing or system error
Aborted Results are accompanied by relevant flags not accompanied by data and are caused by:
• The user choosing to abort the test
No Results are not accompanied by relevant flags and are be caused by:
• General system failures or system errors (see the Troubleshooting section of the appropriate NeuMoDx
System Operator’s Manual) that occur in the pre-analytical stages of sample processing.

Generally, No Result and Indeterminate results are caused by system errors, and Unresolved Results are
caused by test inhibition.
Valid and Invalid Results are used for external controls and user-defined controls:
• Valid: All of the set conditions for the control have been met.
Positive control: Target is amplified, No relevant system errors
Negative control: Target is not amplified, No relevant system errors
• Invalid: Any of the set conditions for the control have not been met
Positive control: Target is not amplified, or relevant system errors, or both
Negative control: Target is amplified, or relevant system errors, or both
Controls Used (If Applicable)

The following information about the controls used during the run will be displayed on the Sample Results
Report, if applicable:
• Control name
• Barcode
• Lot number
• Completed (date and time)
• Test strip lot used for testing the controls
• Result
Consumables
The following information about consumables used during the run will be displayed on the Sample Results
Report:
• Name
• Lot number
• Shelf life remaining
• In-use time
Viewing, Exporting, and Printing a Summary Report

In addition to viewing, printing, and exporting individual Sample Results Reports, you can generate an overall
Summary Report for a group of test results.
1 Use the Filter By option to select the desired tests. See “Filtering Completed Tests” on page 72.
2 Select Report.

3 Select Summary Report.
The Summary Report for the selected samples is displayed.

Example of Results Summary Report
4 To navigate within the Results Summary Report Screen:

• Use the up/down arrows to advance through the pages of the Summary Report
• Use the +/- magnifying glass icons to zoom the Summary Report in or out.
 NOTE: The filters applied are displayed on the Summary Report (see “Filtering Completed Tests” on
page 72).

5 Select Export to export the Summary Report.
 NOTE: Export file type, destination, and access level can be configured in the Report tab under Settings
(see the Report Settings section in the appropriate NeuMoDx System Operator’s Manual).
• Summary Reports are automatically saved to the specified location if a Default Export Type and Default
Output Path for the reports were set up in the Report tab under Settings.
• If a Default Export Type was not specified in the Report tab, the report will be saved as a PDF file.
(e.g., USB drive).
6 Select Export As to Export the Summary Report and select a File Type (CSV and PDF [Default]).
7 Select Print to print the displayed Summary Report.
Exporting Raw Data

The raw data generated in LDT testing can be exported and analyzed on a separate computer. Raw data files can
be exported using the following method, or by selecting Export Raw Data when exporting the Sample Results
Reports or Summary Report.
To export Raw Data

1 Use the Filter By option to select the desired tests. See “Filtering Completed Tests” on page 72.
2 Select Export, then Raw Data.
3 If applicable, select the file path where you want the raw data to be exported.
 NOTE: If a file path for Raw Data Export is specified in the Report tab under Settings, the file will
automatically be exported to that location.
• Raw Data Exports are automatically saved to the specified location if a Default Output Path for Raw Data
Export was set in the Report tab under Settings.

(e.g., USB drive).
Re-Analyzing Raw Data

When optimizing LDT data processing conditions, it may be beneficial to reprocess data under different
parameters, such as Minimum Peak Height and EPR Check Ct Threshold. Using the NeuMoDx System, exported
raw data can be imported and analyzed using a different version of the original ADF.
 NOTE: Re-analyzing data using this method will overwrite the original data. Ensure that you have saved
your data in a trusted location on a USB or separate computer prior to reprocessing.
To re-analyze raw data

1 Select Tools, then Assay.
2 Select Create From Template and choose the ADF that was originally used to process the samples.
3 Select Next.
4 Navigate to the Assay Editor PCR Target Settings screen and make the desired processing parameter changes.
5 Select Next to navigate through the remaining ADF screens. If desired, changes can be made to the Assay tab
in the Assay Editor Localization Settings.
6 Select Export to export the ADF.
7 Select Settings, then Assay to navigate to the Active Only table. Confirm that the new ADF is selected as
Active. Activate the new ADF version if necessary.
8 Select Test Status, then Completed to navigate to the completed tests.

9 Select Import and use the file explorer to navigate to the folder containing the raw data to be reprocessed.
A message informs you that the import was successful.
10 Select Filter By and enter the filter criteria necessary to find your reprocessed samples. Select OK to accept
the criteria and return to the Completed Tests screen.
11 Select Export, then Raw Data to export the reprocessed raw data.
• Raw Data Exports are automatically saved to the specified location if a Default Output Path for Raw Data
Export was set in the Report tab under Settings.
(e.g., USB drive).

Chapter 6
Extraction Only LDT Testing
This chapter outlines the special use case of Extraction Only testing. Extraction Only testing is only available for
use with Research Use Only (RUO) LDT ADFs.
Extraction Only LDT Testing

Extraction Only testing begins with the aspiration of lysis buffer and sample at LHPA but ends the process at
LHPC after the pipette dispenses the entire volume of the eluate into the assigned LDT Primer/Probe Strip well.
Each well that was assigned a sample will contain the user-defined amount of eluate containing extracted DNA
or RNA. The extracted eluate can then be used for further analysis outside of the NeuMoDx Molecular System.
Extraction Only Workflow

EXTRACTION ONLY
IMPLEMENTATION
Define ADF in Assay Editor
Set ADF to Extraction Only in

Assay Settings tab
LOAD GENERIC CONSUMABLES

x NeuMoDx Cartridges
x NeuMoDx Extraction Plates
LOAD TEST SPECIFIC REAGENTS
x 100uL Tips
x NeuMoDx Lysis Buffer
x 300uL Tips
x LDT Primer/Probe Strip
(Extraction Only Test Strip)
Configure
Extraction Only
Test Strip
PROCESS SAMPLES
Unload Eluates in Extraction Only

Test Strip from System Worktable
PN: 40600250 Rev D Extraction Only LDT Testing 81

Extraction Only LDT Implementation Sample Processing Overview
For extraction only LDT implementation, LHPC refers to the process of aspirating the eluted nucleic acid from the
cartridge and dispensing it into an empty LDT Primer/Probe test strip. In this process, the LHR:
• Picks up new 300 μL tips for each discrete eluted nucleic acid sample
• Punctures appropriate foil-covered empty primer/probe test strip wells
• Moves to the appropriate PCR injection port (referred to as the P-port) on the cartridge, and using
independent z-axis control, aspirates all of the eluate (combined with the XPCR Module syringe pump
pushing air)
• Moves to the LDT Primer/Probe Strip and dispenses the entire volume of the eluate into the empty well
 NOTE: Once tips have been used for aspiration of the eluted material, the Liquid Handling Robot does not
Setting Up an ADF for Extraction Only Testing

1 Follow the steps in Chapter 3, “Defining an Assay Definition File (ADF)” to set the parameters for your
Extraction Only test. Ensure that RUO is selected for the ADF Classification in the Assay Editor General
Settings.
2 After defining your ADF, navigate to the next set of settings by selecting Settings and then Assay.

3 Select Edit next to your RUO LDT ADF. The Active Test Settings for that ADF appears.
4 Select Extraction Only to enable this setting for the selected ADF. Select the amount of extraction only
release volume in μL from the available options.
5 Select OK to accept the new settings. Or select Cancel to close the window without saving your settings
changes.

The configured Extraction Only settings appear in the “Enabled Features” column of the Test table.
Configuring an Extraction Only Test Strip

Extraction Only–configured test strips are not configured to a specific Assay Name or Test Name and can be used
for multiple Extraction Only assays simultaneously. It is recommended that test strips be placed on separate
carriers to ensure efficient unloading of the eluates.
1 Insert test strip carriers and select the LDT test strips as explained in the “Loading and Unloading LDT Primer/
Probe Strips” on page 63.
 NOTE: The addition of primer/probe mix to the LDT Primer/Probe test strips is not required prior to
insertion into the system and may be added after extraction has completed.

2 From the LDT Test Strip Configuration window, select Extraction Only as the Type.
3 Select OK to confirm this configuration. Or select Cancel to navigate back to the Carrier Details screen
without making any changes.
The green test strip box displays Extraction Only for the configured test strip.

Initiating and Completing an Extraction Only Test
1 Follow the steps to prepare the specimen tube carriers in “Specimen Loading” on page 69.
2 Define the details for any samples displaying an error.
3 Immediately following the run, a message appear, alerting you that the extraction only test strip is ready to
unload.
4 To export the test strip well mapping to a user-specified file location:

• Select Export PDF to export the test strip well mapping as a PDF file.
• Select Export CSV to export the test strip well mapping as a CSV file.

5 Select the down arrow to eject the carrier.
Select Close if you do not wish to eject the carrier at this time.
 NOTE: It is recommended that the test strip be removed as soon as possible to maintain the integrity of
the eluate.
6 Once the carrier is ejected, immediately remove the test strip from the carrier. Gently pull up on the test strip
so you do not spill any of the eluate in the wells.
7 It is recommended that you use the eluate immediately for further testing. You may cover the strip with foil
and store in the refrigerator or on ice for future use, per your testing requirements. Eluate stability has not
been established.

Completed Extraction Only Tests
To access the Completed tab
The tests that have completed processing are displayed.
• Select Report to export Extraction Only Mappings as PDF files (see “Extraction Only Mapping in PDF File
Format” on page 89).
• Select Export to export Extraction Only Mappings as CSV files (see “Extraction Only Mapping in CSV File
Format” on page 91).

Filtering Completed Tests
Completed Extraction Only tests can be viewed separately from other sample types using the Filter By option.
1 Filter the displayed results by selecting Filter, and then Filter By from the Test Status, Completed tab. Options
available depend on the user login privileges.
2 Select the Date Range Type.
3 Set the Custom Date Range Type, if desired.

The other provided filters can be used to further filter Extraction Only tests.
4 Check the box next to Extraction Only to filter out all other types of tests.
5 Enter all desired filter parameters and select OK to continue. Or select Cancel to exit the window without
applying any filters.
Exporting the Extraction Only Mapping

Extraction Only Mappings can be exported immediately after the samples finish processing, or later from the
Completed Tests tab.
Extraction Only Mapping in PDF File Format

1 Check the boxes next to the desired tests.
2 Select Report.

3 Select Extraction Mapping.
4 If applicable, select the file path where you want the Extraction Mapping file to be exported .
• Extraction Mapping files are automatically saved to the specified location if a Default Output Path for Raw
Data Exports was set in the Report tab under Settings.
(e.g., USB drive).

Extraction Only Mapping in CSV File Format
1 Check the boxes next to the desired tests.
2 Select Export.
3 Select Extraction Mapping.
4 If applicable, select the file path you want the Extraction Mapping to be exported to.
• Extraction Mapping files are automatically saved to the specified location if a Default Output Path for Raw
Data Exports was set in the Report tab under Settings.
(e.g., USB drive).


Chapter 7
Settings
This chapter contains the LDT-specific settings in the Report, Assay, and Controls tabs in Settings. For details on
General, Report, Network, Assay, Controls, Users, and LIS settings, see the appropriate NeuMoDx System
Operator’s Manual.
Report Settings
The Report settings screen allows Supervisor and BioMed-level users to set the default settings for printing and
reporting, as well as default paths for exported data, troubleshooting packages, and screen captures.
To access Report settings

1 Select Settings, then Report.
2 If desired, select Browse for Raw Data Export and navigate to a location to set it as the Default Output Path.
PN: 40600250 Rev D Settings 93

Assay Settings
The Assay tab allows Supervisors and BioMed-level users to set up run and reporting configurations for active
tests.
To access Assay settings

1 Select Settings, then Assay.
A list of assays (IVD and LDT) is displayed.
2 Select:
• Active Only to see only active assays.
• Current to see assays that have not been archived (both active and inactive).
In the Current view, select an assay and then select Deactivate to make the assay inactive. A confirmation
window is displayed.
• Archived to see all assays that have been archived.
 NOTE: Once an assay has been archived, it cannot be recovered.
3 (Optional) Select Import to import an LDT ADF that was exported only (not available for use in the software)
at the Assay Editor Summary screen.
4 For information on standard curves, see “Standard Curves” in the following section.
Standard Curves
NeuMoDx software provides the following functions for standard curves for quantitative assays based on the
user's privilege level:
• Defining a standard curve
• Scanning in a standard curve
• Manually entering a standard curve
• Activating or inactivating a standard curve

 NOTE: For information on scanning in a standard curve and activating or inactivating a standard curve, see
the appropriate NeuMoDx System Operator’s Manual.
To access Standard Curve settings

1 Select Settings, then Assay.
2 Select Standard Curves.
The Standard Curves screen is displayed.
3 Choose the desired test from the drop-down menu.

The available standard curves are listed with the following details for each test:
• Name
• Date
• Expiration (number of days remaining)
• Test strip lot number associated to the standard curve
• Specimen type
• Active status

Defining a Standard Curve
 NOTE: You can only define standard curves for LDTs.
1 Select Define/View Details.
The Define/View Standard Curve screen is displayed.
2 Select optional filters for Lot, Date, or an existing standard curve.

All standards run from the selected conditions will be displayed with the following details for each:
• Name
• Specimen ID
• Date/Time standard was run
• Lot number of test strip used to run the standard
• Concentration of the standard (in Log10)
• Ct value assigned to standard
• Result (Valid or Invalid)
3 To define a standard curve, select the box under the Include column (last) to select each standard to be used
to generate the standard curve.
General guidelines for defining a standard curve:
• At least one replicate from at least four different standards must be chosen to display or create a standard
curve.
• If a standard curve has been manually entered or scanned in using the handheld scanner, no
corresponding standards will be displayed on this screen.
• All selected standards must have the same lot number.
• Invalid standards cannot be included in the standard curve generation.

• NeuMoDx software will automatically generate/regenerate the standard curve upon selection/
deselection of valid standards in the Include column.
• Upon generation/regeneration of a standard curve, the software displays the standard curve equation,
y = mx + b, where m = slope and b = the y intercept value. The R2 value will also be displayed.
4 Once the desired standard curve is displayed, select Create New.
5 Enter a name for the new standard curve (5 to 20 characters, and must be a unique name for the selected
test).
6 Once the standard curve is stored in the system, a standard curve barcode may be printed to the default
printer. Select Print Barcode to print a barcode specific to the selected standard curve.
7 Select Close to exit the Define/View Standard Curve window.

Manually Entering a Standard Curve
When a new standard curve is entered into the system for a given assay and test strip lot number, the previous
standard curve for the same assay (ADF) and lot number will be deactivated, if present. The new standard curve
will become active.
1 Select Enter New from the Standard Curves screen to enter a new standard curve for the selected assay.
The Enter a New Standard Curve window is displayed.
2 To enter a new standard curve, either scan the standard curve barcode using the hand-held scanner, or
manually fill in the displayed fields:
• Name (5 to 20 characters)
• Primer Probe Lot # (user-defined)
• Master Mix Lot # (exactly 6 characters; refers to the NeuMoDx Master Mix Test Strip lot number)
• Date
• Slope (value between -4 and -2 with a maximum of 4 decimal places)
• Y Intercept: Acceptable range is dependent on the number of cycles for the test with a maximum of
4 decimal places
• R2 (between 0.9 and 1 with a maximum of 4 decimal places)
3 Select OK to save the new standard curve, or Cancel to return to the previous screen.

Controls Settings
The Controls settings tab allows you to create user-defined controls. See the appropriate NeuMoDx System
Operator’s Manual for details on creating user-defined controls.
Creating User-Defined Controls

 NOTE: A User-Defined Control cannot be deleted or edited once changes have been made.
1 Select Settings, then Controls.
2 Choose an assay from the Select Assay drop-down menu to define User-Defined Controls.
The User-Defined Controls Settings window is displayed showing only active assays.
3 Select Add to define a user-defined control.

The Specify User-Defined Control window is displayed.
4 Enter the Control Name for the user-defined control (1 to 20 characters).
5 Choose specimen type(s) supported for the selected assay.
6 Select an expected Result for the specimen type.

• Target Not Amplified
• Target Amplified
7 For quantitative user-defined controls you may assign a specific concentration value.
Check this if you want an expected concentration of target to be defined. If not selected, the external control
will be treated as a qualitative control (amplified or not amplified, no specific quantitation required). If this
option is selected, the following three options must be defined:
• Enter the Concentration in Log10.
• Enter the Concentration Lower and Upper Limit in Log10.
8 Select Apply to save the user-defined control. Or select Cancel to exit without saving.
 NOTE: A User-Defined Control cannot be deleted or edited once changes have been made.
The user-defined control is displayed.
To view the controls by lot number, check the box next to View By Lot. To view the controls by time until
due, uncheck the View By Lot box. To show only active user-defined controls for the selected assay, check
the box next to Show Active Only. To view active and inactive user-defined controls, uncheck the Show
Active Only box.
9 If desired, select the user-defined control settings:

• Require Lot Frequency: the system will prompt for controls to be run if a new lot of test strips is used.
• Require Run Frequency: enter the number of days that controls must be run.
10 Select Apply to save changes.
Control Mapping
This feature allows you to map additional specimen ID barcodes to standards and external controls and to define
additional standards and user-defined controls. This can be done by selecting a previously made control or by
importing the mappings as defined in a file. The format of the file is an Excel Workbook with multiple worksheets
(.xlsx).
See the appropriate NeuMoDx System Operator’s Manual for information on creating a control mapping file using
Excel and importing a control mapping file.
 NOTE: It is not possible to create the test order on the NeuMoDx Molecular System; an external computer
must be used.

Mapping User-Defined Controls
1 To add User-Defined Control Sample Mapping(s) for each User-Defined control, select the control you want to
edit and select Edit.
The Specify User-Defined Control screen displays the selected assay.
2 Select Add.
The Create User-Defined Control Mapping screen appears.
3 Enter a Specimen ID and choose the Specimen Type.
4 Click Save to save the Control Mapping. Or click Cancel to close the window without saving.
The User-Defined Control Sample Mapping appears in the table.
The User-Defined Control Sample Mapping appears next to the User-Defined Control.

Specifications for the Control Mapping Excel File
Map Standards Worksheet
The Map Standards worksheet consists of a specimen ID, assay name (for the assay to be performed), control
name (name of existing standard being mapped), and specimen type.
Specimen ID The specimen barcode that the Required (maximum of 20 characters)

standard will be mapped to
Assay Name The name of the assigned assay Required
Control Name The name of the control Required (maximum of 20 characters);
must match an existing standard name
Specimen Type The type of specimen this control Optional; must be blank or one of the
applies to following: Blood, Urine, Transport-
Medium, Serum, Plasma, CSF
Default Specimen will be used if blank.
Define Standards Worksheet

The Define Standards worksheet consists of a sample (specimen) ID, assay name (for the assay to be performed),
new control name (unused name of the new standard), expected concentration, replicates, and specimen type.
Specimen ID The specimen barcode for the standard Required (maximum of 20 characters)
New Control Name The name of the standard Required (maximum of 20 characters);
must be a new standard name
Expected Concentration The concentration of the new standard Required
Replicates The number of replicates assigned to Required
the standard
Specimen Type The type of specimen this standard Required; must be blank or one of the
applies to following: Blood, Urine, Transport-
Medium, Serum, Plasma, CSF
Map User-Defined Controls Worksheet
The Map User-Defined Controls worksheet consists of a specimen ID, assay name (for the assay to be performed),
control name (name of the user-defined control), and specimen type (required for quantitative assays).
Specimen ID The sample barcode for the user- Required (maximum of 20 characters)
defined control
Control Name The name of the control Required (maximum of 20 characters)
Specimen Type The type of specimen this standard Optional; must be blank or one of the
applies to following: Blood, Urine,
TransportMedium, Serum, Plasma, CSF
 NOTE: When defining new user-defined controls, a minimum of one positive and one negative control is
required.

Chapter 8
Troubleshooting
This chapter contains LDT-specific errors and flags reported by the NeuMoDx System. For more troubleshooting
information, see the appropriate NeuMoDx Operator’s Manual.
Flags
Software Flags
Software flags indicate events that occurred during test setup and sample processing. These flags will appear as
both alerts in the System Events Report and flags on the individual Sample Results Reports and Summary
Report.
Flag Code Flag Severity Description
17 Re-analyzed using raw data Informational Re-analyzed using raw data

18 Test Result was calculated with a user Informational Test Result was calculated with a user
modified Ct value modified Ct value
20 Extraction Only Informational Extraction Only
2029 LDT was processed with a Primer Error LDT was processed with a Primer Probe/
Probe/Master Mix lot mismatch Master Mix lot mismatch
Results Processing Flags

Results processing flags are tracked per test to indicate events that occurred during results processing. These
flags will appear on the individual Sample Results Reports and the Summary Report.
Flag Code Flag Severity Description
1032 Modified Ct Threshold Informational Modified Ct Threshold
PN: 40600250 Rev D Troubleshooting 107

Index
A external controls, mapping 101

acronyms 7 extraction only
adding primer/probe mix to LDT strips 62 assay definition file (ADF) 82
completed tests 88
assay definition file (ADF)
configuring test strips 84
defining 13–46
exporting mapping 88, 89
exporting 46
sample processing overview 82
extraction only 82
testing 81–87
importing 94
assay editor settings G
See LDT assay editor
general assay editor settings 13
assay settings 94
I
C
importing
calibrator settings, quantitative assays 38 LDT ADF 94
carriers raw data 79
loading specimen tubes 69 test order file 58
control mapping 101
specifications 104 L
controls LDT
creating user-defined 99 creating from template 47
external 34, 40 defining ADF 13–46
settings 99 extraction only testing 81–87
creating importing ADF 94
LDT from template 47 loading strips 63
test order 49–56 master mix test strips 61
user-defined controls 99 primer/probes 62
sample processing overview 10
D standard curves 94–98
definitions 7 workflow 9
LDT assay editor
E calibrator setting 38
exporting external control settings 40
ADF 46 general settings 13
extraction only mapping 88, 89 localization settings 43
LDT 46 PCR steps 20
Sample Results Reports 74 PCR target settings 24
Summary Reports 78 PCR targets 23
external control settings qualitative external control settings 34
qualitative assays 34 reflex settings 42
qualitative/quantitative assays 40 result codes settings 29
quantitative assays 40 specimen settings 17
PN: 40600250 Rev D 109

standard curve settings 35 exporting Sample Results Reports 74
standard settings 37 settings 93
summary and export 46 Summary 76–78
liquid handling process 10–12 result codes settings 29
loading qualitative assays 31
LDT Primer/Probe strips 63 qualitative/quantitative assays 33
master mix test strips 61 results, sample 75
specimen carriers 69
localization settings 43 S
lysis/binding 11 sample
results 75
M sample processing overview
master mix test strips, loading 61 extraction only tests 82
LDT 10
O Sample Results Report
overview exporting 74
sample processing 10 settings
workflow 9 assay 94
controls 99
P report 93
PCR steps 20 See also LDT assay editor
PCR target settings 24 specimen carriers, loading 69
PCR targets 23 specimen settings 17
primer/probe mix 62 standard curve settings, quantitative assays 35
standard curves 94–98
Q standard settings, quantitative assays 37
qualitative assays Summary Report 76–78
external control settings 34 exporting 78
qualitative/quantitative assays
calibrator settings 38 T
external control settings 40 technical support 8
standard curve settings 35 test order
standards settings 37 creating, importing 58
quantitative assays creating, importing CSV format 57
calibrator settings 38 creating, loaded specimen carrier 52
external control settings 40 creating, manually 49
standard curve settings 35 downloading from LIS 59
standards settings 37 test status
completed tests 71
R completed, extraction only 88
raw data pending tests 49
importing 79 test strips
re-analyzing 79 adding primer/probe mix 62
real-time PCR 12 configuring extraction only 84
reflex settings 42 loading 63
reports 73–74 master mix 61
tests
Sample Results Reports 73–74
troubleshooting 107
U
user-defined controls
creating 99
mapping 101
W
workflow overview 9
X
XPCR extraction 11
PN: 40600250 Rev D 111

Product Ordering and Company Contact Information
1250 Eisenhower Place
Ann Arbor, MI 48108 USA
Tel: 1-888-301-6639
Emergo Europe
Prinsessegracht 20
2514 AP The Hague
The Netherlands
Tel: (31) (0) 70 345-8570
PN 40600250 Rev D
© 2020, NeuMoDx Molecular, Inc. All rights reserved. SN:___________________

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NeuMoDx™ Laboratory Developed

Test (LDT) Supplement

© 2020, NeuMoDx Molecular, Inc. All rights reserved. PN 40600250 Rev D

PN: 40600250 Rev D

About This Supplement . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 5

Chapter 3: Defining an Assay Definition File (ADF) . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 13

PN: 40600250 Rev D iii

Chapter 5: Running an LDT . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 61

Chapter 6: Extraction Only LDT Testing . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 81

Chapter 8: Troubleshooting . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .107

iv NeuMoDx Laboratory Developed Test (LDT) Supplement PN: 40600250 Rev D

PN: 40600250 Rev D 5

6 NeuMoDx Laboratory Developed Test (LDT) Supplement PN: 40600250 Rev D

ADF Assay Definition File

PN: 40600250 Rev D Introduction 7

8 NeuMoDx Laboratory Developed Test (LDT) Supplement PN: 40600250 Rev D

QUALITATIVE LDT QUANTITATIVE LDT

Define ADF in ƐƐĂǇ Editor Define Initial ADF [Qualitative]

LOAD GENERIC CONSUMABLES LOAD GENERIC CONSUMABLES LOAD TEST SPECIFIC

Controls and Calibrators YES

PROCESS ROUTINE SAMPLES

PN: 40600250 Rev D Overview 9

No. Step Description

Liquid Handling Process A (LHPA)

10 NeuMoDx Laboratory Developed Test (LDT) Supplement PN: 40600250 Rev D

Liquid Handling Process B (LHPB)

Liquid Handling Process C (LHPC)

PN: 40600250 Rev D Overview 11

12 NeuMoDx Laboratory Developed Test (LDT) Supplement PN: 40600250 Rev D

Defining an Assay Definition File (ADF)

Assay Editor General Settings

PN: 40600250 Rev D Defining an Assay Definition File (ADF) 13

3 Select the Result Type (Qualitative, Quantitative, or Qualitative/Quantitative).

4 Select a Chemistry Type (RNA or DNA).

6 Enter an Assay Display Name (1 to 30 characters).

7 Enter an Assay Name (1 to 15 characters).

14 NeuMoDx Laboratory Developed Test (LDT) Supplement PN: 40600250 Rev D

9 The Release Date is displayed.

11 (Optional) Enter a barcode.

13 Scroll down to see more General Settings

14 The number of PCR Mixes is set to 1. This cannot be modified.

15 RP (Results Processing) Version is set to V2. This value cannot be changed.

16 Select a Result Processing Algorithm.

17 Choose a desired Ct Calling Algorithm.

PN: 40600250 Rev D Defining an Assay Definition File (ADF) 15

18 Select Starting Fluorescence Start and End Cycles.

General guidelines for Starting Fluorescence Start and End Cycles:

19 Select End Point Fluorescence Start and End Cycles.

20 Enter a time (in hours) for the Open-Life.

16 NeuMoDx Laboratory Developed Test (LDT) Supplement PN: 40600250 Rev D

22 Enter a Fill Check Threshold.

23 Select Next to proceed to the Assay Editor Specimen Settings.

Assay Editor Specimen Settings

PN: 40600250 Rev D Defining an Assay Definition File (ADF) 17

2 Select a Specimen Type.

4 Enter the onboard Specimen Stability time (1 to 48 hours).

6 Select the Specimen Aspirate Volume.

18 NeuMoDx Laboratory Developed Test (LDT) Supplement PN: 40600250 Rev D

8 Enter the Rerun Specimen Mix Volume (5 to 900 μL).

9 Enter the Specimen Mix Count (0 to 10).

10 Enter the Rerun Specimen Mix Count (0 to 10).

11 Scroll down to see more Specimen Settings.

12 Select the Rerun Specimen Aspirate Volume.