The LIAISON NES FLU A/B, RSV & COVID-19 assay is a real-time RT-PCR assay intended for use on the LIAISON NES instrument for the simultaneous in vitro qualitative detection and differentiation of nucleic acid from severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), influenza A (Flu A) virus, influenza B (Flu B) virus, and respiratory syncytial virus (RSV) in anterior nasal swab specimens from individuals with signs and symptoms of respiratory tract infection. Clinical signs and symptoms of respiratory tract infection due to SARS-CoV-2, influenza A, influenza B, and RSV can be similar.

The LIAISON NES FLU A/B, RSV & COVID-19 assay is intended for use as an aid in the differential diagnosis of SARS-CoV-2, influenza A, influenza B, and RSV infections when used in conjunction with other clinical and epidemiological information, and laboratory findings. SARS-CoV-2, influenza A, influenza B, and RSV viral RNA are generally detectable in anterior nasal swab specimens during the acute phase of infection. This test is not intended to detect influenza C virus infections.

Positive results are indicative of the presence of the identified virus, but do not rule out bacterial infection or co-infection with other pathogens not detected by the test. The agent(s) detected by the LIAISON NES FLU A/B, RSV & COVID-19 assay may not be the definite cause of the disease. Negative results do not preclude SARS-CoV-2, influenza A, influenza B or RSV infection and should not be used as the sole basis for patient management decisions.

Device Description

The LIAISON NES Instrument (NES1001) is capable of analysis of a single cartridge containing a single specimen. A set of parameters specific to the assay is included in the instrument software to name target molecules, assign dyes to probes, specify cycling conditions, and to analyze data from runs. Fluorescence intensity is monitored at each PCR cycle by detection modules within the instrument. The instrument software controls the thermocycling and, upon completion of the run, automatically interprets and displays results for the specimen.

The LIAISON NES Instrument is comprised of the following:

Touchscreen User Interface
Status LED Indicator
Audio Speaker
Barcode Scanner

The LIAISON NES software is a graphical user interface (GUI) application that is the end-user interface to the LIAISON NES Instrument. The software is installed in an embedded computer. The LIAISON NES software is responsible for providing the environment in which a user runs assays and obtains results.

The LIAISON NES instrument is intended to accept a cartridge, containing either a quality control (QC) or patient sample, to process and detect for the target nucleic acid.

The LIAISON NES FLU A/B, RSV & COVID-19 assay used on the LIAISON NES instrument is a real-time RT-PCR system that enables the direct amplification, detection, and differentiation of influenza A viral RNA, influenza B viral RNA, RSV RNA, and SARS-CoV-2 RNA directly from nasal swabs. Nasal swabs can be professionally collected by a healthcare provider or self-collected under the healthcare provider's supervision.

The LIAISON NES FLU A/B, RSV & COVID-19 assay consists of the LIAISON NES instrument, the LIAISON NES FLU A/B, RSV & COVID-19 Cartridge containing all the required PCR reagents, the NES Sample Vial containing sample release buffer, and the NES Swab for sample collection.

In the LIAISON NES FLU A/B, RSV & COVID-19 assay, fluorescent probes are used with corresponding forward and reverse primers to amplify influenza A, influenza B, RSV, and SARS-CoV-2, and internal control (IC) RNA. Conserved regions of influenza A viral RNA, influenza B viral RNA, RSV RNA, and SARS-CoV-2 RNA are targeted to identify the viruses in the specimen, while the internal control (IC) RNA is used to detect any PCR failures and/or inhibition.

AI/ML Overview

N/A

Summary

U.S. Food & Drug Administration 510(k) Clearance Letter

Page 1

U.S. Food & Drug Administration
10903 New Hampshire Avenue
Silver Spring, MD 20993
www.fda.gov

Doc ID # 04017.08.02

December 23, 2025

Diasorin Molecular LLC
Sarah Baumann
Principal Regulatory Affairs Specialist
11331 Valley View Street
Cypress, California 90630

Re: K251978
Trade/Device Name: LIAISON NES FLU A/B, RSV & COVID-19
Regulation Number: 21 CFR 866.3981
Regulation Name: Device To Detect And Identify Nucleic Acid Targets In Respiratory Specimens From Microbial Agents That Cause The SARS-Cov-2 Respiratory Infection And Other Microbial Agents When In A Multi-Target Test
Regulatory Class: Class II
Product Code: QOF
Dated: June 27, 2025
Received: June 27, 2025

Dear Sarah Baumann:

We have reviewed your section 510(k) premarket notification of intent to market the device referenced above and have determined the device is substantially equivalent (for the indications for use stated in the enclosure) to legally marketed predicate devices marketed in interstate commerce prior to May 28, 1976, the enactment date of the Medical Device Amendments, or to devices that have been reclassified in accordance with the provisions of the Federal Food, Drug, and Cosmetic Act (the Act) that do not require approval of a premarket approval application (PMA). You may, therefore, market the device, subject to the general controls provisions of the Act. Although this letter refers to your product as a device, please be aware that some cleared products may instead be combination products. The 510(k) Premarket Notification Database available at https://www.accessdata.fda.gov/scripts/cdrh/cfdocs/cfpmn/pmn.cfm identifies combination product submissions. The general controls provisions of the Act include requirements for annual registration, listing of devices, good manufacturing practice, labeling, and prohibitions against misbranding and adulteration. Please note: CDRH does not evaluate information related to contract liability warranties. We remind you, however, that device labeling must be truthful and not misleading.

If your device is classified (see above) into either class II (Special Controls) or class III (PMA), it may be subject to additional controls. Existing major regulations affecting your device can be found in the Code of Federal Regulations, Title 21, Parts 800 to 898. In addition, FDA may publish further announcements concerning your device in the Federal Register.

Additional information about changes that may require a new premarket notification are provided in the FDA guidance documents entitled "Deciding When to Submit a 510(k) for a Change to an Existing Device"

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K251978 - Sarah Baumann Page 2

(https://www.fda.gov/media/99812/download) and "Deciding When to Submit a 510(k) for a Software Change to an Existing Device" (https://www.fda.gov/media/99785/download).

Your device is also subject to, among other requirements, the Quality System (QS) regulation (21 CFR Part 820), which includes, but is not limited to, 21 CFR 820.30, Design controls; 21 CFR 820.90, Nonconforming product; and 21 CFR 820.100, Corrective and preventive action. Please note that regardless of whether a change requires premarket review, the QS regulation requires device manufacturers to review and approve changes to device design and production (21 CFR 820.30 and 21 CFR 820.70) and document changes and approvals in the device master record (21 CFR 820.181).

Please be advised that FDA's issuance of a substantial equivalence determination does not mean that FDA has made a determination that your device complies with other requirements of the Act or any Federal statutes and regulations administered by other Federal agencies. You must comply with all the Act's requirements, including, but not limited to: registration and listing (21 CFR Part 807); labeling (21 CFR Part 801 and Part 809); medical device reporting (reporting of medical device-related adverse events) (21 CFR Part 803) for devices or postmarketing safety reporting (21 CFR Part 4, Subpart B) for combination products (see https://www.fda.gov/combination-products/guidance-regulatory-information/postmarketing-safety-reporting-combination-products); good manufacturing practice requirements as set forth in the quality systems (QS) regulation (21 CFR Part 820) for devices or current good manufacturing practices (21 CFR Part 4, Subpart A) for combination products; and, if applicable, the electronic product radiation control provisions (Sections 531-542 of the Act); 21 CFR Parts 1000-1050.

All medical devices, including Class I and unclassified devices and combination product device constituent parts are required to be in compliance with the final Unique Device Identification System rule ("UDI Rule"). The UDI Rule requires, among other things, that a device bear a unique device identifier (UDI) on its label and package (21 CFR 801.20(a)) unless an exception or alternative applies (21 CFR 801.20(b)) and that the dates on the device label be formatted in accordance with 21 CFR 801.18. The UDI Rule (21 CFR 830.300(a) and 830.320(b)) also requires that certain information be submitted to the Global Unique Device Identification Database (GUDID) (21 CFR Part 830 Subpart E). For additional information on these requirements, please see the UDI System webpage at https://www.fda.gov/medical-devices/device-advice-comprehensive-regulatory-assistance/unique-device-identification-system-udi-system.

Also, please note the regulation entitled, "Misbranding by reference to premarket notification" (21 CFR 807.97). For questions regarding the reporting of adverse events under the MDR regulation (21 CFR Part 803), please go to https://www.fda.gov/medical-devices/medical-device-safety/medical-device-reporting-mdr-how-report-medical-device-problems.

For comprehensive regulatory information about medical devices and radiation-emitting products, including information about labeling regulations, please see Device Advice (https://www.fda.gov/medical-devices/device-advice-comprehensive-regulatory-assistance) and CDRH Learn (https://www.fda.gov/training-and-continuing-education/cdrh-learn). Additionally, you may contact the Division of Industry and Consumer Education (DICE) to ask a question about a specific regulatory topic. See the DICE website (https://www.fda.gov/medical-devices/device-advice-comprehensive-regulatory-assistance/contact-us-division-industry-and-consumer-education-dice) for more information or contact DICE by email (DICE@fda.hhs.gov) or phone (1-800-638-2041 or 301-796-7100).

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K251978 - Sarah Baumann Page 3

Sincerely,

JOSEPH BRIGGS -S

Joseph W. Briggs, Ph.D.
Deputy Director
Division of Microbiology Devices
OHT7: Office of In Vitro Diagnostics
Office of Product Evaluation and Quality
Center for Devices and Radiological Health

Enclosure

Page 4

DEPARTMENT OF HEALTH AND HUMAN SERVICES
Food and Drug Administration

Form Approved: OMB No. 0910-0120
Expiration Date: 07/31/2026
See PRA Statement below.

Indications for Use

510(k) Number (if known): K251978

Device Name: LIAISON NES FLU A/B, RSV & COVID-19

Indications for Use (Describe)

Type of Use (Select one or both, as applicable)

☒ Prescription Use (Part 21 CFR 801 Subpart D)
☐ Over-The-Counter Use (21 CFR 801 Subpart C)

CONTINUE ON A SEPARATE PAGE IF NEEDED.

This section applies only to requirements of the Paperwork Reduction Act of 1995.

DO NOT SEND YOUR COMPLETED FORM TO THE PRA STAFF EMAIL ADDRESS BELOW.

The burden time for this collection of information is estimated to average 79 hours per response, including the time to review instructions, search existing data sources, gather and maintain the data needed and complete and review the collection of information. Send comments regarding this burden estimate or any other aspect of this information collection, including suggestions for reducing this burden, to:

Department of Health and Human Services
Food and Drug Administration
Office of Chief Information Officer
Paperwork Reduction Act (PRA) Staff
PRAStaff@fda.hhs.gov

"An agency may not conduct or sponsor, and a person is not required to respond to, a collection of information unless it displays a currently valid OMB number."

FORM FDA 3881 (8/23) Page 1 of 1 PSC Publishing Services (301) 443-6740 EF

Page 5

510(k) Summary

LIAISON® NES FLU A/B, RSV & COVID-19 (NES4450)
LIAISON® NES FLU A/B, RSV & COVID-19 Control Swab Kit (NES4460)
LIAISON® NES Instrument (NES1001)
Page 1 of 30
Confidential

This summary of 510(k) safety and effectiveness information is being submitted in accordance with the requirements of 21 CRF 807.92.

A. 510(k) Number:
K251978

B. Purpose of Submission:
Traditional 510(k), New Device
CLIA Waived

C. Measurand:
Target RNA sequences for Influenza A virus, Influenza B virus, Respiratory Syncytial Virus, and SARS-CoV-2

D. Type of Test:
Qualitative Real Time Reverse Transcription-Polymerase Chain Reaction (RT-PCR)

E. Applicant:
Sarah Baumann
Diasorin Molecular LLC
11331 Valley View Street
Cypress, CA 90630

F. Proprietary and Established Names:
LIAISON® NES FLU A/B, RSV & COVID-19

G. Regulatory Information:

Product Code	Classification	Regulation Section	Panel
QOF	II	21 CFR 866.3981 – Multi Target Respiratory Specimen Nucleic Acid Test Including SARS-CoV-2 and Other Microbial Agents	Microbiology

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LIAISON® NES FLU A/B, RSV & COVID-19 (NES4450)
LIAISON® NES FLU A/B, RSV & COVID-19 Control Swab Kit (NES4460)
LIAISON® NES Instrument (NES1001)
Page 2 of 30
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H. Intended Use:

1. Intended Use(s):

The LIAISON® NES FLU A/B, RSV & COVID-19 assay is a real-time RT-PCR assay intended for use on the LIAISON® NES instrument for the simultaneous in vitro qualitative detection and differentiation of nucleic acid from severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), influenza A (Flu A) virus, influenza B (Flu B) virus, and respiratory syncytial virus (RSV) in anterior nasal swab specimens from individuals with signs and symptoms of respiratory tract infection. Clinical signs and symptoms of respiratory tract infection due to SARS-CoV-2, influenza A, influenza B, and RSV can be similar.

The LIAISON® NES FLU A/B, RSV & COVID-19 assay is intended for use as an aid in the differential diagnosis of SARS-CoV-2, influenza A, influenza B, and RSV infections when used in conjunction with other clinical and epidemiological information, and laboratory findings. SARS-CoV-2, influenza A, influenza B, and RSV viral RNA are generally detectable in anterior nasal swab specimens during the acute phase of infection. This test is not intended to detect influenza C virus infections.

Positive results are indicative of the presence of the identified virus, but do not rule out bacterial infection or co-infection with other pathogens not detected by the test. The agent(s) detected by the LIAISON® NES FLU A/B, RSV & COVID-19 assay may not be the definite cause of the disease. Negative results do not preclude SARS-CoV-2, influenza A, influenza B or RSV infection and should not be used as the sole basis for patient management decisions.

2. Indication(s) for use:

Same as intended use.

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LIAISON® NES FLU A/B, RSV & COVID-19 (NES4450)
LIAISON® NES FLU A/B, RSV & COVID-19 Control Swab Kit (NES4460)
LIAISON® NES Instrument (NES1001)
Page 3 of 30
Confidential

3. Special conditions for use statement(s):

For prescription use only.

4. Special instrument requirements:

For use with LIAISON® NES Instrument.

I. Device Description:

The LIAISON® NES Instrument (NES1001) is capable of analysis of a single cartridge containing a single specimen. A set of parameters specific to the assay is included in the instrument software to name target molecules, assign dyes to probes, specify cycling conditions, and to analyze data from runs. Fluorescence intensity is monitored at each PCR cycle by detection modules within the instrument. The instrument software controls the thermocycling and, upon completion of the run, automatically interprets and displays results for the specimen.

The LIAISON® NES Instrument is comprised of the following:

Touchscreen User Interface
Status LED Indicator
Audio Speaker
Barcode Scanner

The LIAISON® NES software is a graphical user interface (GUI) application that is the end-user interface to the LIAISON® NES Instrument. The software is installed in an embedded computer. The LIAISON® NES software is responsible for providing the environment in which a user runs assays and obtains results.

The LIAISON® NES instrument is intended to accept a cartridge, containing either a quality control (QC) or patient sample, to process and detect for the target nucleic acid.

The LIAISON® NES FLU A/B, RSV & COVID-19 assay used on the LIAISON® NES instrument is a real-time RT-PCR system that enables the direct amplification, detection, and differentiation of influenza A viral RNA, influenza B viral RNA, RSV RNA, and SARS-CoV-2 RNA directly from nasal swabs. Nasal swabs can be professionally collected by a healthcare provider or self-collected under the healthcare provider's supervision.

The LIAISON® NES FLU A/B, RSV & COVID-19 assay consists of the LIAISON® NES instrument, the LIAISON® NES FLU A/B, RSV & COVID-19 Cartridge containing all the

Page 8

LIAISON® NES FLU A/B, RSV & COVID-19 (NES4450)
LIAISON® NES FLU A/B, RSV & COVID-19 Control Swab Kit (NES4460)
LIAISON® NES Instrument (NES1001)
Page 4 of 30
Confidential

required PCR reagents, the NES Sample Vial containing sample release buffer, and the NES Swab for sample collection.

In the LIAISON® NES FLU A/B, RSV & COVID-19 assay, fluorescent probes are used with corresponding forward and reverse primers to amplify influenza A, influenza B, RSV, and SARS-CoV-2, and internal control (IC) RNA. Conserved regions of influenza A viral RNA, influenza B viral RNA, RSV RNA, and SARS-CoV-2 RNA are targeted to identify the viruses in the specimen, while the internal control (IC) RNA is used to detect any PCR failures and/or inhibition.

J. Substantial Equivalence Information:

1. Predicate device name(s):

Biofire® SPOTFIRE® Respiratory (R) Panel Mini

2. Predicate 510(k) number(s):

K230719

3. Comparison with predicate:

The following table compares the similarities and differences between the LIAISON® NES FLU A/B, RSV & COVID-19 assay and the BIOFIRE® SPOTFIRE® Respiratory (R) Panel Mini (SPOTFIRE R Panel Mini) (K230719).

Device & Predicate Device:	Predicate Device: K230719	Candidate Device
Device Trade Name	BIOFIRE® SPOTFIRE Respiratory (R) Panel Mini (SPOTFIRE R Panel Mini)	LIAISON® NES FLU A/B, RSV & COVID-19
General Device Characteristic Similarities
Intended Use/Indications for Use	The BIOFIRE® SPOTFIRE® Respiratory (R) Panel Mini (SPOTFIRE R Panel Mini) is a multiplexed polymerase chain reaction (PCR) test intended for use with the BIOFIRE®	The LIAISON® NES FLU A/B, RSV & COVID-19 assay is a real-time RT-PCR assay intended for use on the LIAISON® NES instrument for the simultaneous in vitro qualitative detection and

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LIAISON® NES FLU A/B, RSV & COVID-19 (NES4450)
LIAISON® NES FLU A/B, RSV & COVID-19 Control Swab Kit (NES4460)
LIAISON® NES Instrument (NES1001)
Page 5 of 30
Confidential

| | SPOTFIRE® System for the simultaneous, qualitative detection and identification of multiple respiratory viral nucleic acids in nasopharyngeal swab (NPS) specimens obtained from individuals with signs and symptoms of respiratory tract infection, including COVID-19. The following organism types and subtypes are identified and differentiated using the SPOTFIRE R Panel Mini: • Coronavirus SARS-CoV-2 • Human rhinovirus • Influenza A virus • Influenza B virus • Respiratory syncytial virus Nucleic acids from the viral organisms identified by this test are generally detectable in NPS specimens during the acute phase of infection. The detection and identification of specific viral nucleic acids from individuals exhibiting signs and/or symptoms of respiratory infection are indicative of the presence of the identified microorganism and aids in diagnosis if used in conjunction with other clinical and epidemiological information, and laboratory findings. The results of this test should not be used as the sole basis for diagnosis, treatment, or other patient management decisions. | differentiation of nucleic acid from severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), influenza A (Flu A) virus, influenza B (Flu B) virus, and respiratory syncytial virus (RSV) in anterior nasal swab specimens from individuals with signs and symptoms of respiratory tract infection. Clinical signs and symptoms of respiratory tract infection due to SARS-CoV-2, influenza A, influenza B, and RSV can be similar. The LIAISON® NES FLU A/B, RSV & COVID-19 assay is intended for use as an aid in the differential diagnosis of SARS-CoV-2, influenza A, influenza B, and RSV infections when used in conjunction with other clinical and epidemiological information, and laboratory findings. SARS-CoV-2, influenza A, influenza B, and RSV viral RNA are generally detectable in anterior nasal swab specimens during the acute phase of infection. This test is not intended to detect influenza C virus infections. Positive results are indicative of the presence of the identified virus, but do not rule out bacterial infection or co-infection with other pathogens not detected by the test. The agent(s) detected by the LIAISON® NES FLU A/B, RSV & COVID-19 assay may not be the definite cause of the disease. Negative results do not |

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LIAISON® NES FLU A/B, RSV & COVID-19 (NES4450)
LIAISON® NES FLU A/B, RSV & COVID-19 Control Swab Kit (NES4460)
LIAISON® NES Instrument (NES1001)
Page 6 of 30
Confidential

	Negative results in the setting of a respiratory illness may be due to infection with pathogens that are not detected by this test, or lower respiratory tract infection that may not be detected by an NPS specimen. Positive results do not rule out coinfection with other organisms. The agent(s) detected by the SPOTFIRE R Panel Mini may not be the definite cause of disease. Additional laboratory testing (e.g., bacterial and viral culture, immunofluorescence, and radiography) may be necessary when evaluating a patient with possible respiratory tract infection.	preclude SARS-CoV-2, influenza A, influenza B or RSV infection and should not be used as the sole basis for patient management decisions.
Measurand	Nucleic acid from Organisms detected	Same
Patient Population	Individuals with signs and symptoms of respiratory tract infection, including COVID-19.	Individuals with signs and symptoms of respiratory tract infection
Organisms detected	• Coronavirus SARS-CoV-2 • Human rhinovirus • Influenza A virus • Influenza B virus • Respiratory syncytial virus	Same, except no assay for Human rhinovirus
Automated System (Sample to Answer)	Automated test interpretation and reporting. User cannot access raw data.	Same
Reporting Features	System software analyzes processed image data and provides test results for each panel target	Same

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LIAISON® NES FLU A/B, RSV & COVID-19 (NES4450)
LIAISON® NES FLU A/B, RSV & COVID-19 Control Swab Kit (NES4460)
LIAISON® NES Instrument (NES1001)
Page 7 of 30
Confidential

	Negative results in the setting of a respiratory illness may be due to infection with pathogens that are not detected by this test, or lower respiratory tract infection that may not be detected by an NPS specimen. Positive results do not rule out coinfection with other organisms. The agent(s) detected by the SPOTFIRE R Panel Mini may not be the definite cause of disease. Additional laboratory testing (e.g., bacterial and viral culture, immunofluorescence, and radiography) may be necessary when evaluating a patient with possible respiratory tract infection.	preclude SARS-CoV-2, influenza A, influenza B or RSV infection and should not be used as the sole basis for patient management decisions.
Measurand	Nucleic acid from Organisms detected	Same
Patient Population	Individuals with signs and symptoms of respiratory tract infection, including COVID-19.	Individuals with signs and symptoms of respiratory tract infection
Organisms detected	• Coronavirus SARS-CoV-2 • Human rhinovirus • Influenza A virus • Influenza B virus • Respiratory syncytial virus	Same, except no assay for Human rhinovirus
Automated System (Sample to Answer)	Automated test interpretation and reporting. User cannot access raw data.	Same
Reporting Features	System software analyzes processed image data and provides test results for each panel target	Same

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LIAISON® NES FLU A/B, RSV & COVID-19 (NES4450)
LIAISON® NES FLU A/B, RSV & COVID-19 Control Swab Kit (NES4460)
LIAISON® NES Instrument (NES1001)
Page 8 of 30
Confidential

Assay Software Functions	Defines panel-specific parameters, instrument protocols and report requirements.	Same
Time to Result	~15 minutes	~18 minutes
General Device Characteristic Differences
Sample Type	Nasopharyngeal Swab (NPS)	Direct Anterior Nasal Swab (NS)
Instrumentation	BIOFIRE SPOTFIRE System	LIAISON® NES Instrument
Technological Principle	Highly multiplexed PCR with DNA melting analysis	Highly multiplexed real-time RT-PCR with fluorescence detection and analysis

K. Standards/Guidance Documents Referenced:

FDA Guidance – Class II Special Controls Guidance Document: Respiratory Viral Panel Multiplex Nucleic Acid Assay.
AAMI. Principles for medical device security – Risk Management. AAMI document TIR57:2016. Association for the Advancement of Medical Instrumentation; 2016.
AAMI. Principles for medical device security – Postmarket risk management for device manufacturers. AAMI document TIR97:2019. Association for the Advancement of Medical Instrumentation; 2019.
CLSI. Information Technology Security of In Vitro Diagnostic Instruments and Software Systems; Approved Standard – Second Edition. CLSI document AUTO11-A2. Wayne, PA: Clinical Laboratory Standards Institute; 2014.
CLSI. Evaluation of Precision of Quantitative Measurement Procedures; Approved Guideline – Third Edition. CLSI document EP05-A3. Wayne, PA: Clinical Laboratory Standards Institute; 2019.
CLSI. Interference Testing in Clinical Chemistry. 3rd Ed. CLSI Document EP07. Wayne, PA: Clinical Laboratory Standards Institute; 2018.
CLSI. Evaluation of Qualitative, Binary Output Examination Performance; Approved Guideline – Third Edition. CLSI document EP12. Wayne, PA: Clinical Laboratory Standards Institute; 2023.
CLSI. Evaluation of Detection Capability for Clinical Laboratory Measurement Procedures; Approved Guideline – Second Edition. CLSI document EP17-A2. Wayne, PA: Clinical Laboratory Standards Institute; 2012.
CLSI. Assessment of the Diagnostic Accuracy of Laboratory Tests Using Receiver Operating Characteristic Curves; Approved Guideline – Second Edition. CLSI document EP24-A2. Wayne, PA: Clinical Laboratory Standards Institute; 2011.

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LIAISON® NES FLU A/B, RSV & COVID-19 (NES4450)
LIAISON® NES FLU A/B, RSV & COVID-19 Control Swab Kit (NES4460)
LIAISON® NES Instrument (NES1001)
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CLSI. Evaluation of Stability of In Vitro Diagnostic Reagents; Approved Guideline. CLSI document EP25-A. Wayne, PA: Clinical Laboratory Standards Institute; 2009.
CLSI. Collection Transport Preparation and Storage of Specimens for Molecular Methods. 2nd Edition. CLSI Document MM13. Wayne, PA: Clinical Laboratory Standards Institute; 2020.
CLSI. Verification and Validation of Multiplex Nucleic Acid Assays. 2nd Edition. CLSI Document MM17. Wayne, PA: Clinical Laboratory Standards Institute; 2018.
ISTA. Packaged-Products for Parcel Delivery System Shipment 70 kg (150 lb) or Less. ISTA Document 3A. International Safe Transit Association. 2018.
IEC 62366-1 Edition 1.1 2020-06 Consolidated Version; Medical devices – Part 1: Application of usability engineering to medical devices
IEC 61010-1 Edition 3.1 2017-01 Consolidated Version; Safety requirements for electrical equipment for measurement, control, and laboratory use – Part 1: General requirements
IEC 60601-1-2 Edition 4.1 2020-09 Consolidated Version; Medical electrical equipment – Part 1-2: General requirements for basic safety and essential performance – Collateral Standard: Electromagnetic disturbances – Requirements and tests
IEC 61326-1 Edition 3.0 2020-10; Electrical equipment for measurement, control and laboratory use – EMC requirements – Part 1: General requirements
IEC 61326-2 Edition 3.0 2020-10; Electrical equipment for measurement, control and laboratory use – EMC requirements – Part 2-6: Particular requirements – In vitro diagnostic (IVD) medical equipment
IEC 62304 Edition 1.1 2015-06 Consolidated Version; Medical device software – Software life cycle processes
IEC TR 60878 Ed. 4.0 2022-11; Graphical symbols for electrical equipment in medical practice [Including: Corrigendum 1 (2023)]
IEC TR 80001-2-2:2012. Application of risk management for IT Networks incorporating medical devices – Part 2-2: Guidance for the disclosure and communication of medical device security needs, risks and controls
IEC TR 80001-2-8 Edition 1.0 206-05; Application of risk management for IT – networks incorporating medical devices – Part 2-8: Application guidance – Guidance on standards for establishing the security capabilities identified in IEC TR 80001-2-2
ISO 14971:2019 Medical Devices – Application of risk management to medical devices
ISO 15223-1: 2021-07 – Medical Devices- Symbols to be used with information to be supplied by the manufacturer – Part 1: General requirements
UL ANSI 2900-1 First Edition 2017; Standard for Safety, Standard for Software Cybersecurity Network-Connectable Products, Part 1: General Requirements
UL ANSI 2900-2-1 First Edition 2017; Standard for Safety, Software Cybersecurity for Network-Connectable Products, Part 2-1: Particular Requirements for Network Connectable Components of Healthcare and Wellness Systems

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LIAISON® NES FLU A/B, RSV & COVID-19 (NES4450)
LIAISON® NES FLU A/B, RSV & COVID-19 Control Swab Kit (NES4460)
LIAISON® NES Instrument (NES1001)
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L. Test Principle:

The LIAISON® NES Instrument is comprised of the following:

Touchscreen User Interface
Status LED Indicator
Audio Speaker
Barcode Scanner

The LIAISON® NES instrument is intended to accept a cartridge, containing either a quality control (QC) or patient sample, to process and detect for the target nucleic acid.

The LIAISON® NES FLU A/B, RSV & COVID-19 assay consists of the LIAISON® NES instrument, the LIAISON® NES FLU A/B, RSV & COVID-19 Cartridge containing all the required PCR reagents, the NES Sample Vial containing sample release buffer, and the NES Swab for sample collection.

M. Clinical Performance Characteristics:

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LIAISON® NES FLU A/B, RSV & COVID-19 (NES4450)
LIAISON® NES FLU A/B, RSV & COVID-19 Control Swab Kit (NES4460)
LIAISON® NES Instrument (NES1001)
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CLINICAL AGREEMENT

The clinical performance of the LIAISON® NES FLU A/B, RSV & COVID-19 Assay was evaluated using clinical specimens prospectively collected between October 2024 and March 2025 from seven geographically diverse clinical sites within the United States. The clinical study utilized prospective specimens collected from pediatric and adult patients exhibiting clinical signs and symptoms of respiratory tract infection. Nasal swabs were professionally collected by a Healthcare Provider or self-collected under the Healthcare Provider's supervision. All collection and LIAISON® NES FLU A/B, RSV & COVID-19 Assay testing were performed at CLIA-Waived facilities, representative of the intended use, point-of-care environment.

A total of 1692 unique prospectively collected specimens that met the pre-determined inclusion criteria were enrolled in the study. Clinical sample testing using the LIAISON® NES FLU A/B, RSV & COVID-19 Assay was performed using the LIAISON® NES device by untrained operators at all seven collection sites.

Of the 1692 specimens enrolled in the prospective arm of the study, 101 (6.0%) specimens were disqualified and removed from further analysis. After a single test of each specimen, 1571 specimens generated valid LIAISON® NES results for a final success rate of 98.7% (1571/1591).

Demographic data for the prospective study population are presented in Table 1.

Table 1: Demographic Data for the Prospective Study Population (N=1591)

	Site 01	Site 02	Site 03	Site 04	Site 05	Site 06	Site 13	Total
Sex
Male	191 (53.1%)	129 (42.7%)	104 (53.3%)	31 (40.3%)	99 (46.3%)	107 (51.4%)	100 (42.6%)	761 (47.8%)
Female	169 (46.9%)	173 (57.3%)	91 (46.7%)	46 (59.7%)	115 (53.7%)	101 (48.6%)	135 (57.4%)	830 (52.2%)
Total	360 (100.0%)	302 (100.0%)	195 (100.0%)	77 (100.0%)	214 (100.0%)	208 (100.0%)	235 (100.0%)	1591 (100.0%)
Age (years)
0-1	47 (13.1%)	33 (10.9%)	42 (21.5%)	0 (0.0%)	9 (4.2%)	10 (4.8%)	5 (2.1%)	146 (9.2%)
>1-5	87 (24.2%)	47 (15.6%)	60 (30.8%)	0 (0.0%)	5 (2.3%)	32 (15.4%)	23 (9.8%)	254 (16.0%)
>5-21	208 (57.8%)	169 (56.0%)	90 (46.2%)	0 (0.0%)	22 (10.3%)	138 (66.3%)	59 (25.1%)	686 (43.1%)
>21-65	18 (5.0%)	52 (17.2%)	3 (1.5%)	68 (88.3%)	142 (66.4%)	28 (13.5%)	132 (56.2%)	443 (27.8%)
>65	0 (0.0%)	1 (0.3%)	0 (0.0%)	9 (11.7%)	36 (16.8%)	0 (0.0%)	16 (6.8%)	62 (3.9%)
Total	360 (100.0%)	302 (100.0%)	195 (100.0%)	77 (100.0%)	214 (100.0%)	208 (100.0%)	235 (100.0%)	1591 (100.0%)

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	Site 01	Site 02	Site 03	Site 04	Site 05	Site 06	Site 13	Total
Subject Status
Emergency Department	0 (0.0%)	0 (0.0%)	184 (94.4%)	41 (53.2%)	214 (100.0%)	0 (0.0%)	0 (0.0%)	439 (27.6%)
Outpatient	360 (100.0%)	302 (100.0%)	6 (3.1%)	35 (45.5%)	0 (0.0%)	208 (100.0%)	235 (100.0%)	1146 (72.0%)
Other	0 (0.0%)	0 (0.0%)	5 (2.6%)	1 (1.3%)	0 (0.0%)	0 (0.0%)	0 (0.0%)	6 (0.4%)
Total	360 (100.0%)	302 (100.0%)	195 (100.0%)	77 (100.0%)	214 (100.0%)	208 (100.0%)	235 (100.0%)	1591 (100.0%)
Collection Method
Healthcare Professional	285 (79.2%)	258 (85.4%)	186 (95.4%)	37 (48.1%)	153 (71.5%)	183 (88.0%)	155 (66.0%)	1257 (79.0%)
Self-collected	75 (20.8%)	44 (14.6%)	9 (4.6%)	40 (51.9%)	61 (28.5%)	25 (12.0%)	80 (34.0%)	334 (21.0%)
Total	360 (100.0%)	302 (100.0%)	195 (100.0%)	77 (100.0%)	214 (100.0%)	208 (100.0%)	235 (100.0%)	1591 (100.0%)

Comparator testing was performed at two external and one internal testing facility. LIAISON® NES FLU A/B, RSV & COVID-19 Assay results were compared to an FDA-cleared molecular assay to determine the Positive Percent Agreement (PPA) and Negative Percent Agreement (NPA), along with the associated 95% confidence intervals. The results for prospective specimen analysis are summarized in Table 2.

Table 2: Positive Percent Agreement (PPA) and Negative Percent Agreement (NPA) of Prospective Data Set

Pathogen Target	Positive Percent Agreement		Negative Percent Agreement
	TP / (TP+FN)	PPA (%)	95% CI	TN / (TN+FP)
Influenza A	384/398ᵃ	96.5%	94.2%-97.9%	1140/1173ᵇ
Influenza B	34/35ᶜ	97.1%	85.5%-99.5%	1531/1536ᵈ
RSV	102/113ᵉ	90.3%	83.4%-94.5%	1455/1458ᶠ
COVID-19	63/64ᵍ	98.4%	91.7%-99.7%	1498/1507ʰ

ᵃ Eight (8) of the 14 Influenza A False Negative specimens were negative by Standard of Care.
ᵇ Twelve (12) of the 33 Influenza A False Positive specimens were positive by bi-directional sequencing (BDS).
ᶜ The one (1) Influenza B False Negative specimen was negative by BDS.
ᵈ Two (2) of the five (5) Influenza B False Positive specimens were positive by Standard of Care.
ᵉ One (1) of the 11 RSV False Negative specimens was negative by BDS, and another two (2) specimens were negative by Standard of Care.
ᶠ Two (2) of the three (3) RSV False Positive specimens were positive by another FDA-cleared molecular assay.
ᵍ The one (1) COVID-19 False Negative specimen was negative by Standard of Care.
ʰ Five (5) of the nine (9) COVID-19 False Positive specimens were positive by Standard of Care.

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REPRODUCIBILITY

Site-to-site reproducibility of the LIAISON® NES FLU A/B, RSV & COVID-19 assay was evaluated by testing LIAISON® NES FLU A/B, RSV & COVID-19 assay cartridges with a reproducibility panel, blinded to operators, consisting of ten panel members including negative samples (Pooled Negative Human Nasal Matrix spiked onto dry nasal swabs (NTC)), positive control (PC) and eight (8) positive panel members. The eight (8) positive panel members consisted of contrived samples containing one (1) strain of SARS-CoV-2, influenza A, influenza B, or RSV viral particles individually. Each strain was spiked onto dry nasal swabs at two different concentrations [Low Positive (LP) ~ 2X Limit of Detection (LoD), Moderate Positive (MP) ~ 5X LoD].

Each sample panel member was tested by two (2) different operators at each testing site. Each operator tested each panel member in two (2) replicates each day for a total of eight (8) non-consecutive testing days. A total of ninety-six (96) replicates [two (2) replicates X two (2) operators X three (3) sites X eight (8) days] were evaluated for each panel member. A total of seven (7) LIAISON® NES instruments [at least two (2) per site] were used to evaluate the reproducibility study.

Table 3: Reproducibility Panel Member Details

Panel Member	Concentration	Panel Member Replicates per day per site	Total panel member replicates per site (16 days)	Total panel member replicates across study
1	Influenza A Low Positive (LP) 2X LoD	2	32	96
2	Influenza B Low Positive (LP) 2X LoD	2	32	96
3	RSV Low Positive (LP) 2X LoD	2	32	96
4	SARS-CoV-2 Low Positive (LP) 2X LoD	2	32	96
5	Influenza A Moderate Positive (MP) 5X LoD	2	32	96
6	Influenza B Moderate Positive (MP) 5X LoD	2	32	96
7	RSV Moderate Positive (MP) 5X LoD	2	32	96
8	SARS-CoV-2 Moderate Positive (MP) 5X LoD	2	32	96
9	NTC – Pooled Negative Human Nasal Matrix NA	2	32	96
10	PC - LIAISON® NES FLU A/B, RSV & COVID-19 Positive Control Swab (PC) 5-10X LoD	2	32	96
Total		20	320	960

LP = Low Positive, MP = Moderate Positive, LoD = Limit of Detection, NTC = No Template Control, PC = Positive Control

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Table 4: LIAISON® NES FLU A/B, RSV & COVID-19 Reproducibility

Sample	Site 1	Site 2	Site 3	All Sites
	Agreement with expected results	Mean Ct ± SD (%CV)	Agreement with expected results	Mean Ct ± SD (%CV)	Agreement with expected results
Influenza A LP	100% (32/32)	36.6 ± 0.77 (2.1%)	100% (32/32)	36.8 ± 1.28 (3.5%)	100% (32/32)
Influenza A MP	100% (32/32)	34.6 ± 1.09 (3.2%)	100% (32/32)	34.7 ± 1.16 (3.4%)	100% (32/32)
Influenza B LP	100% (32/32)	39.0 ± 1.89 (4.8%)	100% (32/32)	38.2 ± 0.88 (2.3%)	96.9% (31/32)ᵃ
Influenza B MP	100% (32/32)	37.1 ± 1.40 (3.8%)	100% (32/32)	36.1 ± 0.78 (2.2%)	100% (32/32)
Sars-CoV-2 LP	100% (32/32)	37.5 ± 1.41 (3.8%)	100% (32/32)	36.6 ± 0.93 (2.6%)	100% (32/32)
Sars-CoV-2 MP	100% (32/32)	35.2 ± 0.91 (2.6%)	100% (32/32)	35.0 ± 0.67 (1.9%)	100% (32/32)
RSV LP	100% (32/32)	36.7 ± 0.96 (2.6%)	100% (32/32)	37.1 ± 1.05 (2.8%)	100% (32/32)
RSV MP	100% (32/32)	35.2 ± 1.01 (2.9%)	100% (32/32)	35.1 ± 0.85 (2.4%)	100% (32/32)
PC Influenza A	100% (32/32)	28.6 ± 0.43 (1.5%)	100% (32/32)	28.9 ± 0.43 (1.5%)	100% (32/32)
PC Influenza B	100% (32/32)	29.4 ± 0.39 (1.3%)	100% (32/32)	29.4 ± 0.28 (0.9%)	100% (32/32)
PC Sars-CoV-2	100% (32/32)	30.9 ± 1.21 (3.9%)	100% (32/32)	30.8 ± 0.29 (0.9%)	100% (32/32)
PC RSV	100% (32/32)	29.8 ± 0.55 (1.8%)	100% (32/32)	29.8 ± 0.31 (1.0%)	100% (32/32)
NTC	100% (32/32)	NA	100% (32/32)	NA	100% (32/32)

Ct = Cycle threshold, SD= Standard Deviation, %CV = Percent Coefficient of Variation, LP = Low Positive, MP = Moderate Positive, NTC = No Template Control, PC = Positive Control
ᵃ One (1) Influenza B Low Positive replicate tested at Site 3 had an unexpected false negative ("Not Detected") result.

Table 5: LIAISON® NES FLU A/B, RSV & COVID-19 Internal Control Reproducibility

Sample	Site 1	Site 2	Site 3	All Sites
	Agreement with expected results	Mean Ct ± SD (%CV)	Agreement with expected results	Mean Ct ± SD (%CV)	Agreement with expected results
Influenza A LP	100% (32/32)	25.0 ± 0.50 (2.0%)	100% (32/32)	24.7 ± 0.41 (1.7%)	100% (32/32)
Influenza A MP	100% (32/32)	24.8 ± 0.48 (1.9%)	100% (32/32)	24.9 ± 0.40 (1.6%)	96.9% (31/32)ᵃ
Influenza B LP	100% (32/32)	24.9 ± 0.53 (2.1%)	100% (32/32)	24.6 ± 0.49 (2.0%)	100% (32/32)
Influenza B MP	100% (32/32)	24.9 ± 0.79 (3.2%)	100% (32/32)	24.5 ± 0.58 (2.4%)	100% (32/32)
SARS-CoV-2 LP	100% (32/32)	24.9 ± 0.64 (2.6%)	100% (32/32)	24.8 ± 0.49 (2.0%)	100% (32/32)
SARS-CoV-2 MP	100% (32/32)	24.9 ± 0.59 (2.4%)	100% (32/32)	24.8 ± 0.45 (1.8%)	100% (32/32)
RSV LP	100% (32/32)	24.9 ± 0.57 (2.3%)	100% (32/32)	24.6 ± 0.48 (1.9%)	100% (32/32)
RSV MP	100% (32/32)	24.9 ± 0.54 (2.2%)	100% (32/32)	24.6 ± 0.41 (1.7%)	100% (32/32)
PC	100% (32/32)	24.0 ± 0.52 (2.2%)	100% (32/32)	23.8 ± 0.36 (1.5%)	100% (32/32)
NTC	100% (32/32)	25.0 ± 0.44 (1.8%)	100% (32/32)	24.6 ± 0.35 (1.4%)	100% (32/32)

Ct = Cycle threshold, SD= Standard Deviation, %CV = Percent Coefficient of Variation, LP = Low Positive, MP = Moderate Positive, NTC = No Template Control, PC = Positive Control
ᵃ One (1) replicate of Influenza A MP at Site 3 had a "Not Detected" IC result.

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N. Analytical Performance Characteristics

ANALYTICAL SENSITIVITY/LIMIT OF DETECTION

The LoD of the LIAISON® NES FLU A/B, RSV & COVID-19 assay on dry nasal swabs (NS) specimens was determined to be the lowest detectable concentration of quantitated titered viral stocks (copies/swab) at which ≥ 95% of all replicates were detected. Two (2) strains of influenza A, two (2) strains of influenza B, two (2) strains of RSV and two (2) strains of SARS-CoV-2 were serially diluted in pooled negative human nasal matrix and spiked onto dry NS to determine the LoD. The LoD results are shown in Table 6.

Table 6: LIAISON® NES FLU A/B, RSV & COVID-19 assay Limit of Detection

Virus strain/isolate	LoD (copies/swab)
Influenza A Victoria/4897/2022 (H1N1)	6000
Influenza A Darwin/9/21 (H3N2)	8000
Influenza B Austria/1359417/2021 (Victoria)	4000
Influenza B Phuket/3073/2013 (Yamagata)	5000
RSVA A Isolate 2006	8000
RSV B Isolate 12/2014	8000
SARS-CoV-2 USA/WA 1/2020	1000
SARS-CoV-2 USA/MDHP20874/2021 (B.1.1.529 Omicron)	1000

The LoD of the LIAISON® NES FLU A/B, RSV & COVID-19 assay using inactivated SARS-CoV-2 WHO International Standard viral particles on dry NS specimens was determined to be the lowest detectable concentration of quantitated titered viral stock (IU/swab) at which ≥ 95% of all replicates were detected. The Second WHO International Standard for SARS-CoV-2 RNA (NIBSC code: 22/252) was serially diluted in pooled negative human nasal matrix and spiked onto dry NS to determine the LoD. The LoD results are shown in Table 7.

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Table 7: LIAISON® NES FLU A/B, RSV & COVID-19 assay Limit of Detection for WHO International Standard for SARS-CoV-2 RNA

Virus strain	LoD (IU/swab)
Second WHO International Standard for SARS-CoV-2 RNA (NIBSC code: 22/252)	400

ANALYTICAL REACTIVITY/CROSS REACTIVITY

Analytical Reactivity

The analytical reactivity of the LIAISON® NES FLU A/B, RSV & COVID-19 assay on dry NS specimens was evaluated. A total of thirty-six (36) influenza A strains, fourteen (14) influenza B strains, nine (9) RSV strains, and nineteen (19) SARS-CoV-2 strains were tested. Quantified viral material was diluted in pooled negative human nasal matrix and spiked onto dry NS at the concentrations listed in Tables 8-11 below (corresponding to 3X LoD) and tested in triplicate. The results are shown in Tables 8-11. All strains and subtypes were 100% detected with LIAISON® NES FLU A/B, RSV & COVID-19 assay at 3X LoD except for SARS-CoV-2 Isolate hCoV-19/USA/CA-Stanford -109_S21/2022 PANGO Lineage B.1.1.529 (XBB Omicron) which was detected at 4X LoD and influenza A/Hong Kong/8/68 (H3N2) which was detected at 5X LoD.

Table 8: LIAISON® NES FLU A/B, RSV & COVID-19 assay Analytical Reactivity – Influenza A

Influenza A Strain	Tested Concentration (copies/swab)	Agreement with Expected Results: %Detection (#Detected/#Total)
A/Michigan/45/2015 (H1N1)	18000	100% (3/3)
A/Brisbane/59/2007 (H1N1)	18000	100% (3/3)
A/NY/02/2009 (H1N1)	18000	100% (3/3)
A/Mexico/4108/2009 (H1N1)	18000	100% (3/3)
A/New York/18/2009 (H1N1)	18000	100% (3/3)
A/Taiwan/42/2006 (H1N1)	18000	100% (3/3)
A/Brisbane/02/2018 (H1N1)	18000	100% (3/3)
A/Wisconsin/588/2019 (H1N1)	18000	100% (3/3)

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Influenza A Strain	Tested Concentration (copies/swab)	Agreement with Expected Results: %Detection (#Detected/#Total)
A/California/07/2009 (H1N1)	18000	100% (3/3)
A/New Caledonia/20/1999 (H1N1)	18000	100% (3/3)
A/Guangdong-Moanan/SWL1536/2019 (H1N1)	18000	100% (3/3)
A/Puerto Rico/8/1934 (H1N1)	18000	100% (3/3)
A/Victoria/2570/2019 (H1N1)	18000	100% (3/3)
A/Nebraska/14/2018 (H1N1)	18000	100% (3/3)
A/Sydney/05/2021 (H1N1)	18000	100% (3/3)
A/Switzerland/9715293/2013 (H3N2)	24000	100% (3/3)
A/Hong Kong/4801/2014 (H3N2)	24000	100% (3/3)
A/Singapore/INFIMH-16-0019/2016 (H3N2)	24000	100% (3/3)
A/Perth/16/2009 (H3N2)	24000	100% (3/3)
A/Hong Kong/2671/2019 (H3N2)	24000	100% (3/3)
A/Hong Kong/8/1968 (H3N2)	24000 (3x LoD)	67% (2/3)
	32000 (4x LoD)	67% (2/3)
	40000 (5x LoD)	100% (3/3)
A/Cambodia/E0826360/2020 (H3N2)	24000	100% (3/3)
A/Kansas/14/2017 (H3N2)	24000	100% (3/3)
A/Port Chalmers/1/1973 (H3N2)	24000	100% (3/3)
A/Darwin/6/2021 (H3N2)	24000	100% (3/3)
A/Tasmania/503/2020 (H3N2)	24000	100% (3/3)
A/Thailand/08/2022 (H3N2)	24000	100% (3/3)

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Influenza A Strain	Tested Concentration (copies/swab)	Agreement with Expected Results: %Detection (#Detected/#Total)
A/Texas/50/2012 (H3N2)	24000	100% (3/3)
A/Massachusetts/18/2022 (H3N2)	24000	100% (3/3)
A/swine/Ohio/09SW1484E/2009 (Extracted RNA) (H1N2)	18000	100% (3/3)
A/Anhui/01/2005 (H5N1)	18000	100% (3/3)
A/Egypt/N03072/2010 (H5N1)	18000	100% (3/3)
A/Hubei/1/2010 (H5N1)	18000	100% (3/3)
A/Mallard/Netherlands/12/2000 (H7N7)	18000	100% (3/3)
A/Hong Kong/33982/2009 (H9N2)	18000	100% (3/3)
A/bovine/Ohio.B24OSU-439/2024 gRNA (H5N1)	18000	100% (3/3)

Table 9: LIAISON® NES FLU A/B, RSV & COVID-19 assay Analytical Reactivity – Influenza B

Influenza B Strain	Tested Concentration (copies/swab)	Agreement with Expected Results: %Detection (#Detected/#Total)
B/Brisbane/60/2008 (Victoria)	12000	100% (3/3)
B/Brisbane/33/2008 (Victoria)	12000	100% (3/3)
B/Washington/02/2019 (Victoria)	12000	100% (3/3)
B/Colorado/06/2017 (Victoria)	12000	100% (3/3)
B/Florida/02/2006 (Victoria)	12000	100% (3/3)
B/Malaysia/2506/2004 (Victoria)	12000	100% (3/3)
B/Texas/2/2013 (Victoria)	12000	100% (3/3)
B/Maryland/1/1959 (Lineage unknown)	12000	100% (3/3)

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Influenza B Strain	Tested Concentration (copies/swab)	Agreement with Expected Results: %Detection (#Detected/#Total)
B/Massachusetts/02/2012 (Yamagata)	15000	100% (3/3)
B/Wisconsin/1/2010 (Yamagata)	15000	100% (3/3)
B/Florida/07/2004 (Yamagata)	15000	100% (3/3)
B/Florida/04/2006 (Yamagata)	15000	100% (3/3)
B/Panama/45/1990 (Yamagata)	15000	100% (3/3)
B/Utah/9/2014 (Yamagata)	15000	100% (3/3)

Table 10: LIAISON® NES FLU A/B, RSV & COVID-19 assay Analytical Reactivity – RSV

RSV Strain	Tested Concentration (copies/swab)	Agreement with Expected Results: %Detection (#Detected/#Total)
RSV-A 1/2015 Isolate 1	24000	100% (3/3)
RSV-A2	24000	100% (3/3)
RSV-A 3/2015 Isolate 3	24000	100% (3/3)
RSV-A 4/2015 Isolate 1	24000	100% (3/3)
RSV-B 9320	24000	100% (3/3)
RSV-B CH93(18)-18	24000	100% (3/3)
RSV-B 3/2015 Isolate 1	24000	100% (3/3)
RSV-B1	24000	100% (3/3)
RSV-B 3/2015 Isolate 2	24000	100% (3/3)

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Table 11: LIAISON® NES FLU A/B, RSV & COVID-19 assay Analytical Reactivity – SARS-CoV-2

SARS-CoV-2 Strain	Tested Concentration (copies/swab)	Agreement with Expected Results: %Detection (#Detected/#Total)
SARS-CoV-2 Isolate hCoV-19/USA/PHC658/2021 B.1.617.2 (Delta)	3000	100% (3/3)
SARS-CoV-2 Isolate Hong Kong/VM20001061/2020	3000	100% (3/3)
SARS-CoV-2 Isolate hCoV-19/Japan/TY7-503/2021 (Gamma Variant)	3000	100% (3/3)
SARS-CoV-2 Isolate USA/MD-HP24556/2022 BA.2.3 (Omicron)	3000	100% (3/3)
SARS-CoV-2 Isolate South Africa/KRISP-ECK005325/2020 (B.1.351 South Africa variant)	3000	100% (3/3)
SARS-CoV-2 Isolate England/204820464/2020 B.1.1.7 (UK variant)	3000	100% (3/3)
SARS-CoV-2 USA/CA-Stanford-15_S02/2021 B.1.617.1 (Kappa Variant)	3000	100% (3/3)
SARS-CoV-2 USA/NY-Wadsworth-21025952-01/2021 B.1.526_2021 (Iota Variant)	3000	100% (3/3)
SARS-CoV-2 USA-WI 1/2020 (Lineage B)	3000	100% (3/3)
SARS-CoV-2 NY-Wadsworth-21006055-01/2021 Lineage P2_2021 (Zeta Variant)	3000	100% (3/3)
SARS-Related Coronavirus 2 Isolate hCoV19/USA/New York/PV96109/2023 (Omicron Variant JN.1)	3000	100% (3/3)
SARS-CoV-2 Isolate USA-CA3/2020	3000	100% (3/3)
SARS-CoV-2 Isolate Germany/BavPat1/2020 (Lineage B)	3000	100% (3/3)
SARS-CoV-2 Isolate USA/CA/VRLC014/2021 PANGO Lineage B.1.429 (Epsilon)	3000	100% (3/3)
SARS-CoV-2 Isolate hCoV-19/USA/VAFBCH_675/2021 PANGO Lineage AY4.2	3000	100% (3/3)
SARS-CoV-2 Isolate SARS-CoV-2 Peru/un-CDC-2-4069945/2021 PANGO Lineage C.37 (Lambda)	3000	100% (3/3)
SARS-CoV-2 isolate hCoV-191 USA/MDHP38861/2022 PANGO Lineage B.1.1.529, BQ.1.1 (Omicron)	3000	100% (3/3)
	3000 (3x LoD)	67% (2/3)

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SARS-CoV-2 Strain	Tested Concentration (copies/swab)	Agreement with Expected Results: %Detection (#Detected/#Total)
SARS-CoV-2 Isolate hCoV-19/USA/CA-Stanford - 109_S21/2022 PANGO Lineage B.1.1.529, (XBB Omicron)	4000 (4x LoD)	100% (3/3)
SARS-CoV-2 Lineage JN.1.4 Omicron Variant USA/NY-Wadsworth-23068107-01/2023	3000	100% (3/3)

In silico Analytical Reactivity/Inclusivity

An in silico inclusivity analysis of the assay oligo sequences in the LIAISON® NES FLU A/B, RSV & COVID-19 assay was performed. All primer and probe sets were aligned against sequences available in the GISAID EpiCoV (as of May 5, 2025), EpiFlu (between January 1, 2012 and May 4, 2025) or EpiRSV (as of May 4, 2025) databases depending on the target. Specifically, 5,641,034 sequences of SARS-CoV-2, 230093 sequences of influenza A, 54413 sequences of influenza B and 22929 sequences of Respiratory Syncytial Virus were aligned. Based on the in silico analysis, the LIAISON® NES FLU A/B, RSV & COVID-19 assay exhibits ~100% inclusivity to influenza A, influenza B, SARS-CoV-2 and RSV sequences available in the aforementioned databases.

Cross-Reactivity (Analytical Specificity)

Cross-reactivity of the LIAISON® NES FLU A/B, RSV & COVID-19 assay was evaluated by testing whole microorganisms (or purified nucleic acid from microorganisms) that are closely related, cause similar clinical symptoms, or may be present in the same sample type.

Specimens were prepared by diluting cultured isolates, inactivated organisms, or purified nucleic acids (whole genome) in pooled negative human nasal matrix and spiked onto dry NS. Cross-reactivity was determined based on three replicates. For microorganisms not titered in CFU/mL, copies/mL or TCID50/mL, the maximum volume possible was used. Results from cross-reactivity testing are summarized in Table 12. No cross-reactivity was observed with any of the microorganisms tested.

Table 12: LIAISON® NES FLU A/B, RSV & COVID-19 Assay Cross-Reactivity (Analytical Specificity)

Organism	Tested Concentration	Agreement with Expected Results: % Detection (#Detected/#Total)
Bordetella parapertussis	1E6 CFU/mL	0% (0/3)
Bordetella pertussis	1E6 CFU/mL	0% (0/3)
Candida albicans	1E6 CFU/mL	0% (0/3)
Corynebacterium diphtheriae	1E6 CFU/mL	0% (0/3)

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Organism	Tested Concentration	Agreement with Expected Results: % Detection (#Detected/#Total)
Escherichia coli	1E6 CFU/mL	0% (0/3)
Fusobacterium necrophorum	1E6 CFU/mL	0% (0/3)
Haemophilus influenzae	1E6 CFU/mL	0% (0/3)
Lactobacillus casei	1E6 CFU/mL	0% (0/3)
Legionella pneumophila	1E6 CFU/mL	0% (0/3)
Leptospira interrogans	Quantification not provided by the manufacturer. Maximum testable volume used.	0% (0/3)
Moraxella catarrhalis	1E6 CFU/mL	0% (0/3)
Neisseria elongata	1E6 CFU/mL	0% (0/3)
Neisseria gonorrhoeae	1E6 CFU/mL	0% (0/3)
Neisseria meningitidis	1E6 CFU/mL	0% (0/3)
Pneumocystis jirovecii	1E6 CFU/mL	0% (0/3)
Pseudomonas aeruginosa	1E6 CFU/mL	0% (0/3)
Staphylococcus aureus	1E6 CFU/mL	0% (0/3)
Staphylococcus epidermidis BAA3171	1E6 CFU/mL	0% (0/3)
Streptococcus pneumoniae	1E6 CFU/mL	0% (0/3)
Streptococcus pyogenes M1	1E6 CFU/mL	0% (0/3)
Streptococcus salivarius	1E6 CFU/mL	0% (0/3)
Aspergillus fumigatus	1E6 copies/mL	0% (0/3)
Chlamydia pneumoniae	1E6 copies/mL	0% (0/3)
Mycobacterium tuberculosis (genomic DNA)	1E6 copies/mL	0% (0/3)
Mycoplasma genitalium	1E6 copies/mL	0% (0/3)

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Organism	Tested Concentration	Agreement with Expected Results: % Detection (#Detected/#Total)
Mycoplasma pneumoniae	1E6 copies/mL	0% (0/3)
Adenovirus 7A	1E5 TCID50/mL	0% (0/3)
Adenovirus Type 31	1E5 TCID50/mL	0% (0/3)
CMV AD-169	1E5 TCID50/mL	0% (0/3)
Coronavirus 229E	1E5 TCID50/mL	0% (0/3)
Coronavirus HKU1 (synthetic RNA)	1E5 genome copies/mL	0% (0/3)
Coronavirus NL63	1E5 TCID50/mL	0% (0/3)
Coronavirus OC43	1E5 TCID50/mL	0% (0/3)
EBV	1E5 cps/mL	0% (0/3)
Enterovirus Type 68	1E5 TCID50/mL	0% (0/3)
hMPV 9	1E5 TCID50/mL	0% (0/3)
Influenza C SendaiITU2IO8	1E5 TCID50/mL	0% (0/3)
Measles virus	1E5 TCID50/mL	0% (0/3)
MERS Coronavirus (Florida/USA2_Saudi Arabia_2014)	1E5 cps/mL	0% (0/3)
Mumps virus	1E5 TCID50/mL	0% (0/3)
Parainfluenza virus Type 1	1E5 TCID50/mL	0% (0/3)
Parainfluenza virus Type 2	1E5 TCID50/mL	0% (0/3)
Parainfluenza virus Type 3	1E5 TCID50/mL	0% (0/3)
Parainfluenza virus Type 4A	1E5 TCID50/mL	0% (0/3)
Parechovirus Type 1	1E5 TCID50/mL	0% (0/3)
Rhinovirus Type 1A	1E5 TCID50/mL	0% (0/3)
SARS Coronavirus	Quantification not provided by the manufacturer. Maximum testable volume used.	0% (0/3)

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Organism	Tested Concentration	Agreement with Expected Results: % Detection (#Detected/#Total)
Pooled human nasal wash	10%	0% (0/3)

CFU/mL= colony forming units/milliliter
copies/mL= copies/milliliter
TCID50/mL= tissue culture infectious dose/milliliter

In silico Cross Reactivity/Exclusivity

The analytical specificity of the LIAISON® NES FLU A/B, RSV & COVID-19 assay was further evaluated with an extended in silico analysis to predict potential cross-reactivity of the assay oligos through a BLAST comparison of all primers and probes in the assay to the human reference genome and the GenBank nt sequence database. Based on the in silico exclusivity analysis, the assay oligos are predicted to have no cross reactivity to human genome sequences from the Homo sapiens reference genome GRCh38.p14. In silico assessment of the analyzed potential cross-reactive organisms, with sequences available in the GenBank nt database as of May 3, 2025, predicts potential cross-reactivity for assay oligos of SARS-CoV-2 to some bat coronavirus and bat SARS-like coronavirus strains.

INTERFERING SUBSTANCES

Potentially interfering substances from respiratory specimens were tested for the ability to generate false positive or false negative results. Samples were prepared by: 1) diluting each potentially interfering substance into a baseline sample consisting of pooled negative human nasal matrix and influenza A Darwin/9/21 (H3N2), influenza B Phuket 3073/2013 (Yamagata), SARS-CoV-2 USA/WA 1/2020 or RSV B Isolate 12/2014 at 3X LoD and 2) diluting each potentially interfering substance into a baseline sample consisting of pooled negative human nasal matrix. Prepared test samples were spiked onto dry NS and interference based on three replicates was determined. The results are shown in Table 13 and Table 14.

Remdesivir at a concentration of 0.1 mg/mL showed a negative result in one out of the three replicates for RSV. The replicates were retested at a lower concentration of 0.05 mg/mL and no interference was observed for RSV. No other substances tested in Table 12 showed any interference with the detection of SARS-CoV-2, influenza A or influenza B at the concentrations tested. No interference was observed for negative samples.

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Table 13: LIAISON® NES FLU A/B, RSV & COVID-19 assay Interference for Positive Sample

Potentially Interfering Substances	Active Ingredient	Interferent Concentration	Influenza A % Detection (#Detected/#Tested)	Influenza B % Detection (#Detected/#Tested)	SARS-CoV-2 % Detection (#Detected/#Tested)	RSV % Detection (#Detected/#Tested)
Afrin Nasal Spray	Oxymetazoline	15% (v/v)	100% (3/3)	100% (3/3)	100% (3/3)	100% (3/3)
Antibacterial, systemic	Tobramycin	0.004 mg/mL	100% (3/3)	100% (3/3)	100% (3/3)	100% (3/3)
Antibiotic, nasal ointment	Mupirocin	6.6 mg/mL	100% (3/3)	100% (3/3)	100% (3/3)	100% (3/3)
Blood	N/A	2% (v/v)	100% (3/3)	100% (3/3)	100% (3/3)	100% (3/3)
Purified Mucin Protein	Bovine submaxillary gland mucin, type I-S	5 mg/mL	100% (3/3)	100% (3/3)	100% (3/3)	100% (3/3)
Nasal corticosteroid (Beconase AQ)	Beclomethasone	5% (w/v)	100% (3/3)	100% (3/3)	100% (3/3)	100% (3/3)
	Dexamethasone	5% (w/v)	100% (3/3)	100% (3/3)	100% (3/3)	100% (3/3)
	Mometasone	5% (w/v)	100% (3/3)	100% (3/3)	100% (3/3)	100% (3/3)
Nasal corticosteroid (Fluticasone)	Fluticasone	5% (v/v)	100% (3/3)	100% (3/3)	100% (3/3)	100% (3/3)
Tamiflu Antiviral drug	Oseltamivir	0.001 mM	100% (3/3)	100% (3/3)	100% (3/3)	100% (3/3)
Zicam Nasal Gel	Luffa operculata, Galphimia glauca, histaminum hydrochloricum	5 % (v/v)	100% (3/3)	100% (3/3)	100% (3/3)	100% (3/3)
Budesonide	Budesonide	5% (v/v)	100% (3/3)	100% (3/3)	100% (3/3)	100% (3/3)
Flunisolide	Flunisolide	5% (v/v)	100% (3/3)	100% (3/3)	100% (3/3)	100% (3/3)
Relenza Antiviral Drug	Zanamivir	3.3 mg/mL	100% (3/3)	100% (3/3)	100% (3/3)	100% (3/3)
Throat lozenges	Avena sativa, Zinc gluconate, Sambucus nigra, Echinacea, Rose hips, Licorice root	1.25% (w/v)	100% (3/3)	100% (3/3)	100% (3/3)	100% (3/3)
Remdesivir	Remdesivir	0.1 mg/mL	100% (3/3)	100% (3/3)	100% (3/3)	67% (2/3)
		0.05 mg/mL	N/A	N/A	N/A	100% (3/3)
Neo-Synephrine nasal spray	Phenylephrine hydrochloride	15% (v/v)	100% (3/3)	100% (3/3)	100% (3/3)	100% (3/3)

mg/mL= milligram/milliliter
v/v= volume per volume
w/v= weight per volume
mM= millimolar

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Table 14: LIAISON® NES FLU A/B, RSV & COVID-19 Assay Interference for Negative Sample

Potentially Interfering Substances	Active Ingredient	Interferent Concentration	Influenza A % Detection (#Detected/#Tested)	Influenza B % Detection (#Detected/#Tested)	SARS-CoV-2 % Detection (#Detected/#Tested)	RSV % Detection (#Detected/#Tested)	IC % Detection (#Detected/#Tested)
Afrin Nasal Spray	Oxymetazoline	15% (v/v)	0% (0/3)	0% (0/3)	0% (0/3)	0% (0/3)	100% (3/3)
Antibacterial, systemic	Tobramycin	0.004 mg/mL	0% (0/3)	0% (0/3)	0% (0/3)	0% (0/3)	100% (3/3)
Antibiotic, nasal ointment	Mupirocin	6.6 mg/mL	0% (0/3)	0% (0/3)	0% (0/3)	0% (0/3)	100% (3/3)
Blood	N/A	2% (v/v)	0% (0/3)	0% (0/3)	0% (0/3)	0% (0/3)	100% (3/3)
Purified Mucin Protein	Bovine submaxillary gland mucin, type I-S	5 mg/mL	0% (0/3)	0% (0/3)	0% (0/3)	0% (0/3)	100% (3/3)
Nasal corticosteroid (Beconase AQ)	Beclomethasone	5% (w/v)	0% (0/3)	0% (0/3)	0% (0/3)	0% (0/3)	100% (3/3)
	Dexamethasone	5% (w/v)	0% (0/3)	0% (0/3)	0% (0/3)	0% (0/3)	100% (3/3)
	Mometasone	5% (w/v)	0% (0/3)	0% (0/3)	0% (0/3)	0% (0/3)	100% (3/3)
Nasal corticosteroid (Fluticasone)	Fluticasone	5% (v/v)	0% (0/3)	0% (0/3)	0% (0/3)	0% (0/3)	100% (3/3)
Tamiflu Antiviral drug	Oseltamivir	0.001 mM	0% (0/3)	0% (0/3)	0% (0/3)	0% (0/3)	100% (3/3)
Zicam Nasal Gel	Luffa operculata, Galphimia glauca, histaminum hydrochloricum	5 % (v/v)	0% (0/3)	0% (0/3)	0% (0/3)	0% (0/3)	100% (3/3)
Budesonide	Budesonide	5% (v/v)	0% (0/3)	0% (0/3)	0% (0/3)	0% (0/3)	100% (3/3)
Flunisolide	Flunisolide	5% (v/v)	0% (0/3)	0% (0/3)	0% (0/3)	0% (0/3)	100% (3/3)
Relenza Antiviral Drug	Zanamivir	3.3 mg/mL	0% (0/3)	0% (0/3)	0% (0/3)	0% (0/3)	100% (3/3)
Throat lozenges	Avena sativa, Zinc gluconate, Sambucus nigra, Echinacea, Rose hips, Licorice root	1.25% (w/v)	0% (0/3)	0% (0/3)	0% (0/3)	0% (0/3)	100% (3/3)
Remdesivir	Remdesivir	0.1 mg/mL	0% (0/3)	0% (0/3)	0% (0/3)	0% (0/3)	100% (3/3)
		0.05 mg/mL	0% (0/3)	0% (0/3)	0% (0/3)	0% (0/3)	100% (3/3)
Neo-Synephrine nasal spray	Phenylephrine hydrochloride	15% (v/v)	0% (0/3)	0% (0/3)	0% (0/3)	0% (0/3)	100% (3/3)

mg/mL= milligram/milliliter
v/v= volume per volume
w/v= weight per volume
mM= millimolar
IC = Internal Control

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COMPETITIVE INTERFERENCE

Competitive Interference was performed to assess the ability of the assay to detect a low concentration of one (1) target analyte in the presence of a high concentration of another target analyte. Samples were prepared by diluting one (1) assay target analyte at a low concentration (3X LoD) into pooled negative human nasal matrix in the presence of a high concentration (1000X LoD) of one (1) of the other three (3) assay target analytes. Prepared specimens were spiked onto dry NS and tested in triplicate. The results are shown in Table 15. All the combinations tested showed no competitive interference for the detection of low concentrations of influenza A, influenza B, SARS-CoV-2 or RSV in the presence of high concentrations of another assay target analyte.

Table 15: LIAISON® NES FLU A/B, RSV & COVID-19 Assay Competitive Interference

Low Positive Baseline Sample		Competitive Interferent		Agreement with Expected Results: % Detection (#Detected/#Total)
Strain	Copies/swab	Strain	Copies/swab	Influenza A	Influenza B	SARS-CoV-2	RSV
Influenza A Victoria/4897/2022	18000	Influenza B Austria/1359417/2021	4E6	100% (3/3)	100% (3/3)	0% (0/3)	0% (0/3)
		SARS-CoV-2 USA/MDHP20874/2021	1E6	100% (3/3)	0% (0/3)	100% (3/3)	0% (0/3)
		Respiratory Syncytial Virus A 2006 Isolate	8E6	100% (3/3)	0% (0/3)	0% (0/3)	100% (3/3)
Influenza B Austria/1359417/2021	12000	Influenza A Victoria/4897/2022	6E6	100% (3/3)	100% (3/3)	0% (0/3)	0% (0/3)
		SARS-CoV-2 USA/MDHP20874/2021	1E6	0% (0/3)	100% (3/3)	100% (3/3)	0% (0/3)
		Respiratory Syncytial Virus A 2006 Isolate	8E6	0% (0/3)	100% (3/3)	0% (0/3)	100% (3/3)
SARS-CoV-2 USA/MDHP20874/2021	3000	Influenza A Victoria/4897/2022	6E6	100% (3/3)	0% (0/3)	100% (3/3)	0% (0/3)
		Influenza B Austria/1359417/2021	4E6	0% (0/3)	100% (3/3)	100% (3/3)	0% (0/3)
		Respiratory Syncytial Virus A 2006 Isolate	8E6	0% (0/3)	0% (0/3)	100% (3/3)	100% (3/3)
Respiratory Syncytial Virus A 2006 Isolate	24000	Influenza A Victoria/4897/2022	6E6	100% (3/3)	0% (0/3)	0% (0/3)	100% (3/3)
		Influenza B Austria/1359417/2021	4E6	0% (0/3)	100% (3/3)	0% (0/3)	100% (3/3)
		SARS-CoV-2 USA/MDHP20874/2021	1E6	0% (0/3)	0% (0/3)	100% (3/3)	100% (3/3)

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INHIBITION BY OTHER MICROORGANISMS

The LIAISON® NES FLU A/B, RSV & COVID-19 assay was evaluated by testing the ability to detect a low concentration of SARS-CoV-2, influenza A, influenza B, and RSV when other potentially inhibitory microorganisms were present. Specimens were prepared by diluting the potentially inhibitory cultured isolates, inactivated organisms or purified nucleic acids (whole genome) into pooled negative human nasal matrix in the presence of a low concentration (3X LoD) of either influenza A Darwin/9/21 (H3N2), influenza B Phuket 3073/2013 (Yamagata), RSV B Isolate 12/2014 or SARS-CoV-2 USA/WA 1/2020. Forty-eight (48) potentially inhibitory microorganisms were individually spiked and tested in triplicate. For organisms not titered in CFU/mL or TCID50/mL, the maximum volume possible was used.

Staphylococcus aureus and Parainfluenza Virus Type 1 at a high concentration of 1E6 CFU/mL and 1E5 TCID50/mL, respectively, produced a negative result in one (1) out of three (3) replicates for influenza B. A lower concentration of these microorganisms (Staphylococcus aureus at 3.13E5 CFU/mL and Parainfluenza Virus Type 1 at 3.13E4 TCID50/mL) were tested and no interference was observed. No inhibition by other organisms was observed for SARS-CoV-2, influenza A, influenza B or RSV at the concentrations indicated in Table 16.

Table 16: LIAISON® NES FLU A/B, RSV & COVID-19 Assay Microbial Inhibition

Organism	Tested Concentration	Agreement with Expected Results: %Detection (#Detected/#Total)
		Influenza A	Influenza B	SARS-CoV-2	RSV
Bordetella parapertussis	1E6 CFU/mL	100% (3/3)	100% (3/3)	100% (3/3)	100% (3/3)
Bordetella pertussis	1E6 CFU/mL	100% (3/3)	100% (3/3)	100% (3/3)	100% (3/3)
Candida albicans	1E6 CFU/mL	100% (3/3)	100% (3/3)	100% (3/3)	100% (3/3)
Corynebacterium diphtheriae	1E6 CFU/mL	100% (3/3)	100% (3/3)	100% (3/3)	100% (3/3)
Escherichia coli	1E6 CFU/mL	100% (3/3)	100% (3/3)	100% (3/3)	100% (3/3)
Fusobacterium necrophorum	1E6 CFU/mL	100% (3/3)	100% (3/3)	100% (3/3)	100% (3/3)
Haemophilus influenzae	1E6 CFU/mL	100% (3/3)	100% (3/3)	100% (3/3)	100% (3/3)
Lactobacillus casei	1E6 CFU/mL	100% (3/3)	100% (3/3)	100% (3/3)	100% (3/3)
Legionella pneumophila	1E6 CFU/mL	100% (3/3)	100% (3/3)	100% (3/3)	100% (3/3)
Leptospira interrogans	Quantification not provided by the manufacturer.	100% (3/3)	100% (3/3)	100% (3/3)	100% (3/3)

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Organism	Tested Concentration	Agreement with Expected Results: %Detection (#Detected/#Total)
		Influenza A	Influenza B	SARS-CoV-2	RSV
	Maximum testable volume used.
Moraxella catarrhalis	1E6 CFU/mL	100% (3/3)	100% (3/3)	100% (3/3)	100% (3/3)
Neisseria elongata	1E6 CFU/mL	100% (3/3)	100% (3/3)	100% (3/3)	100% (3/3)
Neisseria gonorrhoeae	1E6 CFU/mL	100% (3/3)	100% (3/3)	100% (3/3)	100% (3/3)
Neisseria meningitidis	1E6 CFU/mL	100% (3/3)	100% (3/3)	100% (3/3)	100% (3/3)
Pneumocystis jirovecii	1E6 CFU/mL	100% (3/3)	100% (3/3)	100% (3/3)	100% (3/3)
Pseudomonas aeruginosa	1E6 CFU/mL	100% (3/3)	100% (3/3)	100% (3/3)	100% (3/3)
Staphylococcus aureus	1E6 CFU/mL	100% (3/3)	0% (0/3)	100% (3/3)	100% (3/3)
	3.13E5 CFU/mL	N/A	100% (3/3)	N/A	N/A
Staphylococcus epidermidis BAA-3171	1E6 CFU/mL	100% (3/3)	100% (3/3)	100% (3/3)	100% (3/3)
Streptococcus pneumoniae	1E6 CFU/mL	100% (3/3)	100% (3/3)	100% (3/3)	100% (3/3)
Streptococcus pyogenes M1	1E6 CFU/mL	100% (3/3)	100% (3/3)	100% (3/3)	100% (3/3)
Streptococcus salivarius	1E6 CFU/mL	100% (3/3)	100% (3/3)	100% (3/3)	100% (3/3)
Aspergillus fumigatus	1E6 copies/mL	100% (3/3)	100% (3/3)	100% (3/3)	100% (3/3)
Chlamydia pneumoniae	1E6 copies/mL	100% (3/3)	100% (3/3)	100% (3/3)	100% (3/3)
Mycobacterium tuberculosis (genomic DNA)	1E6 copies/mL	100% (3/3)	100% (3/3)	100% (3/3)	100% (3/3)
Mycoplasma genitalium	1E6 copies/mL	100% (3/3)	100% (3/3)	100% (3/3)	100% (3/3)
Mycoplasma pneumoniae	1E6 copies/mL	100% (3/3)	100% (3/3)	100% (3/3)	100% (3/3)
Adenovirus 7A	1E5 TCID50/mL	100% (3/3)	100% (3/3)	100% (3/3)	100% (3/3)
Adenovirus Type 31	1E5 TCID50/mL	100% (3/3)	100% (3/3)	100% (3/3)	100% (3/3)
CMV AD-169	1E5 TCID50/mL	100% (3/3)	100% (3/3)	100% (3/3)	100% (3/3)

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LIAISON® NES FLU A/B, RSV & COVID-19 (NES4450)
LIAISON® NES FLU A/B, RSV & COVID-19 Control Swab Kit (NES4460)
LIAISON® NES Instrument (NES1001)
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Confidential

Organism	Tested Concentration	Agreement with Expected Results: %Detection (#Detected/#Total)
		Influenza A	Influenza B	SARS-CoV-2	RSV
Coronavirus 229E	1E5 TCID50/mL	100% (3/3)	100% (3/3)	100% (3/3)	100% (3/3)
Coronavirus HKU1 (synthetic RNA)	1E5 genome copies/mL	100% (3/3)	100% (3/3)	100% (3/3)	100% (3/3)
Coronavirus NL63	1E5 TCID50/mL	100% (3/3)	100% (3/3)	100% (3/3)	100% (3/3)
Coronavirus OC43	1E5 TCID50/mL	100% (3/3)	100% (3/3)	100% (3/3)	100% (3/3)
EBV	1E5 cps/mL	100% (3/3)	100% (3/3)	100% (3/3)	100% (3/3)
Enterovirus Type 68	1E5 TCID50/mL	100% (3/3)	100% (3/3)	100% (3/3)	100% (3/3)
hMPV 9	1E5 TCID50/mL	100% (3/3)	100% (3/3)	100% (3/3)	100% (3/3)
Influenza C SendaiITU2IO8	1E5 TCID50/mL	100% (3/3)	100% (3/3)	100% (3/3)	100% (3/3)
Measles virus	1E5 TCID50/mL	100% (3/3)	100% (3/3)	100% (3/3)	100% (3/3)
MERS Coronavirus (Florida/USA2_Saudi Arabia_2014)	1E5 cps/mL	100% (3/3)	100% (3/3)	100% (3/3)	100% (3/3)
Mumps virus	1E5 TCID50/mL	100% (3/3)	100% (3/3)	100% (3/3)	100% (3/3)
Parainfluenza virus Type 1	1E5 TCID50/mL	100% (3/3)	33% (1/3)	100% (3/3)	100% (3/3)
	3.13E4 TCID50/mL	N/A	100% (3/3)	N/A	N/A
Parainfluenza virus Type 2	1E5 TCID50/mL	100% (3/3)	100% (3/3)	100% (3/3)	100% (3/3)
Parainfluenza virus Type 3	1E5 TCID50/mL	100% (3/3)	100% (3/3)	100% (3/3)	100% (3/3)
Parainfluenza virus Type 4A	1E5 TCID50/mL	100% (3/3)	100% (3/3)	100% (3/3)	100% (3/3)
Parechovirus Type 1	1E5 TCID50/mL	100% (3/3)	100% (3/3)	100% (3/3)	100% (3/3)
SARS Coronavirus	Quantification not provided by the manufacturer. Maximum testable volume used.	100% (3/3)	100% (3/3)	100% (3/3)	100% (3/3)
Rhinovirus Type 1A	1E5 TCID50/mL	100% (3/3)	100% (3/3)	100% (3/3)	100% (3/3)
Pooled human nasal wash	10%	100% (3/3)	100% (3/3)	100% (3/3)	100% (3/3)

CFU/mL= colony forming units/milliliter
Cps/mL= copies/milliliter
TCID50/mL= tissue culture infectious dose/milliliter

CARRY-OVER CONTAMINATION

Amplification carry-over and cross-contamination for the LIAISON® NES FLU A/B, RSV & COVID-19 assay has been assessed. The study was designed by processing the samples alternating between highly positive and negative samples. High Positive (HP) samples were formulated by spiking SARS-CoV-2 USA/MDHP20874/2021 in pooled negative human nasal matrix onto dry NS at a final concentration of 1000X LoD (1E8 copies/mL). Pooled negative nasal matrix onto dry NS was used as negative sample. The same alternating order was maintained when loading the corresponding cartridges into the instruments. No evidence of carry-over contamination was observed.

ASSAY CUT-OFF

For each channel, the respective target is detected if (1) the fluorescence signal crosses the fluorescence threshold, and (2) the cycle at which it crosses is at/before the last cycle (the Ct threshold, 45 cycles).

MATRIX COMPARISON

Not applicable

O. Proposed Labeling:

The labeling provided in the submission satisfies the requirements of 21 CFR 809.10.

P. Conclusion:

The submitted information in this premarket notification is complete and supports a substantial equivalence decision.

Regulation Number and Section

§ 866.3981 Device to detect and identify nucleic acid targets in respiratory specimens from microbial agents that cause the SARS-CoV-2 respiratory infection and other microbial agents when in a multi-target test.

(a)
Identification. A device to detect and identify nucleic acid targets in respiratory specimens from microbial agents that cause the SARS-CoV-2 respiratory infection and other microbial agents when in a multi-target test is an in vitro diagnostic device intended for the detection and identification of SARS-CoV-2 and other microbial agents when in a multi-target test in human clinical respiratory specimens from patients suspected of respiratory infection who are at risk for exposure or who may have been exposed to these agents. The device is intended to aid in the diagnosis of respiratory infection in conjunction with other clinical, epidemiologic, and laboratory data or other risk factors.(b)
Classification. Class II (special controls). The special controls for this device are:(1) The intended use in the labeling required under § 809.10 of this chapter must include a description of the following: Analytes and targets the device detects and identifies, the specimen types tested, the results provided to the user, the clinical indications for which the test is to be used, the specific intended population(s), the intended use locations including testing location(s) where the device is to be used (if applicable), and other conditions of use as appropriate.
(2) Any sample collection device used must be FDA-cleared, -approved, or -classified as 510(k) exempt (standalone or as part of a test system) for the collection of specimen types claimed by this device; alternatively, the sample collection device must be cleared in a premarket submission as a part of this device.
(3) The labeling required under § 809.10(b) of this chapter must include:
(i) A detailed device description, including reagents, instruments, ancillary materials, all control elements, and a detailed explanation of the methodology, including all pre-analytical methods for processing of specimens;
(ii) Detailed descriptions of the performance characteristics of the device for each specimen type claimed in the intended use based on analytical studies including the following, as applicable: Limit of Detection, inclusivity, cross-reactivity, interfering substances, competitive inhibition, carryover/cross contamination, specimen stability, precision, reproducibility, and clinical studies;
(iii) Detailed descriptions of the test procedure(s), the interpretation of test results for clinical specimens, and acceptance criteria for any quality control testing;
(iv) A warning statement that viral culture should not be attempted in cases of positive results for SARS-CoV-2 and/or any similar microbial agents unless a facility with an appropriate level of laboratory biosafety (
e.g., BSL 3 and BSL 3+, etc.) is available to receive and culture specimens; and(v) A prominent statement that device performance has not been established for specimens collected from individuals not identified in the intended use population (
e.g., when applicable, that device performance has not been established in individuals without signs or symptoms of respiratory infection).(vi) Limiting statements that indicate that:
(A) A negative test result does not preclude the possibility of infection;
(B) The test results should be interpreted in conjunction with other clinical and laboratory data available to the clinician;
(C) There is a risk of incorrect results due to the presence of nucleic acid sequence variants in the targeted pathogens;
(D) That positive and negative predictive values are highly dependent on prevalence;
(E) Accurate results are dependent on adequate specimen collection, transport, storage, and processing. Failure to observe proper procedures in any one of these steps can lead to incorrect results; and
(F) When applicable (
e.g., recommended by the Centers for Disease Control and Prevention, by current well-accepted clinical guidelines, or by published peer-reviewed literature), that the clinical performance may be affected by testing a specific clinical subpopulation or for a specific claimed specimen type.(4) Design verification and validation must include:
(i) Detailed documentation, including performance results, from a clinical study that includes prospective (sequential) samples for each claimed specimen type and, as appropriate, additional characterized clinical samples. The clinical study must be performed on a study population consistent with the intended use population and compare the device performance to results obtained using a comparator that FDA has determined is appropriate. Detailed documentation must include the clinical study protocol (including a predefined statistical analysis plan), study report, testing results, and results of all statistical analyses.
(ii) Risk analysis and documentation demonstrating how risk control measures are implemented to address device system hazards, such as Failure Modes Effects Analysis and/or Hazard Analysis. This documentation must include a detailed description of a protocol (including all procedures and methods) for the continuous monitoring, identification, and handling of genetic mutations and/or novel respiratory pathogen isolates or strains (
e.g., regular review of published literature and periodic in silico analysis of target sequences to detect possible mismatches). All results of this protocol, including any findings, must be documented and must include any additional data analysis that is requested by FDA in response to any performance concerns identified under this section or identified by FDA during routine evaluation. Additionally, if requested by FDA, these evaluations must be submitted to FDA for FDA review within 48 hours of the request. Results that are reasonably interpreted to support the conclusion that novel respiratory pathogen strains or isolates impact the stated expected performance of the device must be sent to FDA immediately.(iii) A detailed description of the identity, phylogenetic relationship, and other recognized characterization of the respiratory pathogen(s) that the device is designed to detect. In addition, detailed documentation describing how to interpret the device results and other measures that might be needed for a laboratory diagnosis of respiratory infection.
(iv) A detailed device description, including device components, ancillary reagents required but not provided, and a detailed explanation of the methodology, including molecular target(s) for each analyte, design of target detection reagents, rationale for target selection, limiting factors of the device (
e.g., saturation level of hybridization and maximum amplification and detection cycle number, etc.), internal and external controls, and computational path from collected raw data to reported result (e.g., how collected raw signals are converted into a reported signal and result), as applicable.(v) A detailed description of device software, including software applications and hardware-based devices that incorporate software. The detailed description must include documentation of verification, validation, and hazard analysis and risk assessment activities, including an assessment of the impact of threats and vulnerabilities on device functionality and end users/patients as part of cybersecurity review.
(vi) For devices intended for the detection and identification of microbial agents for which an FDA recommended reference panel is available, design verification and validation must include the performance results of an analytical study testing the FDA recommended reference panel of characterized samples. Detailed documentation must be kept of that study and its results, including the study protocol, study report for the proposed intended use, testing results, and results of all statistical analyses.
(vii) For devices with an intended use that includes detection of Influenza A and Influenza B viruses and/or detection and differentiation between the Influenza A virus subtypes in human clinical specimens, the design verification and validation must include a detailed description of the identity, phylogenetic relationship, or other recognized characterization of the Influenza A and B viruses that the device is designed to detect, a description of how the device results might be used in a diagnostic algorithm and other measures that might be needed for a laboratory identification of Influenza A or B virus and of specific Influenza A virus subtypes, and a description of the clinical and epidemiological parameters that are relevant to a patient case diagnosis of Influenza A or B and of specific Influenza A virus subtypes. An evaluation of the device compared to a currently appropriate and FDA accepted comparator method. Detailed documentation must be kept of that study and its results, including the study protocol, study report for the proposed intended use, testing results, and results of all statistical analyses.
(5) When applicable, performance results of the analytical study testing the FDA recommended reference panel described in paragraph (b)(4)(vi) of this section must be included in the device's labeling under § 809.10(b) of this chapter.
(6) For devices with an intended use that includes detection of Influenza A and Influenza B viruses and/or detection and differentiation between the Influenza A virus subtypes in human clinical specimens in addition to detection of SARS-CoV-2 and similar microbial agents, the required labeling under § 809.10(b) of this chapter must include the following:
(i) Where applicable, a limiting statement that performance characteristics for Influenza A were established when Influenza A/H3 and A/H1-2009 (or other pertinent Influenza A subtypes) were the predominant Influenza A viruses in circulation.
(ii) Where applicable, a warning statement that reads if infection with a novel Influenza A virus is suspected based on current clinical and epidemiological screening criteria recommended by public health authorities, specimens should be collected with appropriate infection control precautions for novel virulent influenza viruses and sent to State or local health departments for testing. Viral culture should not be attempted in these cases unless a BSL 3+ facility is available to receive and culture specimens.
(iii) Where the device results interpretation involves combining the outputs of several targets to get the final results, such as a device that both detects Influenza A and differentiates all known Influenza A subtypes that are currently circulating, the device's labeling must include a clear interpretation instruction for all valid and invalid output combinations, and recommendations for any required followup actions or retesting in the case of an unusual or unexpected device result.
(iv) A limiting statement that if a specimen yields a positive result for Influenza A, but produces negative test results for all specific influenza A subtypes intended to be differentiated (
i.e., H1-2009 and H3), this result requires notification of appropriate local, State, or Federal public health authorities to determine necessary measures for verification and to further determine whether the specimen represents a novel strain of Influenza A.(7) If one of the actions listed at section 564(b)(1)(A) through (D) of the Federal Food, Drug, and Cosmetic Act occurs with respect to an influenza viral strain, or if the Secretary of Health and Human Services determines, under section 319(a) of the Public Health Service Act, that a disease or disorder presents a public health emergency, or that a public health emergency otherwise exists, with respect to an influenza viral strain:
(i) Within 30 days from the date that FDA notifies manufacturers that characterized viral samples are available for test evaluation, the manufacturer must have testing performed on the device with those influenza viral samples in accordance with a standardized protocol considered and determined by FDA to be acceptable and appropriate.
(ii) Within 60 days from the date that FDA notifies manufacturers that characterized influenza viral samples are available for test evaluation and continuing until 3 years from that date, the results of the influenza emergency analytical reactivity testing, including the detailed information for the virus tested as described in the certificate of authentication, must be included as part of the device's labeling in a tabular format, either by:
(A) Placing the results directly in the device's labeling required under § 809.10(b) of this chapter that accompanies the device in a separate section of the labeling where analytical reactivity testing data can be found, but separate from the annual analytical reactivity testing results; or
(B) In a section of the device's label or in other labeling that accompanies the device, prominently providing a hyperlink to the manufacturer's public website where the analytical reactivity testing data can be found. The manufacturer's website, as well as the primary part of the manufacturer's website that discusses the device, must provide a prominently placed hyperlink to the website containing this information and must allow unrestricted viewing access.