Predicate Devices

Reference Devices

Not Found

AI/ML (Artificial Intelligence / Machine Learning)

No
The description focuses on standard molecular diagnostic techniques (RT-PCR, array hybridization) and automated processing, with no mention of AI or ML in the analysis or interpretation of results. The "Flex Software" allows for custom panel selection, which is a configuration feature, not an AI/ML capability.

Therapeutic

No.
Explanation: This device is an in vitro diagnostic (IVD) test that detects and identifies bacterial and viral nucleic acids to aid in the diagnosis of respiratory infections. It does not provide treatment or directly apply therapy, which are hallmarks of a therapeutic device. The "Intended Use" explicitly states that the results "should not be used as the sole basis for diagnosis, treatment, or patient management decisions," further clarifying its diagnostic, not therapeutic, role.

Diagnostic

Yes
The "Intended Use / Indications for Use" section explicitly states that the device "aids in the diagnosis of respiratory infection."

SaMD (Software as a Medical Device)

No

The device description explicitly states that the system is composed of the LIAISON PLEX Instrument, the LIAISON PLEX® System Software (preinstalled on the LIAISON PLEX Instrument), the LIAISON PLEX® Respiratory Flex Assay cartridge, and the LIAISON PLEX® Respiratory Flex Assay File. This includes significant hardware components (Instrument, cartridge) in addition to the software.

IVD (In Vitro Diagnostic)

Yes, this device is an IVD (In Vitro Diagnostic).

Here's why:

Intended Use: The "Intended Use / Indications for Use" section explicitly states that the LIAISON PLEX Respiratory Flex (RSP Flex) Assay is a "multiplexed qualitative test for the simultaneous in vitro detection and identification of multiple bacterial and viral nucleic acids in nasopharyngeal swabs (NPS) obtained from individuals with clinical signs and symptoms of respiratory tract infection, including SARS-CoV-2." The phrase "in vitro detection and identification" is a key characteristic of IVDs.
Device Description: The description details a system that processes biological specimens (nasopharyngeal swabs) to detect specific analytes (bacterial and viral nucleic acids) using laboratory techniques (RT, PCR, array hybridization). This is consistent with the definition of an IVD.
Performance Studies: The document describes clinical performance studies using patient specimens (nasopharyngeal swabs) to evaluate the device's ability to detect the target organisms. This type of testing is performed to demonstrate the clinical utility of an IVD.
Predicate Device: The mention of a "Predicate Device(s)" with a K number (DEN200031) and name (BioFire Respiratory Panel 2.1 (RP2.1)) indicates that this device is being compared to a previously cleared IVD, which is a standard process for regulatory submission of new IVDs.

All of these points strongly support the classification of this device as an In Vitro Diagnostic.

PCCP (Predetermined Change Control Plan) Authorized

N/A

Overview

Intended Use / Indications for Use

The LIAISON PLEX Respiratory Flex (RSP Flex) Assay is a multiplexed qualitative test for the simultaneous in vitro detection and identification of multiple bacterial and viral nucleic acids in nasopharyngeal swabs (NPS) obtained from individuals with clinical signs and symptoms of respiratory tract infection, including SARS-CoV-2. The test is performed on the automated LIAISON PLEX System utilizing reverse transcription (RT), polymerase chain reaction (PCR), and array hybridization to detect specific nucleic acid gene sequences of the following organism types and subtypes:

Viruses: Adenovirus Human Coronavirus (HKU1, NL63, OC43, and 229E not differentiated) Human Enterovirus/Rhinovirus (not differentiated) Human Metapneumovirus, Influenza A Influenza A (subtype H1) Influenza A (subtype H3) Influenza B Parainfluenza 1 Parainfluenza 2 Parainfluenza 3 Parainfluenza 4 Respiratory Syncytial Virus Severe Acute Respiratory Syndrome Coronavirus (SARS-CoV-2)

Bacteria: Bordetella holmesi Bordetella parapertussis Bordetella pertussis Chlamydia pneumoniae Mycoplasma pneumoniae

Nucleic acids from the bacterial and viral organisms identified by this test are generally detectable in NPS specimens during the acute phase of infection. Detecting and identifying specific bacterial and viral nucleic acids from individuals exhibiting signs and symptoms of respiratory infection aids in the diagnosis of respiratory infection, if used in conjunction with other clinical, epidemiological, and laboratory findings. The results of this test should not be used as the sole basis for diagnosis, treatment, or patient management decisions.

Negative results in the presence of a respiratory illness may be due to infection with pathogens that are not detected by this test or due to lower respiratory that is not detected by an NPS specimen. Conversely, positive results do not rule out infection or co-infection with organisms not detected by the LIAISON PLEX Respiratory Flex (RSP Flex) Assay. The agent(s) detected may not be the definite cause of disease.

The use of additional laboratory testing (e.g., bacterial and viral culture, immunofluorescence, and radiography), may be necessary when evaluating a patient with possible respiratory tract infection.

Product codes (comma separated list FDA assigned to the subject device)

QOF, OEM, OOU, OTG, OZE, OZX, OZY, OZZ, OCC, NSU

Device Description

The LIAISON PLEX® Respiratory Flex Assay is a multiplexed nucleic acid test system composed of the LIAISON PLEX Instrument, the LIAISON PLEX® System Software (preinstalled on the LIAISON PLEX Instrument), the LIAISON PLEX® Respiratory Flex Assay cartridge, and the LIAISON PLEX® Respiratory Flex Assay File. The LIAISON PLEX® Respiratory Flex Assay cartridge contains the reagents to perform nucleic acid extraction and purification, reverse transcription, PCR, and array hybridization. Specifically, the LIAISON PLEX® Respiratory Flex Assay detects bacteria and viruses from nasopharyngeal swab (NPS) specimens collected from individuals with signs and symptoms of respiratory infection.

The LIAISON PLEX System consists of a touchscreen user interface that includes the software for running and analyzing assay results, one to six processing/imaging LIAISON PLEX modules, and a handheld barcode reader. Each LIAISON PLEX module processes one sample at a time under the control of the LIAISON PLEX System software.

LIAISON PLEX® automates the sample processing through analysis within a single cartridge. Processing steps include 1.) Sample Preparation: Nucleic acid extraction from organisms by chemical and mechanical means and isolation of nucleic acid on magnetic beads 2.) Target Amplification: Multiplex PCR and RT-PCR based amplification of extracted nucleic acid to generate target specific amplicons 3.) Hybridization: Amplicons hybridize with their target specific DNA probe arranged in a microarray format and that are attached to mediator and gold nanoparticles 4.) Analysis: Gold nanoparticles specifically bound to target amplicons are silver enhanced and the light scatter from microarray spot is measured and analyzed to confirm presence (Detected) or absence (not Detected) of a target.

The LIAISON PLEX Respiratory Flex Assay has the option of creating and processing results for custom panels using Flex® Software. Flex Software allows users to randomly select and group targets in tiers for result processing. Up to 7 targets may be selected for the initial test tier. After the first tier, each additional tier requires a specific number of credits. Flex™ credits allow the end-user to create custom panels and pay for a smaller subset of results tailored to the individual patient's clinical presentation. Alternatively, a laboratory may choose the fixed price option where all target results are processed at the same time.

Mentions image processing

Not Found

Mentions AI, DNN, or ML

Not Found

Input Imaging Modality

Not Found

Anatomical Site

nasopharyngeal swabs (NPS)

Indicated Patient Age Range

pediatric and adult patients

Intended User / Care Setting

For prescription use only. For in vitro diagnostic use only. For use with LIAISON PLEX Systems only.

Description of the training set, sample size, data source, and annotation protocol

Not Found

Description of the test set, sample size, data source, and annotation protocol

Prospective Clinical Evaluation:
Sample Size: 1911 unique prospective specimens enrolled initially, 1843 specimens for evaluation.
Data Source: Clinical specimens prospectively collected between October 2022 to April 2023 from six geographically diverse clinical sites within the United States. Remnant, de-identified specimens from pediatric and adult patients exhibiting clinical signs and symptoms of respiratory tract infections.
Annotation Protocol: Specimens were stored refrigerated at 2-8°C for up to 72 hours before testing (Category I) or frozen at -70℃ (Category II). Clinical runs were performed by trained operators. Comparator methods were FDA-cleared molecular respiratory panels for most analytes, FDA-cleared molecular SARS-CoV-2 assay for SARS-CoV-2, and analytically validated Fragment Analysis (FA) assays followed by PCR/Bi-Directional Sequencing (PCR/BDS) assays for Bordetella species.

Testing of Preselected Archived Specimens:
Sample Size: 256 pre-selected left-over frozen, de-identified specimens.
Data Source: Four sites/vendors in the United States, collection dates from November 2013 through June 2023.
Annotation Protocol: Specimens were characterized by the same comparator methods as the prospective study. Tested in a randomized, blinded manner with negative specimens.

Contrived Specimen Testing:
Sample Size: 300 contrived specimens.
Data source: Contrived specimens for low prevalence targets (Bordetella holmesii, Bordetella parapertussis, Bordetella pertussis, Chlamydia pneumoniae, Mycoplasma pneumoniae) and Influenza A H1N1 pdm09.
Annotation Protocol: Specimens were contrived, blinded, randomized, and tested along with negative specimens at two testing sites.

Summary of Performance Studies (study type, sample size, AUC, MRMC, standalone performance, key results)

1. Analytical performance:
a. Precision/Reproducibility:
Within Laboratory Precision:
Study type: Within laboratory precision study.
Sample size: 45 total replicates (3 lots, 5 non-consecutive days, 3 replicates per day).
Key results:
Bordetella pertussis: 93.3% (1.5X LoD), 100% (5X LoD), 100% (Negative).
Adenovirus: 97.8% (1.5X LoD), 97.8% (5X LoD), 100% (Negative).
Influenza B: 100% (1.5X LoD), 100% (5X LoD), 100% (Negative).
hMPV: 100% (1.5X LoD), 97.8% (5X LoD), 100% (Negative).
SARS-CoV-2: 100% (1.5X LoD), 100% (5X LoD), 100% (Negative).

    **Reproducibility:**
    Study type: Multi-site reproducibility study.
    Sample size: 90 replicates per target (3 sites, 2 operators per site, 5 non-consecutive days, 3 replicates per day).
    Key results:
    Adenovirus: 97.8% (low positive), 100% (moderate positive).
    Bordetella pertussis: 96.7% (low positive), 100% (moderate positive).
    Influenza B: 96.7% (low positive), 100% (moderate positive).
    Human Metapneumovirus: 94.4% (low positive), 97.8% (moderate positive).
    SARS-CoV-2: 98.9% (low positive), 100% (moderate positive).
    Negative NPS: 100%.

**c. Traceability, Stability, Expected values (controls, calibrators, or methods):**
    **Specimen Stability:**
    Study type: Specimen stability evaluation.
    Key results: Specimens stored frozen (≤ -70°C) are stable for up to 30 days, refrigerated (2°C - 8°C) for up to 72 hours, and at room temperature (15°C - 30°C) for up to 8 hours.

    **Fresh vs. Frozen Specimen Stability:**
    Study type: Fresh vs. frozen and freeze/thaw cycle assessment.
    Key results: NPS specimens in UVT/UTM can undergo up to two freeze/thaw cycles prior to testing.

**d. Detection Limit:**
    Study type: Limit of Detection (LoD) study.
    Sample size: 39 strains/isolates tested individually; 20 replicates for confirmed LoD.
    Key results: Confirmed LoD for each target organism is provided in Table 5 (pages 17-18). For WHO International Standard for SARS-CoV-2: 7.7x10^5 IU/mL.

**e. Analytical Reactivity (Inclusivity):**
    **Laboratory (Wet Testing):**
    Study type: Analytical reactivity (inclusivity) evaluation.
    Sample size: 181 isolates and clinical samples (34 bacteria, 147 viruses).
    Key results: 176 strains detected with 100% positivity at 3X LoD. Four strains (influenza A Brisbane (02/18), coronavirus NL63 (NR-470), coronavirus 229E (VR-740) and SARS-CoV-2 (Stanford)) were detected at 9X LoD and one strain (B. holmesii (CIP 104396)) was detected at 27X LoD. (Table 7, pages 19-25)

    **In Silico Inclusivity for SARS-CoV-2 and Influenza:**
    Study type: In silico analysis.
    Sample size: SARS-CoV-2: 5,622,325 sequences; Influenza: 112,056 (Influenza A), 54,364 (H1), 104,428 (H3), 26,470 (Influenza B).
    Key results: SARS-CoV-2: 100% of sequences evaluated are expected to be detected. Influenza: 99.9% of Influenza A, 98.9% of H1, 99.9% of H3, and 99.9% of Influenza B sequences are expected to be detected based on >90% homology criterion. (Table 8, page 26).

**f. Analytical specificity:**
    **Cross-Reactivity and Microbial Interference:**
        **Off-Panel Cross Reactivity (Laboratory (Wet) Testing):**
        Study type: Cross-reactivity assessment.
        Sample size: 60 off-panel viral, fungal, and bacterial organisms.
        Key results: 59 organisms yielded negative results. Mycoplasma genitalium cross-reacted with Mycoplasma pneumoniae assay at 4x10^6 CCU/mL, but not at 1x10^6 CCU/mL. (Table 9, page 27)

        **In Silico Cross-Reactivity:**
        Study type: In silico specificity/exclusivity analysis (BLAST comparison).
        Sample size: 83 off-panel organisms (68 bacteria/fungi, 15 viruses) and all on-panel organisms.
        Key results: Predicted cross-reactions listed in Table 10 (page 28-29): Adenovirus G, bat/pangolin coronavirus with SARS-CoV-2, B. bronchiseptica (with IS1001) with B. parapertussis, certain swine/avian influenza strains with Influenza A H1/H3.

        **On-Panel Cross Reactivity:**
        Study type: Intra-panel cross-reactivity evaluation.
        Sample size: 28 on-panel organisms.
        Key results: All 28 on-panel organisms yielded expected results and did not cross-react with other target assays. (Table 11, page 29).

    **Microbial Interference:**
    Study type: Microbial interference study.
    Sample size: 16 potentially interfering non-panel microbial organisms.
    Key results: Most organisms did not interfere. Streptococcus pyogenes and Legionella pneumophila interfered with Adenovirus detection in 1 of 6 replicates.

    **Interfering Substances:**
    Study type: Evaluation of inhibitory effect of non-microbial substances.
    Sample size: 36 interfering substances.
    Key results: Human Sputum/Mucus, Human Whole Blood (at higher concentrations), Leukocytes (at higher concentrations), Mupirocin, and Tobramycin showed interference with certain target organisms at specific concentrations. (Table 14, page 32).

    **Competitive Inhibition/Co-infection:**
    Study type: Competitive inhibition/co-infection study.
    Sample size: 27 pairings of clinically prevalent co-infections.
    Key results: 48 combinations showed no interference. Interference was observed for Parainfluenza 3 (low concentration) with human coronavirus OC43 and adenovirus 37D; RSV A (low concentration) with adenovirus 37D; Flu A H3N2 (low concentration) with adenovirus 37D; Human coronavirus 229E (low concentration) with SARS-CoV-2; SARS-CoV-2 (low concentration) with human coronavirus OC43. Interference was resolved for some pairs at lower high-concentration levels of the interfering agent. (Table 15, pages 33-34).

    **Carry-Over and Cross-Contamination:**
    Study type: Carry-over and cross-contamination evaluation.
    Sample size: 30 positive and 30 negative tests across 5 consecutive runs using two systems.
    Key results: No carry-over or cross-contamination was observed.

2. Clinical Performance:
a. Prospective Clinical Evaluation:
Study type: Multi-site prospective clinical study.
Sample size: 1843 evaluable clinical specimens.
Key results:
Initial invalid rate: 4.8% (88/1843). Final invalid rate: 0.6% (11/1843).
Overall PPA and NPA are summarized in Table 18 (pages 38-39).
Examples by target (Overall):
Adenovirus: PPA 100%, NPA 95.7%.
Human Coronavirus: PPA 90.0%, NPA 99.5%.
Enterovirus/Rhinovirus: PPA 93.7%, NPA 97.8%.
SARS-CoV-2: PPA 96.5%, NPA 99.5%.
Bordetella parapertussis: PPA 80.0%, NPA 99.8%.

**b. Testing of Preselected Archived Specimens:**
Study type: Evaluation of preselected archived retrospective NPS specimens.
Sample size: 256 specimens.
Key results:
First-attempt valid rate: 94.1%. Final success rate after retest: 100%.
Performance summarized in Table 22 (pages 46-47).
Examples by target:
Bordetella parapertussis: PPA 100%, NPA 98.7%.
Bordetella pertussis: PPA 100%, NPA 98.6%.
Mycoplasma pneumoniae: PPA 95.8%, NPA 97.4%.

**c. Contrived Specimen Testing:**
Study type: Contrived specimen testing to supplement low prevalence targets.
Sample size: 300 specimens.
Key results:
First-attempt valid rate: 97.0%. Final success rate after retest: 100%.
Performance summarized in Table 23 (pages 47-48).
Examples by target (Combined):
Bordetella holmesii: PPA 100%, NPA 100%.
Bordetella parapertussis: PPA 98.0%, NPA 100%.
Bordetella pertussis: PPA 100%, NPA 100%.
Chlamydia pneumoniae: PPA 100%, NPA 100%.
Influenza A H1N1 pdm09: PPA 98.0%, NPA 100%.
Mycoplasma pneumoniae: PPA 98.0%, NPA 99.6%.

Key Metrics (Sensitivity, Specificity, PPV, NPV, etc.)

Key metrics are presented as Positive Percent Agreement (PPA) and Negative Percent Agreement (NPA).

Prospective Clinical Evaluation (Overall):

Adenovirus: PPA = 100% (96.1-100%), NPA = 95.7% (94.7-96.6%)
Bordetella holmesii: PPA = NE, NPA = 100% (99.8-100%)
Bordetella parapertussis: PPA = 80.0% (37.6-96.4%), NPA = 99.8% (99.5-99.9%)
Bordetella pertussis: PPA = NE, NPA = 100% (99.8-100%)
Chlamydia pneumoniae: PPA = NE, NPA = 100% (99.8-100%)
Human Coronavirus: PPA = 90.0% (83.6-94.1%), NPA = 99.5% (99.1-99.8%)
Enterovirus/Rhinovirus: PPA = 93.7% (90.5-95.8%), NPA = 97.8% (96.9-98.4%)
hMPV: PPA = 95.4% (90.4-97.9%), NPA = 99.6% (99.2-99.8%)
Influenza A: PPA = 100% (97.1-100%), NPA = 99.1% (98.5-99.4%)
Influenza A Subtype H1: PPA = 100% (90.6-100%), NPA = 99.9% (99.7-100%)
Influenza A Subtype H3: PPA = 97.2% (92.1-99.0%), NPA = 99.8% (99.4-99.9%)
Influenza B: PPA = 100% (67.6-100%), NPA = 100% (99.8-100%)
Mycoplasma pneumoniae: PPA = NE, NPA = 100% (99.8-100%)
Parainfluenza 1: PPA = 91.7% (64.6-98.5%), NPA = 100% (99.8-100%)
Parainfluenza 2: PPA = 92.3% (66.7-98.6%), NPA = 100% (99.8-100%)
Parainfluenza 3: PPA = 93.2% (81.8-97.7%), NPA = 99.9% (99.7-100%)
Parainfluenza 4: PPA = 88.9% (56.5-98.0%), NPA = 99.9% (99.7-100%)
Respiratory Syncytial Virus: PPA = 95.9% (90.8-98.3%), NPA = 100% (99.8-100%)
SARS-CoV-2: PPA = 96.5% (93.4-98.1%), NPA = 99.5% (99.0-99.7%)

Archived Performance Summary:

Adenovirus: PPA = 100% (61.0-100%), NPA = 96.4% (93.3-98.1%)
Bordetella holmesii: PPA = NE, NPA = 100% (98.4-100%)
Bordetella parapertussis: PPA = 100% (67.6-100%), NPA = 98.7% (96.3-99.6%)
Bordetella pertussis: PPA = 100% (85.7-100%), NPA = 98.6% (96.0-99.5%)
Chlamydia pneumoniae: PPA = 92.9% (68.5-98.7%), NPA = 99.6% (97.7-99.9%)
Human Coronavirus: PPA = 100% (51.0-100%), NPA = 98.8% (96.6-99.6%)
Enterovirus/Rhinovirus: PPA = 88.9% (71.9-96.1%), NPA = 97.4% (94.4-98.8%)
hMPV: PPA = 100% (20.7-100%), NPA = 100% (98.5-100%)
Influenza A: PPA = 100% (20.7-100%), NPA = 99.6% (97.8-99.9%)
Influenza A Subtype H1: PPA = 100% (20.7-100%), NPA = 99.6% (97.8-99.9%)
Influenza A Subtype H3: PPA = NE, NPA = 100% (98.5-100%)
Influenza B: PPA = 100% (85.7-100%), NPA = 99.6% (97.6-99.9%)
Mycoplasma pneumoniae: PPA = 95.8% (79.8-99.3%), NPA = 97.4% (94.5-98.8%)
Parainfluenza 1: PPA = 100% (82.4-100%), NPA = 99.6% (97.7-99.9%)
Parainfluenza 2: PPA = 95.0% (76.4-99.1%), NPA = 99.6% (97.6-99.9%)
Parainfluenza 3: PPA = 100% (34.2-100%), NPA = 100% (98.5-100%)
Parainfluenza 4: PPA = 100% (85.7-100%), NPA = 98.7% (96.3-99.6%)
Respiratory Syncytial Virus: PPA = 100% (70.1-100%), NPA = 99.6% (97.7-99.9%)

Contrived Specimens (Combined):

Bordetella holmesii: PPA = 100% (92.9-100%), NPA = 100% (98.5-100%)
Bordetella parapertussis: PPA = 98.0% (89.5-99.6%), NPA = 100% (98.5-100%)
Bordetella pertussis: PPA = 100% (92.9-100%), NPA = 100% (98.5-100%)
Chlamydia pneumoniae: PPA = 100% (92.9-100%), NPA = 100% (98.5-100%)
Influenza A H1N1 pdm09: PPA = 98.0% (89.5-99.6%), NPA = 100% (98.5-100%)
Mycoplasma pneumoniae: PPA = 98.0% (89.5-99.6%), NPA = 99.6% (97.8-99.9%)

Predicate Device(s): If the device was cleared using the 510(k) pathway, identify the Predicate Device(s) K/DEN number used to claim substantial equivalence and list them here in a comma separated list exactly as they appear in the text. List the primary predicate first in the list.

DEN200031

Reference Device(s): Identify the Reference Device(s) K/DEN number and list them here in a comma separated list exactly as they appear in the text.

Not Found

Predetermined Change Control Plan (PCCP) - All Relevant Information for the subject device only (e.g. presence / absence, what scope was granted / cleared under the PCCP, any restrictions, etc).

Not Found

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.

Summary

0

March 1, 2024

Image /page/0/Picture/1 description: The image shows the logo of the U.S. Food and Drug Administration (FDA). On the left is the Department of Health & Human Services logo. To the right of that is the FDA logo, which is a blue square with the letters "FDA" in white. To the right of the blue square is the text "U.S. FOOD & DRUG ADMINISTRATION" in blue.

Luminex Corporation Tara Viviani Sr. Director Molecular Regulatory Affairs 4088 Commercial Avenue Northbrook, Illinois 60062

Re: K233410

Trade/Device Name: LIAISON PLEX Respiratory Flex Assay 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, OEM, OOU, OTG, OZE, OZX, OZY, OZZ, OCC, NSU Dated: October 6, 2023 Received: October 6, 2023

Dear Tara Viviani:

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"

1

(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 OS 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 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-safetyreporting-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.

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-device-safety/medical-device-reportingmdr-how-report-medical-device-problems.

For comprehensive regulatory information about mediation-emitting products, including information about labeling regulations, please see Device Advice (https://www.fda.gov/medicaldevices/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-device-advice-comprehensive-regulatoryassistance/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).

Sincerely.

Joseph Briggs -S

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

Enclosure

2

Indications for Use

510(k) Number (if known) K233410

Device Name LIAISON PLEX Respiratory Flex Assay

Indications for Use (Describe)

The LIAISON PLEX Respiratory Flex (RSP Flex) Assay is a multiplexed qualitative test for the simultaneous in vitro detection and identification of multiple bacterial and viral nucleic acids in nasopharyngeal swabs (NPS) obtained from individuals with clinical signs and symptoms of respiratory tract infection, including SARS-CoV-2. The test is performed on the automated LIAISON PLEX System utilizing reverse transcription (RT), polymerase chain reaction (PCR), and array hybridization to detect specific nucleic acid gene sequences of the following organism types and subtypes:

Viruses: Adenovirus Human Coronavirus (HKU1, NL63, OC43, and 229E not differentiated) Human Enterovirus/Rhinovirus (not differentiated) Human Metapneumovirus, Influenza A Influenza A (subtype H1) Influenza A (subtype H3) Influenza B Parainfluenza 1 Parainfluenza 2 Parainfluenza 3 Parainfluenza 4 Respiratory Syncytial Virus Severe Acute Respiratory Syndrome Coronavirus (SARS-CoV-2)

Bacteria: Bordetella holmesii Bordetella parapertussis Bordetella pertussis Chlamydia pneumoniae Mycoplasma pneumoniae

Nucleic acids from the bacterial and viral organisms identified by this test are generally detectable in NPS specimens during the acute phase of infection. Detecting and identifying specific bacterial and viral nucleic acids from individuals exhibiting signs and symptoms of respiratory infection aids in the diagnosis of respiratory infection, if used in conjunction with other clinical, epidemiological, and laboratory findings. The results of this test should not be used as the sole basis for diagnosis, treatment, or patient management decisions.

Negative results in the presence of a respiratory illness may be due to infection with pathogens that are not detected by this test or due to lower respiratory that is not detected by an NPS specimen. Conversely, positive results do not rule out infection or co-infection with organisms not detected by the LIAISON PLEX Respiratory Flex (RSP Flex) Assay. The agent(s) detected may not be the definite cause of disease.

The use of additional laboratory testing (e.g., bacterial and viral culture, immunofluorescence, and radiography), may be necessary when evaluating a patient with possible respiratory tract infection.

3

X Prescription Use (Part 21 CFR 801 Subpart D)

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4

510(k) Summary

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

Preparation Date: 21-February-2024

A. 510(k) Number:

K233410

B. Purpose for Submission:

Traditional 510(k), New Device

C. Measurand:

Adenovirus, Bordetella holmesii, Bordetella parapertussis, Bordetella pertussis, Chlamydia pneumoniae, Human Coronavirus (HKU1, NL63, OC43, and 229E not differentiated), Human Enterovirus (Rhinovirus (not differentiated), Human Metapneumovirus, Influenza A (subtype H1), Influenza A (subtype H3), Influenza B, Mycoplasma pneumoniae, Parainfluenza 1, Parainfluenza 2, Parainfluenza 3, Parainfluenza 4, Respiratory Syncytial Virus (RSV), and Severe Acute Respiratory Syndrome Coronavirus (SARS-CoV-2) nucleic acid target sequences

D. Type of Test:

Qualitative Multiplexed Nucleic Acid Test that Utilizes Reverse Transcription, Real Time Polymerase Chain Reaction (PCR), and Array Hybridization.

E. Applicant:

Tara Viviani, Luminex Corporation 4088 Commercial Avenue Northbrook, IL 60062 (847) 400-9000

F. Proprietary and Established Names:

LIAISON PLEX® Respiratory Flex Assay

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G. Regulatory Information:

| Primary
Product

Code	Classification	Regulation Section
QOF	II	21 CFR 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	MI - Microbiology

H. Intended Use:

Intended use(s):

The LIAISON PLEX Respiratory Flex (RSP Flex) Assay is a multiplexed qualitative test for the simultaneous in vitro detection and identification of multiple bacterial and viral nucleic acids in nasopharyngeal swabs (NPS) obtained from individuals with clinical signs and symptoms of respiratory tract infection, including SARS-CoV-2. The test is performed on the automated LIAISON PLEX System utilizing reverse transcription (RT), polymerase chain reaction (PCR), and array hybridization to detect specific nucleic acid gene sequences of the following organism types and subtypes:

Viruses:

Adenovirus Human Coronavirus (HKU1, NL63, OC43, and 229E not differentiated) Human Enterovirus/Rhinovirus (not differentiated) Human Metapneumovirus, Influenza A Influenza A (subtype H1) Influenza A (subtype H3) Influenza B Parainfluenza 1 Parainfluenza 2 Parainfluenza 3 Parainfluenza 4 Respiratory Syncytial Virus Severe Acute Respiratory Syndrome Coronavirus (SARS-CoV-2)

Bacteria:

Bordetella holmesii Bordetella parapertussis Bordetella pertussis Chlamydia pneumoniae Mycoplasma pneumoniae

6

Nucleic acids from the bacterial and viral organisms identified by this test are generally detectable in NPS specimens during the acute phase of infection. Detecting and identifying specific bacterial and viral nucleic acids from individuals exhibiting signs and symptoms of respiratory infection aids in the diagnosis of respiratory infection, if used in conjunction with other clinical, epidemiological, and laboratory findings. The results of this test should not be used as the sole basis for diagnosis, treatment, or patient management decisions.

Negative results in the presence of a respiratory illness may be due to infection with pathogens that are not detected by this test or due to lower respiratory tract infection that is not detected by an NPS specimen. Conversely, positive results do not rule out infection or coinfection with organisms not detected by the LIAISON PLEX Respiratory Flex (RSP Flex) Assay. The agent(s) detected may not be the definite cause of disease.

The use of additional laboratory testing (e.g., bacterial and viral culture, immunofluorescence, and radiography), may be necessary when evaluating a patient with possible respiratory tract infection.

Indication(s) for use:

Same as intended use.

Special conditions for use statement(s):

For prescription use only.

For in vitro diagnostic use only

Special instrument requirements:

For use with LIAISON PLEX Systems only

l. Device Description:

The LIAISON PLEX® Respiratory Flex Assay is a multiplexed nucleic acid test system composed of the LIAISON PLEX Instrument, the LIAISON PLEX® System Software (preinstalled on the LIAISON PLEX Instrument), the LIAISON PLEX® Respiratory Flex Assay cartridge, and the LIAISON PLEX® Respiratory Flex Assay File. The LIAISON PLEX® Respiratory Flex Assay cartridge contains the reagents to perform nucleic acid extraction and purification, reverse transcription, PCR, and array hybridization. Specifically, the LIAISON PLEX® Respiratory Flex Assay detects bacteria and viruses from nasopharyngeal swab (NPS) specimens collected from individuals with signs and symptoms of respiratory infection.

The LIAISON PLEX System consists of a touchscreen user interface that includes the software for running and analyzing assay results, one to six processing/imaging LIAISON PLEX modules, and a handheld barcode reader. Each LIAISON PLEX module processes one sample at a time under the control of the LIAISON PLEX System software.

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LIAISON PLEX® automates the sample processing through analysis within a single cartridge. Processing steps include 1.) Sample Preparation: Nucleic acid extraction from organisms by chemical and mechanical means and isolation of nucleic acid on magnetic beads 2.) Target Amplification: Multiplex PCR and RT-PCR based amplification of extracted nucleic acid to generate target specific amplicons 3.) Hybridization: Amplicons hybridize with their target specific DNA probe arranged in a microarray format and that are attached to mediator and gold nanoparticles 4.) Analysis: Gold nanoparticles specifically bound to target amplicons are silver enhanced and the light scatter from microarray spot is measured and analyzed to confirm presence (Detected) or absence (not Detected) of a target.

The LIAISON PLEX Respiratory Flex Assay has the option of creating and processing results for custom panels using Flex® Software. Flex Software allows users to randomly select and group targets in tiers for result processing. Up to 7 targets may be selected for the initial test tier. After the first tier, each additional tier requires a specific number of credits. Flex™ credits allow the end-user to create custom panels and pay for a smaller subset of results tailored to the individual patient's clinical presentation. Alternatively, a laboratory may choose the fixed price option where all target results are processed at the same time.

Substantial Equivalence Information: J.

Predicate device name(s):

BioFire Respiratory Panel 2.1 (RP2.1)

Predicate 510(k) number(s):

DEN200031

Comparison with predicate:

The following table compares Luminex's LIAISON PLEX® Respiratory Flex Assay to the BioFire Respiratory Panel 2.1 (RP2.1) (DEN200031).

Comparison to Predicate
-------------------------	--	--	--

| Comparison to
Predicate Device | Predicate Device:
BioFire Respiratory Panel 2.1 (RP2.1)
(DEN200031) | Candidate Device:
LIAISON PLEX® Respiratory Flex
Assay |
|----------------------------------------|-------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------|---------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------|
| Product Code | QOF | QOF |
| Regulation Number | 21 CFR 866.3981 | 21 CFR 866.3981 |
| Comparison to
Predicate Device | Predicate Device:
BioFire Respiratory Panel 2.1 (RP2.1)
(DEN200031) | Candidate Device:
LIAISON PLEX® Respiratory Flex
Assay |
| Organisms Detected | Adenovirus, Coronavirus 229E,
Coronavirus HKU1, Coronavirus NL63,
Coronavirus OC43, Severe Acute
Respiratory Syndrome Coronavirus
(SARS-CoV-2), Human
Metapneumovirus, Human
Rhinovirus/Enterovirus, Influenza A,
including subtypes H1, H1-2009, and
H3, Influenza B, Parainfluenza Virus 1,
Parainfluenza Virus 2, Parainfluenza
Virus 3, Parainfluenza Virus 4,
Respiratory Syncytial Virus, Bordetella
parapertussis (IS1001), Bordetella
pertussis (ptxP), Chlamydia
pneumoniae, and Mycoplasma
pneumoniae | Adenovirus, Bordetella holmesii,
Bordetella parapertussis, Bordetella
pertussis, Chlamydia pneumoniae,
Coronavirus 229E, Coronavirus
HKU1, Coronavirus NL63,
Coronavirus OC43,
Enterovirus/Rhinovirus, Human
Metapneumovirus, Influenza A,
Influenza A (subtype H1), Influenza A
(subtype H3), Influenza B,
Mycoplasma pneumoniae,
Parainfluenza 1, Parainfluenza 2,
Parainfluenza 3, Parainfluenza 4,
Respiratory Syncytial Virus, and
SARS-CoV-2 |
| Measurand | Nucleic acid from Organisms detected | Nucleic acid from Organisms detected |
| Intended Use | The BioFire Respiratory Panel 2.1 (RP2.1)
is a PCR-based multiplexed nucleic acid
test intended for use with the BioFire
FilmArray 2.0 or BioFire FilmArray Torch
systems for the simultaneous qualitative
detection and identification of multiple
respiratory viral and bacterial nucleic
acids in nasopharyngeal swabs (NPS)
obtained from individuals suspected of
respiratory tract infections, including
COVID-19.
The following organism types and
subtypes are identified using the BioFire
RP2.1: | The LIAISON PLEX Respiratory Flex
(RSP Flex) Assay is a multiplexed
qualitative test for the simultaneous in
vitro detection and identification of
multiple bacterial and viral nucleic
acids in nasopharyngeal swabs (NPS)
obtained from individuals with clinical
signs and symptoms of respiratory
tract infection, including SARS-CoV-2.
The test is performed on the
automated LIAISON PLEX System
utilizing reverse transcription (RT),
polymerase chain reaction (PCR), and
array hybridization to detect specific
nucleic acid gene sequences of the
following organism types and
subtypes: |
| | Adenovirus,
Coronavirus 229E,
Coronavirus HKU1,
Coronavirus NL63,
Coronavirus OC43,
Severe Acute Respiratory Syndrome
Coronavirus (SARS-CoV-2),
Human Metapneumovirus,
Human Rhinovirus/Enterovirus,
Influenza A, including subtypes H1, H1-
2009, and H3,
Influenza B,
Parainfluenza Virus 1,
Parainfluenza Virus 2,
Parainfluenza Virus 3,
Parainfluenza Virus 4,
Respiratory Syncytial Virus,
Bordetella parapertussis (IS1001), | Viruses:
Adenovirus
Human Coronavirus (HKU1, NL63,
OC43, and 229E not differentiated)
Human Enterovirus/Rhinovirus (not
differentiated)
Human Metapneumovirus,
Influenza A
Influenza A (subtype H1)
Influenza A (subtype H3)
Influenza B
Parainfluenza 1
Parainfluenza 2
Parainfluenza 3
Parainfluenza 4
Respiratory Syncytial Virus
Severe Acute Respiratory Syndrome
Coronavirus (SARS-CoV-2) |
| Comparison to
Predicate Device | Predicate Device:
BioFire Respiratory Panel 2.1 (RP2.1)
(DEN200031) | Candidate Device:
LIAISON PLEX® Respiratory Flex
Assay |
| | Bordetella pertussis (ptxP),
Chlamydia pneumoniae, and
Mycoplasma pneumoniae
Nucleic acids from the respiratory viral
and bacterial organisms identified by this
test are generally detectable in NPS
specimens during the acute phase of
infection. The detection and identification
of specific viral and bacterial nucleic acids
from individuals exhibiting signs and/or
symptoms of respiratory infection is
indicative of the presence of the identified
microorganism and aids in the diagnosis
of respiratory infection if used in
conjunction with other clinical and
epidemiological information. The results of
this test should not be used as the sole
basis for diagnosis, treatment, or other
patient management decisions.
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
BioFire RP2.1 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. | Bacteria:
Bordetella holmesii
Bordetella parapertussis
Bordetella pertussis
Chlamydia pneumoniae
Mycoplasma pneumoniae
Nucleic acids from the bacterial and
viral organisms identified by this test
are generally detectable in NPS
specimens during the acute phase of
infection. Detecting and identifying
specific bacterial and viral nucleic
acids from individuals exhibiting signs
and symptoms of respiratory infection
aids in the diagnosis of respiratory
infection, if used in conjunction with
other clinical, epidemiological, and
laboratory findings. The results of this
test should not be used as the sole
basis for diagnosis, treatment, or
patient management decisions.
Negative results in the presence of a
respiratory illness may be due to
infection with pathogens that are not
detected by this test or due to lower
respiratory tract infection that is not
detected by an NPS specimen.
Conversely, positive results do not rule
out infection or co-infection with
organisms not detected by the
LIAISON PLEX Respiratory Flex (RSP
Flex) Assay. The agent(s) detected
may not be the definite cause of
disease.
The use of additional laboratory testing
(e.g., bacterial and viral culture,
immunofluorescence, and radiography),
may be necessary when evaluating a
patient with possible respiratory tract
infection. |
| Automated System
(Sample to Answer) | Automated | Same |
| Instrumentation | BioFire® FilmArray® 2.0 or BioFire®
FilmArray® Torch Systems | LIAISON PLEX® |
| Sample Types | Nasopharyngeal Swab (NPS) Specimens | Same |
| Technological Principles | Highly multiplexed nested nucleic acid
amplification with melt analysis. | Highly multiplexed nucleic acid PCR
and RT-PCR test with microarray
detection |
| Internal Controls | Two controls are included in each reagent
pouch to control for sample processing | Multiple internal controls contained in
the cartridge monitor sample
processing and RT and PCR |
| Comparison to
Predicate Device | Predicate Device:
BioFire Respiratory Panel 2.1 (RP2.1)
(DEN200031) | Candidate Device:
LIAISON PLEX® Respiratory Flex
Assay |
| | and both stages of PCR and melt analysis. | functions. |
| Bordetella Species
Detected | Bordetella parapertussis
Bordetella pertussis | Bordetella parapertussis Bordetella pertussis Bordetella holmesii |
| Human Coronavirus
Result Reporting | Each target human coronavirus species
(i.e., HKU1, OC43, 229E, NL63) is
reported independently. | The human coronavirus target species
(i.e., HKU1, OC43, 229E, NL63) are
not differentiated. |
| Influenza A Subtyping | Influenza A subtypes H1, H1-2009, and H3
detected/reported. | Influenza A subtypes H1 and H3
detected/reported. |
| Time to Result | ~45 minutes | ~2 hours |

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9

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K. Standards/Guidance Documents Referenced:

Standards

. CLSI. User Protocol for Evaluation of Qualitative Test Performance; Approved Guideline -Second Edition. CLSI document EP12-A2. Wayne, PA: Clinical and Laboratory Standards Institute; 2008.
CLSI. Evaluation of Stability of In Vitro Diagnostic Reagents; Approved Guideline. CLSI document EP25-A. Wayne, PA: Clinical and Laboratory Standards Institute; 2009.
. CLSI. Interference Testing in Clinical Chemistry. 3rd ed. CLSI guideline EP07. Wayne, PA: Clinical and Laboratory Standards Institute; 2018.
ISO 14971:2019 Medical devices - Application of risk management to medical devices
IEC 62366-1:2015 Medical devices - Part 1: Application of usability engineering to medical devices
ISO 62304:2006 Medical device software - Software life-cycle processes
ISO 15223-1:2016: Medical Devices - Symbols to be used with medical device labels, labeling and information to be supplied - Part 1: General requirements
. IEC 61010-1 Ed. 3.0 2010: Safety requirements for electrical equipment for measurement, control, and laboratory use - Part 1: General requirements
EN 61010-2-101:2002/IEC 61010-2-101:2015: Safety requirements for electrical equipment for measurement, control and laboratory use - Part 2-101: Particular requirements for in vitro diagnostic (IVD) medical equipment.
IEC 60601-1-2:2014 (Edition 4.0): Medical electrical equipment - Part 1-2: General requirements for basic safety and essential performance - Collateral Standard: Electromagnetic disturbances - Requirements and tests
. ISO 13485:2016/EN ISO 13485:2016; Medical devices - Quality Management System -Requirements for regulatory purposes
ISO 20916:2019; In vitro diagnostic medical devices. Clinical performance studies using specimens from human subjects. Good study practice
. EN ISO 18113-1:2011; In vitro diagnostic medical devices - Information supplied by the manufacturer (labeling). Terms, definition and general requirements
. EN ISO 18113-2:2011; In vitro diagnostic medical devices - Information supplied by the manufacturer (labeling) – Part 2: In vitro diagnostic reagents for professional use

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. EN ISO 18113-3:2011; In vitro diagnostic medical devices - Information supplied by the manufacturer (labeling) – Part 3: In vitro diagnostic instruments for professional use
EN ISO 23640:2015; In vitro diagnostic medical devices - Evaluation of stability of in vitro
. IEC 61326-1:2012; Electrical equipment for measurement control and laboratory use -EMC requirements - Part 1: General requirements
. EN 61326-2-6:2006/IEC 61326-2-6:2012; Electrical equipment for measurement control and laboratory use - EMC requirements - Part 2-6: Particular requirements - In vitro diagnostic (IVD) medical equipment

Special Controls

Class II Special Controls as per 21 CFR 866.3981 ●

Guidance Documents

Electronic Submission Template for Medical Device 510(k) Submissions Guidance for Industry ● and Food and Drug Administration Staff (October 2, 2023).
. Respiratory Viral Panel Multiplex Nucleic Acid Assay - Class II Special Controls Guidance for Industry and FDA Staff (October 9, 2009).
. Content of Premarket Submissions for Device Software Functions - Guidance for Industry and Food and Drug Administration Staff (June 14, 2023).
. Cybersecurity in Medical Devices: Quality System Considerations and Content of Premarket Submissions - Guidance for Industry and Food and Drug Administration Staff (September 23, 2023).
Statistical Guidance on Reporting Results from Studies Evaluating Diagnostic Tests - Guidance for Industry and FDA Staff (March 13, 2007).

L. Test Principle:

The LIAISON PLEX® Respiratory Flex Assay is performed on nasopharyngeal swab (NPS) specimens collected in Copan Universal Transport Medium™ or BD™ Universal Viral Transport Media. The system consists of an instrument and a single-use, disposable test cartridge and a transfer pipette. The user loads a portion of the sample into the sample port of the LIAISON PLEX Respiratory Flex Assay Cartridge. Next, the user sets up the sample order on the LIAISON PLEX System by first entering the sample information or scanning the barcode ID located on the sample tube, then scanning the barcode ID located on the test cartridge. Last, the user inserts the test cartridge into the processing module to initiate the test. The LIAISON PLEX System identifies the assay being run and automatically initiates the proper testing protocol to process the sample, analyze the data, and generate test results.

The LIAISON PLEX System automates the LIAISON PLEX Respiratory Flex Assay sample analysis through the following steps: a) Sample Preparation: Nucleic acid extraction via mechanical and chemical cell lysis and magnetic bead- based nucleic acid isolation of prepared specimens obtained from patients; b) Target Amplification: Multiplex PCR- and RT-PCR-based amplification of the extracted nucleic acids to generate target-specific amplicons; c) Hybridization: Amplicons hybridize to target-specific capture DNA on a microarray format, and target-specific mediator and gold nanoparticle probes hybridize to captured amplicons; d) Signal Analysis: Gold nanoparticle probes bound specifically to target-containing spots in the microarray are silver-enhanced, and light scatter from the spots is measured and further

12

analyzed to determine the presence (Detected) or absence (Not Detected) of a target.

M. Performance Characteristics:

1. Analytical performance:
- a. Precision/Reproducibility:

Within Laboratory Precision

Within laboratory precision of the LIAISON PLEX Respiratory Flex Assay was evaluated by testing three lots of LIAISON PLEX Respiratory Flex Assay cartridges at a single site over five non-consecutive days. Three target concentrations were prepared and tested. Targets consisted of a negative sample and a positive sample comprised of five targets (B. pertussis, Adenovirus, influenza B, human metapneumovirus, and SARS-CoV-2). All positive samples were diluted in a simulated NPS matrix to a low positive concentration (1.5X LoD) and a moderate positive concentration (5X LoD). Targets were randomized and blinded to the operators in an order that each operator tested each target (negative, 1.5X LoD) in triplicate on each of the testing days. Qualitative results of the within laboratory precision study are summarized in Table 1.

| Target | Panel Concentration | % Positive | % Agreement with
Expected Results
(95% CI) |
|-------------------------|----------------------------|---------------|--------------------------------------------------|
| Bordetella
pertussis | Low Positive (1.5X LoD) | 93.3% (42/45) | 93.3% (82.1-97.7%) |
| | Moderate Positive (5X LoD) | 100% (45/45) | 100% (92.1-100%) |
| | Negative | 0% (0/45) | 100% (92.1-100%) |
| Adenovirus | Low Positive (1.5X LoD) | 97.8% (44/45) | 97.8% (88.4-99.6%) |
| | Moderate Positive (5X LoD) | 97.8% (44/45) | 97.8% (88.4-99.6%) |
| | Negative | 0% (0/45) | 100% (92.1-100%) |
| Influenza B | Low Positive (1.5X LoD) | 100% (45/45) | 100% (92.1-100%) |
| | Moderate Positive (5X LoD) | 100% (45/45) | 100% (92.1-100%) |
| | Negative | 0% (0/45) | 100% (92.1-100%) |
| hMPV | Low Positive (1.5X LoD) | 100% (45/45) | 100% (92.1-100%) |
| | Moderate Positive (5X LoD) | 97.8% (44/45) | 97.8% (88.4-99.6%) |
| | Negative | 0% (0/45) | 100% (92.1-100%) |
| SARS-CoV-2 | Low Positive (1.5X LoD) | 100% (45/45) | 100% (92.1-100%) |
| | Moderate Positive (5X LoD) | 100% (45/45) | 100% (92.1-100%) |
| | Negative | 0% (0/45) | 100% (92.1-100%) |

Table 1 - Within Laboratory Precision

Note: Results are shown only for the intended targets. Panel members co-spiked with 5 different targets 3 lots and tested over 5 non-consecutive days (45 total replicates).

Reproducibility

Reproducibility of the LIAISON PLEX Respiratory Flex Assay was evaluated by testing one lot of LIAISON PLEX Respiratory Flex Assay cartridges with two operators at each of three sites over five non-consecutive days. Three target concentrations were prepared and tested across all

13

sites and operators to evaluate site-to-site reproducibility. Targets consisted of a negative sample and a positive sample comprised of five assay targets (B. pertussis, Adenovirus, influenza B, human metapneumovirus, and SARS-CoV-2). All positive samples were diluted in a simulated NPS matrix to a low positive concentration (1.5X LoD) and a moderate positive concentration (5X LoD). Targets were randomized and blinded to the operators in an order that each operator tested each target (negative, 1.5X LoD, and 5X LoD) in triplicate on each of the testing days. Qualitative results of the reproducibility study are summarized in Table 2.

		% Agreement with Expected Results
Organism	Target
Concentration	Site 1	Site 2	Site 3	All Sites
(95% Confidence)
Adenovirus	Low Positive
(1.5X LoD)	96.7%
(29/30)	100%
(30/30)	96.7%
(29/30)	97.8% (88/90)
(92.3% - 99.4%)
	Moderate Positive
(5X LoD)	100%
(30/30)	100%
(30/30)	100%
(30/30)	100% (90/90)
(95.9% - 100%)
Bordetella pertussis	Low Positive
(1.5X LoD)	93.3%
(28/30)	100%
(30/30)	96.7%
(29/30)	96.7% (87/90)
(90.7% - 98.9%)
	Moderate Positive
(5X LoD)	100%
(30/30)	100%
(30/30)	100%
(30/30)	100% (90/90)
(95.9% - 100%)
Influenza B	Low Positive
(1.5X LoD)	93.3%
(28/30)	100%
(30/30)	96.7%
(29/30)	96.7% (87/90)
(90.7%- 98.9%)
	Moderate Positive
(5X LoD)	100%
(30/30)	100%
(30/30)	100%
(30/30)	100% (90/90)
(95.9% - 100%)
Human
Metapneumovirus	Low Positive
(1.5X LoD)	90.0%
(27/30)	100%
(30/30)	93.3%
(28/30)	94.4% (85/90)
(87.6% - 97.6%)
	Moderate Positive
(5X LoD)	100%
(30/30)	93.3%
(28/30)	100%
(30/30)	97.8% (88/90)
(92.3% - 99.4%)
SARS-CoV-2	Low Positive
(1.5X LoD)	96.7%
(29/30)	100%
(30/30)	100%
(30/30)	98.9% (89/90)
(93.9% - 99.8%)
	Moderate Positive
(5X LoD)	100%
(30/30)	100%
(30/30)	100%
(30/30)	100% (90/90)
(95.9% - 100%)
Negative NPS	Negative	100%
(30/30)	100%
(30/30)	100%
(30/30)	100% (90/90)
(95.9% - 100%)

Table 2. Reproducibility Results

b. Linearity/assay reportable range:

Not applicable. The LIAISON PLEX® Respiratory Flex Assay is a qualitative assay.

Traceability, Stability, Expected values (controls, calibrators, or methods): C.

Controls:

Each LIAISON PLEX Respiratory Flex Assay cartridge includes internal controls (extraction control, amplification control, and hybridization control) to ensure performance of sample preparation, amplification, and detection. Extraction control is automatically added to the sample prior to initiation of sample preparation and assesses extraction, nucleic acid recovery, amplification of RNA targets, and detection. Additionally, an amplification control present in the lyophilized PCR

14

master mix, serves as an independent amplification and detection control for DNA targets. Finally, a post-amplification hybridization control serves as an indicator of successful hybridization. Internal control results are reported as Pass, Fail, or N/A on the printed reports (see Table 3 for detailed explanations of each control result). Internal controls must either (1) generate a signal above threshold in each internal reaction for the system to report a valid test result, or (2) the amplification or extraction control result can be below the signal threshold if a DNA or RNA target pathogen is detected, respectively.

If the Test Result is "No Call" for reasons other than failure of internal controls, the Internal Control Result is reported as "N/A" and the user should repeat the test with a new cartridge. For additional assistance regarding assay failures unrelated to internal controls, please refer to Chapter 8 (Troubleshooting Unexpected Results/Failures) of the LIAISON PLEX® System User Manual.

| Internal

Control Result	Explanation	Suggested Action
Pass	The hybridization control was detected, indicating successful
hybridization.
The amplification control was detected, indicating successful
amplification.
The extraction control was detected, indicating successful
extraction.	Review and report
results
N/A	The hybridization control was detected, indicating successful
hybridization.
A DNA pathogen target was detected, indicating successful
amplification. If a DNA pathogen target is detected, the
amplification control result is ignored.
The extraction control was detected, indicating successful
extraction.	Review and report
results
N/A	The hybridization control was detected, indicating successful
hybridization.
The amplification control was detected, indicating successful
amplification.
A RNA pathogen target was detected, indicating successful
extraction. If a RNA pathogen target is detected, the extraction
control result is ignored.	Review and report
results
Fail	The hybridization control was not detected indicating hybridization
was not successful.
The amplification control, or a DNA pathogen was detected,
indicating successful amplification.
The extraction control, or a RNA pathogen was detected, indicating
successful extraction.	Repeat test with a
new cartridge

Table 3. Interpretation of Controls on the LIAISON PLEX Respiratory Flex Assay Report

15

| Internal

Control Result	Explanation	Suggested Action
Fail	The hybridization control was detected indicating successful
hybridization.
The amplification control, or a DNA pathogen was not detected,
indicating amplification was not successful.
The extraction control, or a RNA pathogen was detected, indicating
successful extraction.	Repeat test with a
new cartridge
Fail	The hybridization control was detected indicating successful
hybridization.
The amplification control, or a DNA pathogen was detected,
indicating successful amplification.
The extraction control, or a RNA pathogen, was not detected,
indicating extraction was not successful.	Repeat test with a
new cartridge

External Controls

Positive and negative external controls should be tested with each new lot or shipment of reagents, or monthly, (whichever occurs first), or in accordance with updated local, regional, state, and/or federal guidelines. Positive and negative external controls are not provided with the LIAISON PLEX Respiratory Flex Assay. Verified negative nasopharyngeal swab (NPS) specimens can be used as the negative control. Previously characterized positive samples or verified negative NPS specimens spiked with well characterized organisms may be used as the external positive control. External controls should be used in accordance with laboratory protocols and in accordance with local, state, and federal accrediting organizations, as applicable.

Stability:

Specimen Stability

Contrived specimen stability at room temperature (15°C - 30°C), refrigerated (2°C - 8°C), and frozen (90% homology), 99.9% (112,034/112,056) of influenza A sequences are expected to be detected, 98.9% (53,778/54,364) of influenza A H1 sequences are expected to be detected, 99.9% (104,315/104,428) of influenza A H3 sequences are expected to be detected, and 99.9% (26,433/26,470) of influenza B sequences are expected to be detected.

| Reportable
Target | Target Gene | of Sequences in
Alignment | # of Sequences with
Percent Oligo Identify

90% |
|----------------------|-----------------------------|------------------------------|-------------------------------------------------------|
| Influenza A | Matrix protein (MP) | 112,056 | 112,0341 |
| Influenza A H1 | HA | 54,364 | 53,778 |
| Influenza A H3 | HA | 104,428 | 104,315 |
| Influenza B | Non-structural protein (NS) | 26,470 | 26,4332 |

Table 8. Influenza In Silico Inclusivity Results

1 Analysis included influenza A subtype H0, H1, H3, H5, H7, H9, and H10 strains.

2 Analysis included 17,509 Victoria lineage strains, 8,167 Yamagata lineage strains of unknown lineage.

26

f. Analytical specificity
Cross-Reactivity and Microbial Interference:

Off-Panel Cross Reactivity

Laboratory (Wet Testing)

Cross-reactivity was assessed in the laboratory by wet testing 60 off-panel viral, fungal, and bacterial organisms that may be found in a respiratory tract clinical specimen. The potential cross-reacting organisms were spiked into simulated NPS matrix that was negative for all targets on the assay. Bacteria were tested at concentrations ≥ 1x10° CFU/mL (or equivalent) and viruses were tested at ≥ 1x105 TCID50/mL (or equivalent), or the highest available concentration. For Mycobacterium tuberculosis, genomic DNA (quantified in ng/uL) was evaluated to minimize pathogenic exposure to the test operator.

Of the 60 potential cross-reacting off-panel organisms tested in the laboratory (Table 9), 59 organisms yielded negative results for all targets and are considered non-reactive with the LIAISON PLEX Respiratory Flex Assay. Mycoplasma qenitalium cross-reacted with the Mycoplasma pneumoniae assay at a concentration of 4x10 CCU/mL. No cross-reactivity was observed when Mycoplasma genitalium was tested at a lower concentration of 1x10f CCU/mL.

Organism	Conc./Unit	Organism	Conc./Unit
Acinetobacter baumannii	1x106 CFU/mL	Legionella pneumophila	4x105 CFU/mL3
Aspergillus flavus	1x106 CFU/mL	Listeria innocua	1x106 CFU/mL
Aspergillus fumigatus	4x105 CFU/mL3	Listeria monocytogenes	1x106 CFU/mL
Bordetella avium	1x106 CFU/mL	Measles	1x105 TCID50/mL
Bordetella bronchiseptica	1x106 CFU/mL	MERS-CoV	NA2
Bordetella hinzii	1x106 CFU/mL	Moraxella catarrhalis	1x106 CFU/mL
Bordetella petrii	1x106 CFU/mL	Mumps Virus	1x105 TCID50/mL
Bordetella trematum	1x106 CFU/mL	Mycobacterium
tuberculosis
(H37Rv gDNA)	2.88 ng/uL
Bordetella parapertussis
Bpp5 (synthetic DNA)1	1x106 copies/mL	Mycoplasma
genitalium3	4x106 CCU/mL
			1x106 CCU/mL
			4x105 CCU/mL
Candida albicans	1x106 CFU/mL	Mycoplasma hominis	1x106 CFU/mL
Candida glabrata	1x106 CFU/mL	Nasal Wash (pooled)	NA2
Chlamydia trachomatis
Serovar D	1x106 IFU/mL	Neisseria elongata	1x106 CFU/mL
Coronavirus-SARS	NA2	Neisseria gonorrhoeae	1x106 CFU/mL
Corynebacterium
diphtheriae	1x106 CFU/mL	Neisseria lactamica	1x106 CFU/mL
Corynebacterium
pseudodiphtheriticum	1x106 CFU/mL	Neisseria meningitidis	1x106 CFU/mL
Corynebacterium striatum	1x106 CFU/mL	Neisseria mucosa	1x106 CFU/mL
Cytomegalovirus	1x105 TCID50/mL	Neisseria sicca	1x106 CFU/mL
Epstein Barr Virus	1x105 copies/mL	Pneumocystis jiroveci	1x106 CFU/mL

Table 9. Organisms Tested for Potential Cross-Reactivity (Off-Panel)

27

Organism	Conc./Unit	Organism	Conc./Unit
Escherichia coli	1x106 CFU/mL	Proteus vulgaris	1x106 CFU/mL
Fluoribacter bozemanae	4x106 CFU/mL	Pseudomonas
aeruginosa	1x106 CFU/mL
Fusobacterium
necrophorum	1x106 CFU/mL	Serratia marcescens	1x106 CFU/mL
Haemophilus influenzae	1x106 CFU/mL	Staphylococcus aureus	1x106 CFU/mL
Haemophilus
parainfluenzae	1x106 CFU/mL	Staphylococcus
epidermidis	1x106 CFU/mL
Herpes Simplex Virus
Type 1	1x105 TCID50/mL	Staphylococcus
haemolyticus	1x106 CFU/mL
Klebsiella pneumoniae	1x106 CFU/mL	Streptococcus
agalactiae	1x106 CFU/mL
Lactobacillus acidophilus	1x106 CFU/mL	Streptococcus
pneumoniae	1x106 CFU/mL
Lactobacillus plantarum	1x106 CFU/mL	Streptococcus
pyogenes	1x106 CFU/mL
Legionella anisa	1x106 CFU/mL	Streptococcus
salivarius	1x106 CFU/mL
Legionella feeleii	1x106 CFU/mL	Ureaplasma
urealyticum	1x106 CCU/mL
Legionella longbeachae	1x106 CFU/mL	Varicella-Zoster Virus	2.34x104
TCID50/mL3

CFU = Colony Forming Units; CCU = Colony Changing Units; TCID50 = Median Tissue Culture Infectious Dose.

14 portion of the B. parapertussis Bpp5 genome was identified by in-silico analysis as potentially cross-reactive with B. pertussis. Synthetic DNA was tested that matched the region of high homology in the assay. Testing was included in the off-panel testing since the targeted sequence was not expected to be detected as B. parapertussis by the assay.

2No concentration provided by the supplier.

3The highest possible concentration was tested.

In Silico Cross-Reactivity

In silico analysis of assay specificity/exclusivity was performed by conducting a BLAST comparison of the assay's oligos sequences to the GenBank nt sequence database, as of July 14, 2023. Sequences for 83 off-panel organisms (68 bacteria/fungi and 15 viruses) that can be found in a respiratory specimen were included. Additionally, sequences for all on-panel organisms were included to evaluate intra-panel cross-reactivity. A summary of the results from the analysis is provided in Table 10. The LIAISON PLEX Respiratory Flex assays were shown to be specific for their respective analytes with the following exceptions:

Cross-reaction of the Adenovirus assays with closely related Adenovirus G (serotype 52) strains.
Cross-reaction of the SARS-CoV-2 assays with closely related bat and pangolin coronavirus sequences;
Cross-reaction of the B. parapertussis assay with strains of B. bronchiseptica that carry IS1001;
Cross-reaction of the influenza A H1 subtyping assay with 3 swine H3N2 strains and 1 avian H6N1 strain;
Cross-reaction of the influenza A H3 subtyping assay with 59 swine H1N1 and swine H1N2 strains, 1 duck H5N2 strain, 1 ostrich H7N1 strain, 1 avian H7N9 strain, 1 avian H8N4 strain, and 1 avian H11N9 strain.

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Reportable Target	Predicted Cross-Reaction
Adenovirus	Adenovirus G (serotype 52) - strains
SARS-CoV-2	Bat coronavirus and Bat SARS-like coronavirus (accessions MG772933,
MG772934, and MN996532)1
Bordetella parapertussis	Bordetella bronchiseptica containing IS1001 element (accessions JX013523
to JX013527 and CP022962)²
Influenza A H1	H5N1 (accession CY110922)3;
swine H3N2 (accessions KM110061, KM110062, KM110063, and
OM935891);
avian H6N1 (accession OP888980)
Influenza A H3	swine H1N1 and swine H1N2 – 59 strains;
duck H5N2 (accession OK103962);
ostrich H7N1 (accession AF202244);
avian H7N9 (accession KP413675);
avian H8N4 (accession OK103964);
avian H11N9 (accession OK103956)

Table 10. Organisms Predicted by In Silico Analyses to Cross-React with the Respiratory Flex Assay.

1 is unlikely that Bat coronavirus and Bat SARS-like coronavirus would be present in human clinical NPS specimens; but if present, the cross-reactive product(s) produced by the LIAISON PLEX Respiratory Flex Assay will be reported as SARS-CoV-2.

²The LIAISON PLEX Respiratory Flex Assay contains primers designed to target the B. parapertussis IS1001 insertion sequence. Some strains of Bordetella bronchiseptica, which is rarely isolated from humans, carry the same sequence that is targeted by LIALSON PLEX Respiratory Flex primers for B. parapertussis. If present, the LIAISON PLEX Respiratory Flex Assay will report these specimens as B. parapertussis.

³This H5N1 human strain sequence containing H1N1 sequence fragments. Therefore, detection of this sequence by the H1 oligos is not considered a cross-reaction.

On-Panel Cross Reactivity

Potential intra-panel cross-reactivity was evaluated using 28 on-panel organisms. The potential cross-reacting on-panel organisms were spiked into simulated NPS matrix that was negative for all targets on the assay. Bacteria were tested at concentrations ≥ 1x10 CFU/mL (or equivalent) and viruses were tested at ≥ 1x105 TCID50/mL (or equivalent), or the highest available concentration.

All 28 on-panel organisms tested (Table 11) yielded the expected on-panel result and did not cross-react with other target assays of the LIAISON PLEX Respiratory Flex Assay.

Organism	Conc./Unit
Adenovirus	1x105 TCID50/mL
Bordetella holmesii	1x106 CFU/mL
Bordetella parapertussis	1x106 CFU/mL
Bordetella pertussis	1x106 CFU/mL
Chlamydia pneumoniae	1x106 IFU/mL
Human Coronavirus 229E	1x105 TCID50/mL
Human Coronavirus HKU1	6.62x104 copies/mL
Human Coronavirus NL63	1x105 TCID50/mL
Human Coronavirus OC43	1x105 TCID50/mL

Table 11. Organisms Tested for Potential Cross-Reactivity (On-Panel)

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Conc./Unit Organism Echovirus 1x105 TCID50/mL (Enterovirus/Rhinovirus) Human Metapneumovirus 1x105 TCID50/mL Influenza B (Washington/02/2019/Victoria 1x105 TCID50/mL Lineage) Influenza B (Phuket/3073/13/Yamagata 1x105 TCID50/mL Lineage) 1x106 CCU/mL Mycoplasma pneumoniae Parainfluenza 1 1x105 TCID50/mL Parainfluenza 2 1x105 TCID50/mL Parainfluenza 3 1x108 TCID50/mL Parainfluenza 4 1x105 TCID50/mL RSV A 1x105 PFU/mL RSV B 1x10> TCID50/mL Influenza A H1N1 1x105 TCID50/mL Influenza A H3N2 1x10 TCID50/mL Influenza A H5N11 2x107 copies/mL Influenza A H7N21 6x106 copies/mL Influenza A H7N71 2x107 copies/mL Influenza A H9N21 4x108 copies/mL Influenza A H1N2² 3x107 copies/mL 1x105 TCID50/mL SARS-CoV-2

LIAISON PLEX® Respiratory Flex Assay Traditional 510(k) Submission

CFU = Colony Forming Units; IFU = Inclusion Forming Units; TCID50 = Median Tissue Culture Infectious Dose; PFU = Plaque Forming Units

"These influenza A non-subtype H1/H3 strains are expected to be inclusive to the influenza A matrix target, only (i.e., are expected to be reported as influenza A positive, subtype H3 negative). All were negative for both subtype H1 and subtype H3, as anticipated.

²This influenza A H1N2 strain is expected to be influenza A matrix target and influenza A subtype H1 target. This strain was positive for both the influenza A matrix target and influenza A subtype H1 target, as expected.

Microbial Interference

The impact of 16 potentially interfering non-panel microbial organisms commonly found in nasopharyngeal swab samples were tested (Table 12) in the presentative assay panel targets. Each potentially interfering non-panel organism was tested at a high concentration of ≥ 1x10° CFU/mL (or equivalent) for bacteria and 1x107 TCIDsg/mL (or equivalent) for viruses, or the highest concentration available, in the presence of a multi-analyte panel consisting of five assay targets (B. pertussis, Adenovirus, influenza B, human metapneumovirus, and SARS-CoV-2) at a low concentration (3X LoD).

None of the potentially interfering organisms tested at high concentration interfered with detection for panel targets at a low concentration, except for Streptococcus pyogenes at 1x10° CFU/mL and Legionella pneumophilia at 4x105 CFU/mL, which both interfered with detection of Adenovirus in 1 of 6 replicates (83.3% positivity).

30

Candida albicans	Pseudomonas aeruginosa
Corynebacterium diphtheriae	Streptococcus pneumoniae
Cytomegalovirus	Streptococcus pyogenes
Haemophilus influenzae	SARS1
Herpes Simplex Virus 1	Legionella pneumophilia2
MERS1	Measles Virus
Neisseria meningitidis	Moraxella catarrhalis
Staphylococcus aureus	Mumps

Table 12. Organisms Tested for Potential Microbial Interference

'No concentration was provided for material. Material was tested at the highest available concentration by diluting stock material directly into representative multi-analyte panel preparation.

2Tested at the highest available concentration of 4x105 CFU/mL

Interfering Substances:

The potential inhibitory effect of non-microbial substances (endogenous and exogenous) expected to be found in nasopharyngeal swab (NPS) specimens or introduced during sample handling, were evaluated for the LIAISON PLEX Respiratory Flex Assay. Potential interference from 36 interfering substances were evaluated in the presence and absence of a multi-analyte panel of targets (Table 13). Each interfering substance tested was diluted to a clinically relevant concentration and tested in the presence of a positive and negative target in triplicate. The positive target was a multianalyte panel consisting of five assay targets (B. pertussis, Adenovirus, influenza B, human metapneumovirus, and SARS-CoV-2) at a low concentration (3X LoD), while the negative target was a simulated NPS matrix. Interference was observed for the substances/concentrations shown in Table 14.

Substance/Class	Description/Active Ingredient	Concentration Tested
Nasal Corticosteroid	Beclomethasone dipropionate	25 µg/mL
Anesthetic	Benzocaine	10% w/v
Nasal Corticosteroid	Budesonide	$3.4x10^{-2}$ µmol/L
Nasal Corticosteroid	Dexamethasone	30.6 µmol/L
Nasal Corticosteroid	Flunisolide	25 µg/mL
FLONASE Sensimist Allergy Relief	Fluticasone furoate	$2.84x10^{-3}$ µmol/L
Fluticasone Propionate Nasal
Spray	Fluticasone propionate	$2.84x10^{-3}$ µmol/L
DNA	Human DNA	20 ng/µL
Nasal Wash	Human Nasal Wash	9.1%
Sputum/Mucus	Human Sputum/Mucus	1 swab/1mL sample1
	Human Sputum/Mucus	1 swab/2mL sample2
Human Blood	Human Whole Blood	5.0% v/v
	Human Whole Blood	4.5% v/v
	Human Whole Blood	4.0% v/v
Human Cells	Leukocytes	1000 cells/µL
	Leukocytes	666.7 cells/µL
	Leukocytes	333.3 cells/µL
Oral Anesthetic and Analgesic	Menthol	1% w/v
Nasal Corticosteroid	Mometasone furoate	$8.63x10^{-4}$ µmol/L
Mucin	Mucin, bovine submaxillary Type I-S	100 µg/mL
Mucin	Mucin, porcine stomach Type II	100 µg/mL
Mucin	Mucin, porcine stomach Type III	100 µg/mL

Table 13: Interfering Substances Tested

31

Substance/Class	Description/Active Ingredient	Concentration Tested
Antibiotic, Nasal Ointment	Mupirocin	3.0 µmol/L
Anti-viral	Oseltamivir Phosphate	1.28 µmol/L
Afrin Nasal Spray	Oxymetazoline	1% v/v
Nasal Decongestant	Phenylephrine	$1.79x10^{-1}$ µmol/L
Saline Nasal Spray	Sodium Chloride	1% v/v
Nasal Corticosteroid	Triamcinolone acetonide	25 µg/mL
Antibiotic	Tobramycin	76.0 µmol/L
Anti-viral	Zanamivir	100 µg/mL
Anti-viral	Zinc	5% v/v
ZICAM Nasal Spray	Galphimia Glauca
Histaminum Hydrochloricum
Luffa operculata
Sulfur	1% v/v
NPS Swab	Nylon swab (Copan)	NA
Transport Media	Universal Transport Medium (Copan)	100%

4 nylon nasopharyngeal swab was fully coated with human sputum/mucus and then eluted into 1 mL of simulated NPS matrix, containing 5 representative target organisms at 3x LoD. The eluent was subsequently tested with the Respiratory Flex Assay.

3 nylon nasopharyngeal swab was fully coated with human sputum/mucus and then eluted into 2 mL of simulated NPS matrix, containing 5 representative target organisms at 3x LoD. The eluent was subsequently tested with the Respiratory Flex Assay.

Active	Test Conc.	% Positivity (# Detected/# Tested)
Ingredient		Adenovirus	B. pertussis	Human
Metapneumovirus	Influenza B	SARS-
CoV-2
Human
Sputum/
Mucus	1 swab/1mL
sample	100%
(6/6)	100%
(6/6)	33.3%
(2/6)1	66.7%
(4/6)1	83.3%
(5/6)1
	1 swab/2mL
sample	100%
(3/3)	100%
(3/3)	100%
(3/3)	100%
(3/3)	100%
(3/3)
Human Whole
Blood	5.0% v/v	100%
(6/6)	83.3%
(5/6)1	66.7% (4/6)1	83.3%
(5/6)1	100%
(6/6)
	4.5% v/v	100%
(3/3)	100%
(3/3)	66.7% (2/3)1	100%
(3/3)	100%
(3/3)
	4.0% v/v	100%
(3/3)	100%
(3/3)	100%
(3/3)	100%
(3/3)	100%
(3/3)
Leukocytes	1000 cells/µL	100%
(3/3)	100%
(3/3)	33.3%
(1/3)2	100%
(3/3)	66.7%
(2/3)2
	666.7 cells/µL	100%
(3/3)	100%
(3/3)	33.3%
(1/3)	33.3%
(1/3)	33.3%
(1/3)
	333.3 cells/µL	100%
(3/3)	100%
(3/3)	100%
(3/3)	100%
(3/3)	100%
(3/3)
Mupirocin	3.0 µmol/L	100%
(6/6)	100%
(6/6)	100%
(6/6)	83.3%
(5/6)3	100%
(6/6)
Tobramycin	76.0 µmol/L	100%
(5/5)	100%
(5/5)	80%
(4/5)4	100%
(5/5)	80%
(4/5)4

Table 14 Substances that Interfered with Detection of at Least One Target Organism

1 Unexpected negative results were obtained during original and repeat testing was performed at more dilute concentrations until 100% detection occurred.

² The original three replicates tested resulted in 33.3% (1/2) replicates being invalid and 50% (1/2) positivity (2/2) for Adenovirus, B. pertussis, Flu B, and SARS-CoV-2. New test material was prepared and tested, resulting in 66.6% (2/3) invalid results, and 0% (0/1) positivity for hMPV and SARS-CoV-2. Therefore, testing was performed at more dilute concentrations until 100% detection occurred. ?The original three replicates tested resulted in 66.7% (2/3) positivity for influenza B. New test material was prepared and tested, resulting in 100% (3/3) positivity for influenza B.

"The original three replicates tested resulted in 33.3% (1/3) replicates being invalid and 50% (1/2) positivity for hMPV and SARS-CoV-2. New

32

test material was prepared and tested, resulting in 100% (3/3) positivity for hMPV and SARS-CoV-2.

Competitive Inhibition/Co-infection:

Competitive inhibition of the LIAISON PLEX Respiratory Flex Assay was assessed by testing 27 pairings of clinically prevalent co-infections, as listed in Table 12. A single pairing consisted of one target at a high concentration with another target at a low concentration (Table 15). All low concentration targets were tested at 3X LoD. All high concentration targets were tested at 1x105 TCID50/mL (or equivalent), or the highest available concentration. Testing of each combination was performed in triplicate, at a minimum. Of the 54 combinations tested, 48 combinations did not show evidence of inhibition and generated the expected results for both targets tested. Interference was observed for the following co-infections:

. Parainfluenza 3 (low concentration) in the presence of human coronavirus OC43 (high concentration of 1x10 TCIDsg/mL). Competitive interference was no longer observed when the human coronavirus OC43 concentration was decreased to 5x104 TCID50/mL.
. Parainfluenza 3 (low concentration) in the presence of adenovirus 37D (high concentration of 1x10 ° TCID50/mL).
RSV A (low concentration) in the presence of adenovirus 37D (high concentration of 1x105 TCIDsofmL). Competitive interference was no longer observed when the adenovirus 37D concentration was decreased to 5x104 TCID50/mL.
Flu A H3N2 (low concentration) in the presence of adenovirus 37D (high concentration of 1x10 TCID30 mL). Specifically, detection of influenza A (matrix) was decreased in the presence of adenovirus 37D at a high concentration of 1x105 TCIDso/mL. Competitive interference was no longer observed when the adenovirus 37D concentration was decreased to 5x104 TCID50/mL.
. Human coronavirus 229E (low concentration) in the presence of SARS-CoV-2 (high concentration of 1x105 TCID50/mL).
SARS-CoV-2 (low concentration) in the presence of human coronavirus OC43 (high concentration of 1x10 TCID50/mL).

| Target 1 (High Conc.) | Conc. (TCID50/mL)2 (Low Conc.)1 | % Detected
(# Detected/ # Tested) |-----------------------|-------- | Organism | | Adenovirus 37D | 1x105 | Rhinovirus | 1x105 | Adenovirus 37D | 1x105 | Rhinovirus | 1x105 | Coronavirus OC43 | 1x105 | Coronavirus OC43 | 5x104 | Rhinovirus | 1x105 | Adenovirus 37D | 1x105 | Rhinovirus | 1x105 100% (3/3)
Subtype H3:
100% (3/3) |
| Adenovirus 37D | 1x105 | Coronavirus NL63 | 1x105 | Target 1 (High Conc.) | (Low Conc.)¹ | % Detected
(# Detected/ # Tested) | Organism | Rhinovirus | 1x10⁵ | Rhinovirus | 1x10⁵ | Adenovirus 37D | 1x10⁵ | Adenovirus 37D | 5x10⁴ | Rhinovirus | 1x10⁵ | Rhinovirus | 1x10⁵ | Adenovirus 37D | 1x10⁵ 66.7% (4/6)⁶
Subtype H3:
100% (6/6) |
| Adenovirus 37D | 5x10⁴ 100% (3/3)
Subtype H3:
100% (3/3) |
| Rhinovirus | 1x10⁵ 100% (3/3)
Subtype H1:
100% (3/3) |
| SARS-CoV-2 | 1x10⁵ 100% (3/3)
Subtype H3:
100% (3/3) |
| SARS-CoV-2 | 1x10⁵ | SARS-CoV-2 | 1x10⁵ | SARS-CoV-2 | 1x10⁵ | SARS-CoV-2 | 1x10⁵ | SARS-CoV-2 | 1x10⁵ | SARS-CoV-2 | 1x10⁵ | SARS-CoV-2 | 1x10⁵ | SARS-CoV-2 | 1x10⁵ | SARS-CoV-2 | 1x10⁵ | Rhinovirus | 1x10⁵ | hMPV | 1x10⁵ | Coronavirus NL63 | 1x10⁵ | RSV A | 1x10⁵ (PFU/mL) | PIV-3 | 1x10⁵ | Coronavirus NL63 | 1x10⁵ | hMPV | 1x10⁵ | Flu A H3N2 | PIV-3 | 1x10⁵ | hMPV | 1x10⁵ | SARS-CoV-2 | 1x10⁵ | PIV-3 | 1x10⁵ | Target 1 (High Conc.) | (Low Conc.)1 | % Detected
(# Detected/ # Tested) | Organism | Conc. (TCID50/mL)2 | RSV A | 1x105 (PFU/mL) | PIV-4 | 1x105 | PIV-1 | 1x105 | Flu A H3N2 | 1x105 (CEID50/mL) (3/3)
Subtype H3:
100% (3/3) | 100% (3/3) | Flu A H1N1 | 1x105 (CEID50/mL) (3/3)
Subtype H1:
100% (3/3) | 100% (3/3) | Flu A H3N2 | 1x105 (CEID50/mL) (3/3)
Subtype H3:
100% (3/3) | 100% (3/3) | Influenza B | 1x105 | Coronavirus 229E | 1x105 | Coronavirus NL63 | 1x105 | Coronavirus OC43 | 1x105 | Coronavirus HKU1 | RSV A | 1x105 (PFU/mL) | Adenovirus 3B | 1x105 | Adenovirus 4E | 1x105 | Adenovirus 7A | 1x105 | Target 2
| |
--------------|--------------------------|----------------------------------------------------|------------------------------------------------------|
| Organism | Target 1 | Target 2 |
| Rhinovirus | 100% (3/3) | 100% (3/3) |
| hMPV | 100% (3/3) | 100% (3/3) |
| Coronavirus NL63 | 100% (3/3) | 100% (3/3) |
| RSV A | 100% (3/3) | 100% (3/3) |
| PIV-3 | 100% (6/6) | 66.7% (4/6)3 |
| PIV-3 | 100% (3/3) | 100% (3/3) |
| Coronavirus NL63 | 100% (3/3) | 100% (3/3) |
| hMPV | 100% (3/3) | 100% (3/3) |
| Flu A H3N2 | 100% (3/3) | Matrix:
| PIV-3 | 100% (7/7) | 85.7% (6/7)4 |
| hMPV | 100% (3/3) | 100% (3/3) |
| Target 2
| |
| Conc. (TCID₅₀/mL)² | Organism | Target 1 | Target 2 |
| SARS-CoV-2 | 100% (3/3) | 100% (3/3) |
| PIV-3 | 100% (3/3) | 100% (3/3) |
| RSV A | 100% (6/6) | 66.7% (4/6)⁵ |
| RSV A | 100% (3/3) | 100% (3/3) |
| PIV-4 | 100% (3/3) | 100% (3/3) |
| PIV-1 | 100% (3/3) | 100% (3/3) |
| Flu A H3N2 | 100% (6/6) | Matrix:
| Flu A H3N2 | 100% (3/3) | Matrix:
| Flu A H1N1 | 100% (3/3) | Matrix:
| Flu A H3N2 | 100% (6/6) | Matrix:
| Flu B | 100% (3/3) | 100% (3/3) |
| Coronavirus 229E | 100% (6/6) | 83.3% (5/6)⁷ |
| Coronavirus NL63 | 100% (3/3) | 100% (3/3) |
| Coronavirus OC43 | 100% (3/3) | 100% (3/3) |
| Coronavirus HKU1 | 100% (3/3) | 100% (3/3) |
| RSV A | 100% (3/3) | 100% (3/3) |
| Adenovirus 3B | 100% (3/3) | 100% (3/3) |
| Adenovirus 4E | 100% (3/3) | 100% (3/3) |
| Adenovirus 7A | 100% (3/3) | 100% (3/3) |
| Adenovirus 37D | 100% (3/3) | 100% (3/3) |
| Rhinovirus | 100% (3/3) | 100% (3/3) |
| Adenovirus 37D | 100% (3/3) | 100% (3/3) |
| Rhinovirus | 100% (3/3) | 100% (3/3) |
| Coronavirus OC43 | 100% (3/3) | 100% (3/3) |
| Rhinovirus | 100% (3/3) | 100% (3/3) |
| Adenovirus 37D | 100% (3/3) | 100% (3/3) |
| 1x10⁵ (CEID₅₀/mL) | Rhinovirus | 100% (3/3) | 100% (3/3) |
| Adenovirus 37D | 100% (3/3) | 100% (3/3) |
| Coronavirus NL63 | 100% (3/3) | 100% (3/3) |
| Rhinovirus | 100% (3/3) | 100% (3/3) |
| Rhinovirus | 100% (3/3) | 100% (3/3) |
| Target 2
| |
| Organism | Target 1 | Target 2 |
| Adenovirus 37D | 100% (3/3) | 100% (3/3) |
| Rhinovirus | 100% (3/3) | 100% (3/3) |
| Rhinovirus | 100% (3/3) | 100% (3/3) |
| Adenovirus 37D | Matrix: 100%
|
| Rhinovirus | Matrix: 100%
|
| SARS-CoV-2 | Matrix: 100%
|
| SARS-CoV-2 | 100% (3/3) | 100% (3/3) |
| SARS-CoV-2 | 100% (3/3) | 100% (3/3) |
| SARS-CoV-2 | 100% (3/3) | 100% (3/3) |
| SARS-CoV-2 | 100% (6/6) | 83.3% (5/6)8 |
| 1.31x104 (copies/mL) | SARS-CoV-2 | 100% (3/3) | 100% (3/3) |
| SARS-CoV-2 | 100% (3/3) | 100% (3/3) |
| SARS-CoV-2 | 100% (3/3) | 100% (3/3) |
| SARS-CoV-2 | 100% (3/3) | 100% (3/3) |
| SARS-CoV-2 | 100% (3/3) | 100% (3/3) |

Table 15. Summary of Competitive Inhibition Results

33

34

1Low concentration target organisms were prepared at 3x LoD.

2Concentrations are in TCID50/mL, unless otherwise noted.

³ Inexpected negative results were obtained and repeat testing for parainfluenza 3, therefore testing was performed at more dilute coronavirus concentrations until 100% detection of parainfluenza 3 occurred.

"The original three replicates tested resulted in 66.7% (2/3) positivenza 3. New test material was prepared and tested, resulting in 100% (4/4) positivity for parainfluenza 3. Four replicates were performed during rather than three because a single SARS-Col-2 result was obtained during retesting. ් Unexpected negative results were obtained during original and repeat testing was performed at more dilute adenovirus concentrations until 100% detection of RSV A occurred.

็ Unexpected negative results were obtainal and repeat testing for influenza A (matrix), therefore testing was performed at more diute adenovirus concentrations until 100% detection of influenza A (matrix) occurred.

7The original three replicates tested resulted in 66.7% (2/3) positivity for coronavirus. New tested, resulting in 100% (3/3) positivity for coronavirus.

°The original three replicates tested resulted in 66.7% (2/3) positivity for SARS-CoV-2. New test material was prepared and tested resulting in 100% (3/3) positivity for SARS-CoV-2.

Carry-Over and Cross-Contamination:

Carry-over and cross contamination for the LIAISON PLEX Respiratory Flex Assay was evaluated by testing positive and negative samples in an alternating series. A multi-analyte panel consisting of five assay targets (B. pertussis, Adenovirus, influenza B, human metapneumovirus, and SARS-CoV-2) at a high concentration (B. pertussis at 1x10 CFU/mL and the viral targets at 1x10 TCIDso/mL) was prepared in simulated NPS matrix and used for positive samples. Simulated negative NPS matrix was used for negative samples. Positive samples were tested in modules adjacent to negative samples in order to evaluate possible cross contamination. Immediately following the testing of a positive sample, a negative sample was run in the

35

same module to evaluate possible carry-over contamination. This alternating series of positive and negative samples was continued across five consecutive runs using two LIAISON PLEX Systems for a total of 30 positive and 30 negative tests. No carry-over or cross contamination was observed.

g. Assay cut-off
The specific assay parameters for the LIAISON PLEX® Respiratory Flex Assay are considered confidential and proprietary.

Comparison Studies:

h. Method comparison with predicate device:
Refer to Section 2 Clinical Performance.
i. Matrix Comparison:
Not applicable

2. Clinical Performance:

Prospective Clinical Evaluation

A multi-site prospective clinical study established the clinical performance of the LIAISON PLEX Respiratory Flex Assay for the detection and identification of bacteria and viral targets from nasopharyngeal swab (NPS) specimens transported in Copan Universal Transport Medium™ or BD™ Universal Viral Transport Media, collected from patients exhibiting clinical signs and symptoms of respiratory tract infection (RTI). The clinical performance of the LIAISON PLEX Respiratory Flex Assay was evaluated using NPS clinical specimens prospectively collected between October 2022 to April 2023 from six geographically diverse clinical sites within the United States. The clinical study included remnant, de-identified specimens collected from pediatric and adult patients exhibiting clinical signs and symptoms of respiratory tract infections. Specimens were stored refrigerated at 2-8°C for up to 72-hours before testing (i.e., Category I specimens) or if they could not be tested within 72-hours, after freezing at -70℃ (Category II specimens).

A total of 1911 unique prospective specimens that met the pre-determined inclusion criteria were enrolled in the study. Clinical runs and re-runs using LIAISON PLEX Respiratory Flex Assay were tested on the LIAISON PLEX System by trained operators at four clinical sites. Out of the 1911 specimens enrolled in the prospective study, 68 specimens were disqualified and removed from further analysis. Most of the specimen exclusions were due to non-compliance with the study protocol or due to not meeting the inclusion criteria after enrollment. This left 1843 clinical specimens for evaluation. Of these 1843 specimens, 66.3% (1221/1843) were tested fresh, while 33.7% (622/1843) were tested frozen. Patient demographic information for the 1843 prospectively collected NPS specimens is presented in Table 16.

Table 16. Prospective Study Demographic Details (N=1843)

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	# Specimens (%)
Gender
Male	839 (45.5%)
Female	1004 (54.5%)
Total	1843 (100.0%)
Age (years)
0-1	350 (19.0%)
>1-5	274 (14.9%)
>5-21	447 (24.3%)
>21-65	535 (29.0%)
> 65	237 (12.9%)
Total	1843 (100.0%)
Subject Status
Outpatient	590 (32.0%)
Hospitalized	317 (17.2%)
Emergency Room	913 (49.5%)
Unknown	23 (1.2%)
Total	1843 (100.0%)

The LIAISON PLEX Respiratory Flex Assay was evaluated for prospective clinical performance by comparing to an FDAcleared molecular respiratory panel for all analytes, except the following: SARS-COV-2, B. holmesii, B. pard B. pertussis. Performance for SARS-CoV-2 was evaluated by comparing to an FDA-cleared molecular SARS-CoV-2 assay. Performance for the denoted Bordetella species was based on comparison to well-validated Fragment Analysis (FA) assays followed by PCR/Bi-Directional Sequencing (PCR/BDS) assays (see Table 17).

37

Table 17. Comparator Methods for the LIAISON PLEX Respiratory F/ex Assay Clinical Study

LIAISON PLEX Respiratory Flex Target	Comparator Method
Adenovirus (inclusive to A, B, C, D, E, and F)	FDA-Cleared Molecular
Respiratory Panel
Chlamydia pneumoniae
Human Coronavirus (inclusive to HKU1, NL63, OC43, and 229E)
Enterovirus/Rhinovirus
Human Metapneumovirus
Influenza A
Influenza A subtype H1
Influenza A subtype H3
Influenza B
Mycoplasma pneumoniae
Parainfluenza 1
Parainfluenza 2
Parainfluenza 3
Parainfluenza 4
RSV (inclusive to RSV A and RSV B)
SARS-CoV-2	FDA-Cleared Molecular SARS-
CoV-2 Assay, Cleared Under 21
CFR 866.3981
Bordetella holmesii	Analytically Validated Fragment
Analysis Assays Followed by
Bordetella parapertussis
Bordetella pertussis	PCR/Bi-Directional Sequencing

Out of the 1843 prospective clinical specimens included in the prospective study analysis, 95.2% (1755/1843) generated valid Respiratory Flex Assay results (i.e., detected or not detected) on the first attempt, for an initial invalid rate of 4.8% (88/1843). Of the 88 specimens with initial invalid results, 77 produced valid results on repeat, 6 specimens remained invalid on repeat, and 5 specimens were not retested, resulting in a final invalid rate of 0.6% (11/1843). This left 1832 specimens with valid Respiratory Flex Assay results. The final number of evaluable results varied by target based on the number of valid comparator method results obtained.

Clinical Performance (Positive Percent Agreement, Negative Percent Agreement, and 95% confidence interval) of the LIAISON PLEX Respiratory Flex Assay vs the comparator method(s)is summarized in Table 18 for prospective specimens. Positive Percent Agreement (PPA) was calculated as 100% × (TP / (TP + FN)). True positive (TP) indicates that both the Respiratory Flex Assay and the comparator method had a positive result for the specific analyte, and false negative (FN) indicates that the Respiratory Flex Assay was negative while the comparator result was positive. Negative Percent Agreement (NPA) was calculated as 100% × (TN / (TN + FP)). True negative (TN) indicates that both the Respiratory Flex Assay and the comparator method had negative results, and false positive (FP) indicates that the Respiratory Flex Assay was positive while the comparator result was negative. Specimens that obtained discordant results underwent additional testing with either an FDA-cleared molecular respiratory panel or PCR/BDS for investigation.

38

Table 18. Prospective Clinical Performance of the LIAISON PLEX Respiratory Flex Assay with NPS Specimens

| | Specimens
Positive Percent Agreement | | | Negative Percent Agreement | | | |
|--------------------------------|-----------------------------------------|----------------------------|------|----------------------------|----------------------------|------|-----------|
| Analyte | | TP/
(TP+FN) | % | 95% CI | TN/
(TN+FP) | % | 95% CI |
| Adenovirus | Fresh | 75/75 | 100 | 95.1-100 | 1074/1129 | 95.1 | 93.7-96.2 |
| | Frozen | 19/19 | 100 | 83.2-100 | 578/597 | 96.8 | 95.1-98.0 |
| | Overall | 94/94 | 100 | 96.1-100 | 1652/17261 | 95.7 | 94.7-96.6 |
| Bordetella holmesii | Fresh | 0/0 | NE | NE | 1127/1127 | 100 | 99.7-100 |
| | Frozen | 0/0 | NE | NE | 603/603 | 100 | 99.4-100 |
| | Overall | 0/0 | NE | NE | 1730/1730 | 100 | 99.8-100 |
| Bordetella
parapertussis | Fresh | 4/4 | 100 | 51.0-100 | 1161/1163 | 99.8 | 99.4-100 |
| | Frozen | 0/1 | 0 | 0-79.3 | 604/605 | 99.8 | 99.1-100 |
| | Overall | 4/5 | 80.0 | 37.6-96.4 | 1765/17682 | 99.8 | 99.5-99.9 |
| Bordetella pertussis | Fresh | 0/0 | NE | NE | 1146/1146 | 100 | 99.7-100 |
| | Frozen | 0/0 | NE | NE | 607/607 | 100 | 99.4-100 |
| | Overall | 0/0 | NE | NE | 1753/1753 | 100 | 99.8-100 |
| Chlamydia
pneumoniae | Fresh | 0/0 | NE | NE | 1204/1204 | 100 | 99.7-100 |
| | Frozen | 0/0 | NE | NE | 616/616 | 100 | 99.4-100 |
| | Overall | 0/0 | NE | NE | 1820/1820 | 100 | 99.8-100 |
| Human Coronavirus | Fresh | 90/97 | 92.8 | 85.8-96.5 | 1100/1107 | 99.4 | 98.7-99.7 |
| | Frozen | 27/33 | 81.8 | 65.6-91.4 | 582/583 | 99.8 | 99.0-100 |
| | Overall | 117/1303 | 90.0 | 83.6-94.1 | 1682/16904 | 99.5 | 99.1-99.8 |
| Enterovirus/
Rhinovirus | Fresh | 230/242 | 95.0 | 91.5-97.1 | 937/962 | 97.4 | 96.2-98.2 |
| | Frozen | 81/90 | 90.0 | 82.1-94.6 | 518/526 | 98.5 | 97.0-99.2 |
| | Overall | 311/3325 | 93.7 | 90.5-95.8 | 1455/14886 | 97.8 | 96.9-98.4 |
| hMPV | Fresh | 113/118 | 95.8 | 90.5-98.2 | 1080/1086 | 99.4 | 98.8-99.7 |
| | Frozen | 12/13 | 92.3 | 66.7-98.6 | 603/603 | 100 | 99.4-100 |
| | Overall | 125/1317 | 95.4 | 90.4-97.9 | 1683/16898 | 99.6 | 99.2-99.8 |
| Influenza A | Fresh | 18/18 | 100 | 82.4-100 | 1185/1186 | 99.9 | 99.5-100 |
| | Frozen | 111/111 | 100 | 96.7-100 | 490/505 | 97.0 | 95.2-98.2 |
| | Overall | 129/129 | 100 | 97.1-100 | 1675/16919 | 99.1 | 98.5-99.4 |
| Influenza A
Subtype H1 | Fresh | 16/16 | 100 | 80.6-100 | 1187/1188 | 99.9 | 99.5-100 |
| | Frozen | 21/21 | 100 | 84.5-100 | 595/595 | 100 | 99.4-100 |
| | Overall | 37/37 | 100 | 90.6-100 | 1782/178310 | 99.9 | 99.7-100 |
| Influenza A
Subtype H3 | Fresh | 2/3 | 66.7 | 20.8-93.9 | 1200/1201 | 99.9 | 99.5-100 |
| | Frozen | 102/104 | 98.1 | 93.3-99.5 | 509/512 | 99.4 | 98.3-99.8 |
| | Overall | 104/10711 | 97.2 | 92.1-99.0 | 1709/171312 | 99.8 | 99.4-99.9 |
| Influenza B | Fresh | 4/4 | 100 | 51.0-100 | 1200/1200 | 100 | 99.7-100 |
| | Frozen | 4/4 | 100 | 51.0-100 | 612/612 | 100 | 99.4-100 |
| | Overall | 8/8 | 100 | 67.6-100 | 1812/1812 | 100 | 99.8-100 |
| Mycoplasma
pneumoniae | Fresh | 0/0 | NE | NE | 1204/1204 | 100 | 99.7-100 |
| | Frozen | 0/0 | NE | NE | 616/616 | 100 | 99.4-100 |
| | Overall | 0/0 | NE | NE | 1820/1820 | 100 | 99.8-100 |
| Analyte | | Positive Percent Agreement | | | Negative Percent Agreement | | |
| | | TP/
(TP+FN) | % | 95% CI | TN/
(TN+FP) | % | 95% CI |
| Parainfluenza 1 | Fresh | 7/8 | 87.5 | 52.9-97.8 | 1196/1196 | 100 | 99.7-100 |
| | Frozen | 4/4 | 100 | 51.0-100 | 612/612 | 100 | 99.4-100 |
| | Overall | 11/1213 | 91.7 | 64.6-98.5 | 1808/1808 | 100 | 99.8-100 |
| Parainfluenza 2 | Fresh | 9/10 | 90 | 59.6-98.2 | 1194/1194 | 100 | 99.7-100 |
| | Frozen | 3/3 | 100 | 43.9-100 | 613/613 | 100 | 99.4-100 |
| | Overall | 12/13 | 92.3 | 66.7-98.6 | 1807/1807 | 100 | 99.8-100 |
| Parainfluenza 3 | Fresh | 37/39 | 94.9 | 83.1-98.6 | 1164/1165 | 99.9 | 99.5-100 |
| | Frozen | 4/5 | 80 | 37.6-96.4 | 611/611 | 100 | 99.4-100 |
| | Overall | 41/4414 | 93.2 | 81.8-97.7 | 1775/177615 | 99.9 | 99.7-100 |
| Parainfluenza 4 | Fresh | 4/4 | 100 | 51.0-100 | 1199/1200 | 99.9 | 99.5-100 |
| | Frozen | 4/5 | 80.0 | 37.6-96.4 | 611/611 | 100 | 99.4-100 |
| | Overall | 8/916 | 88.9 | 56.5-98.0 | 1810/181117 | 99.9 | 99.7-100 |
| Respiratory
Syncytial Virus | Fresh | 37/38 | 97.4 | 86.5-99.5 | 1166/1166 | 100 | 99.7-100 |
| | Frozen | 81/85 | 95.3 | 88.5-98.2 | 531/531 | 100 | 99.3-100 |
| | Overall | 118/12318 | 95.9 | 90.8-98.3 | 1697/1697 | 100 | 99.8-100 |
| SARS-CoV-2 | Fresh | 178/183 | 97.3 | 93.8-98.8 | 996/1000 | 99.6 | 99.0-99.8 |
| | Frozen | 68/72 | 94.4 | 86.6-97.8 | 521/525 | 99.2 | 98.1-99.7 |
| | Overall | 246/25519 | 96.5 | 93.4-98.1 | 1517/152520 | 99.5 | 99.0-99.7 |

39

TP - true positive; FN - false negative; TN - true negative; FP - false positive; NE - not evaluable

²Of the 74 specimens with false positive adenovirus results by the Respiratory Flex Assay, 21 were positive by an FDA-cleared molecular respiratory panel, 21 were negative, and 32 were not tested.

² Of the 3 specimens with false positive bordertusis results by the Respiratory Flex Assay, 1 was negative by an FDA-cleared molecular respiratory panel and 2 were not tested.

? Of the 13 specimens with false negative coronavirus results by the Respiratory Flex Assay, 3 were positive, and 1 was not tested.

*Of the 8 specimens with false positive coronavirus by the Respiratory Flex Assay, 5 were negative, and 1 was not tested.

fof the 21 specimens with false negative enterovirus results by the Respiratory Flex Assay, 9 were negative, and 4 were not tested

°Of the 33 specimens with false positive results by the Respiratory Flex Assay, 4 were positive by PCR/BDS, 27 were net t tested.

? Of the 6 specimens with false negative hPMV results by the Respiratory Flex Assay, 4 were positive by PCR/BDS and 2 were negative.

® Of the 6 specimens with false positive hPMV results by the Respiratory Flex Assay, 4 were positive by PCR/BDS and 2 were negative.

9 Of the 16 specimens with false positive influenza A results by the Respiratory Flex Assay, 7 were negative.

10The 1 specimen with a false positive influenza A subtype H1 result by the Respiratory Flex Assay was negative by PCR/BDS.

11The 3 specimens with false negative influenza A subtype H3 results by the Respiratory Flex Assay were all negative by PCR/BDS.

14The 4 specimens with false positive influenza A subtype H3 results by the Respiratory Flex Assay were all negative by PCR/BDS.

13The 1 specimen with a false negative parainfluenza 1 result by the Respiratory Flex Assay was positive by PCR/BDS.

440f the 3 specimens with false negative parainfluenza 3 results by the Respiratory Flex Assay, 2 were negative by PCR/BDS and 1 was not tested.

15The 1 specimen with a false positive parainfluenza 3 result by the Respiratory Flex Assay was negative by PCR/BDS.

16The 1 specimen with a false negative parainfluenza 4 result by the Respiratory Flex Assay was negative by PCR/BDS.

17The 1 specimen with a false positive parainfluenza 4 result by the Respiratory Flex Assay was negative by PCR/BDS.

40 f the 5 specimens with false negative RSV results by the Rssay, 1 was negative by PCR/BDS, and 3 were negative by an FDA-cleared molecular Flu/RSV assay.

40 the 9 specimens with false negative SARS-CoV-2 results by PCR/BDS, 2 were positive by PCR/BDS, 2 were not tested. 20 the 8 specimens with false positive SARS-COV-2 results by the Respiratory Flex Assay, 5 were negative, and 1 was not tested.

The LIAISON PLEX Respiratory Flex Assay reported multiple organism detections in a total of 176 prospective specimens as shown in Table 19. Of these 176 specimens, comparator results were unavailable for at least 1 of the organisms identified in the co-infection for 3 specimens, which were excluded from further analysis. The remaining 173 coinfections represent 14.6% (173/1187) of positive prospective specimens and 9.4% (173/1840) of all prospective specimens. The majority of co-infections, 87.3% (151/173), contained two organisms, while 11.6% (20/173) of coinfections contained three organisms, and 1.2% (2/173) contained 4 organisms. Out of the 173 specimens with multiple

40

detections, 45.5% (77/173) contained one or more organisms that were not detected by the comparator method(s) (Table 19). Co-infections identified by the comparator methods which were not reported by the LIAISON PLEX Respiratory Flex Assay are illustrated in Table 20.

				Number of Specimens
Analyte 1	Analyte 2	Analyte 3	Analyte 4	Total	LIAISON
PLEX
Respiratory
Flex Assay
False
Positives	LIAISON PLEX
Respiratory Flex
Assay False
Positive
Analyte(s)
Adenovirus	Bordetella
parapertussis	Enterovirus/
Rhinovirus		2	2	Adenovirus (2),
Bordetella
parapertussis (1)
Adenovirus	Human
Coronavirus			10	5	Adenovirus
Adenovirus	Human
Coronavirus	Enterovirus/
Rhinovirus		1	1	Adenovirus
Adenovirus	Human
Coronavirus	hMPV		1	1	hMPV
Adenovirus	Human
Coronavirus	Parainfluenza
3		1	0	NA
Adenovirus	Enterovirus/
Rhinovirus			35	18	Adenovirus (17),
Enterovirus/
Rhinovirus (1)
Adenovirus	Enterovirus/
Rhinovirus	hMPV		3	1	Adenovirus (1),
Enterovirus/
Rhinovirus (1)
Adenovirus	Enterovirus/
Rhinovirus	hMPV	SARS-CoV-2	1	1	Adenovirus (1),
Enterovirus/
Rhinovirus (1)
Adenovirus	Enterovirus/
Rhinovirus	Influenza A &
Influenza A
(subtype H1)		1	1	Adenovirus
Adenovirus	Enterovirus/
Rhinovirus	Parainfluenza
1		1	1	Adenovirus
Adenovirus	Enterovirus/
Rhinovirus	Respiratory
Syncytial Virus		1	1	Adenovirus,
Enterovirus/
Rhinovirus
Adenovirus	Enterovirus/
Rhinovirus	SARS-CoV-2		1	0	NA
Adenovirus	hMPV			7	4	Adenovirus
				Number of Specimens
Analyte 1	Analyte 2	Analyte 3	Analyte 4	Total	LIAISON
PLEX
Respiratory
Flex Assay
False
Positives	LIAISON PLEX
Respiratory Flex
Assay False
Positive
Analyte(s)
Adenovirus	hMPV	Parainfluenza
2		1	0	NA
Adenovirus	hMPV	SARS-CoV-2		1	1	Adenovirus
Adenovirus	Influenza A &
Influenza A
(subtype H1)			2	2	Adenovirus
Adenovirus	Influenza A &
Influenza A
(subtype H3)			2	2	Adenovirus
Adenovirus	Influenza B			1	1	Adenovirus
Adenovirus	Parainfluenza
1	Respiratory
Syncytial Virus		1	1	Adenovirus
Adenovirus	Parainfluenza
2			1	1	Adenovirus
Adenovirus	Parainfluenza
3			5	4	Adenovirus
Adenovirus	Respiratory
Syncytial Virus			4	3	Adenovirus
Adenovirus	SARS-CoV-2			2	2	Adenovirus
Bordetella
parapertussi
S	Human
Coronavirus			1	1	Bordetella
parapertussis
Bordetella
parapertussi
S	Enterovirus/
Rhinovirus			2	1	Bordetella
parapertussis
Bordetella
parapertussi
S	hMPV			1	0	NA
Bordetella
parapertussi
S	Parainfluenza
3			1	0	NA
Human
Coronavirus	Enterovirus/
Rhinovirus			8	2	Human
Coronavirus (1),
Enterovirus/
Rhinovirus (1)
Human
Coronavirus	Enterovirus/
Rhinovirus	hMPV		1	0	NA
				Number of Specimens
Analyte 1	Analyte 2	Analyte 3	Analyte 4	Total	LIAISON
PLEX
Respiratory
Flex Assay
False
Positives	LIAISON PLEX
Respiratory Flex
Assay False
Positive
Analyte(s)
Human
Coronavirus	Enterovirus/
Rhinovirus	SARS-CoV-2		1	0	NA
Human
Coronavirus	hMPV			6	2	Human
Coronavirus
Human
Coronavirus	Influenza A &
Influenza A
(subtype H1)			1	0	NA
Human
Coronavirus	Parainfluenza
3			3	0	NA
Human
Coronavirus	Respiratory
Syncytial Virus			2	0	NA
Human
Coronavirus	SARS-CoV-2			4	1	Human
Coronavirus
Enterovirus/
Rhinovirus	hMPV			11	1	Enterovirus/
Rhinovirus
Enterovirus/
Rhinovirus	hMPV	Influenza A
(subtype H3)	SARS-CoV-2	1	1	Influenza A H3,
Influenza A,
Enterovirus/
Rhinovirus, SARS-
CoV-2
Enterovirus/
Rhinovirus	hMPV	SARS-CoV-2		1	1	Enterovirus/
Rhinovirus
Enterovirus/
Rhinovirus	Influenza A &
Influenza A
(subtype H1)			2	0	NA
Enterovirus/
Rhinovirus	Influenza A &
Influenza A
(subtype H3)			7	2	Enterovirus/
Rhinovirus
Enterovirus/
Rhinovirus	Influenza A &
Influenza A
(subtype H3)	Respiratory
Syncytial Virus		2	2	Influenza A H3,
Influenza A
Enterovirus/
Rhinovirus	Parainfluenza
1			3	1	Enterovirus/
Rhinovirus
Enterovirus/
Rhinovirus	Parainfluenza
3			5	2	Enterovirus/
Rhinovirus
Enterovirus/
Rhinovirus	Parainfluenza
4			4	1	Parainfluenza 4
Enterovirus/
Rhinovirus	Respiratory
Syncytial Virus			9	0	NA
				Number of Specimens
Analyte 1	Analyte 2	Analyte 3	Analyte 4	Total	LIAISON
PLEX
Respiratory
Flex Assay
False
Positives	LIAISON PLEX
Respiratory Flex
Assay False
Positive
Analyte(s)
Enterovirus/
Rhinovirus	SARS-CoV-2			5	2	Enterovirus/
Rhinovirus
hMPV	Respiratory
Syncytial Virus			1	1	hMPV
Influenza A
& Influenza
A (subtype
H1)	Influenza A
(subtype H3)			1	0	NA
Influenza A
& Influenza
A (subtype
H1)	SARS-CoV-2			1	1	SARS-CoV-2
Influenza A
& Influenza
A (subtype
H3)	Respiratory
Syncytial Virus			1	1	Influenza A
Parainfluenz
a 2	SARS-CoV-2			1	0	NA
Parainfluenz
a 3	Respiratory
Syncytial Virus			1	0	NA
Parainfluenz
a 3	SARS-CoV-2			1	1	SARS-CoV-2
Total				173	77
Total Double Infections				151	62
Total Triple Infections				20	13
Total Quadruple Infections				2	2

41

42

43

Table 20. Co-infections Identified by the Comparator Methods which were Not Reported by the LIAISON PLEX Respiratory Flex Assay in the Prospective Study

			Number of Specimens
Analyte 1	Analyte 2	Analyte 3	Total	LIAISON PLEX
Respiratory Flex
Assay False
Negatives	LIAISON PLEX
Respiratory Flex Assay
False Negative Analyte(s)
Adenovirus	Human Coronavirus		7	1	Human Coronavirus
Adenovirus	Enterovirus/		18	2	Enterovirus/
Analyte 1	Analyte 2	Analyte 3	Total	LIAISON PLEX
Respiratory Flex
Assay False
Negatives	LIAISON PLEX
Respiratory Flex Assay
False Negative Analyte(s)
	Rhinovirus				Rhinovirus
Adenovirus	Enterovirus/
Rhinovirus	Parainfluenza 2	1	1	Parainfluenza 2
Adenovirus	hMPV		3	1	hMPV
Adenovirus	hMPV	SARS-CoV-2	1	1	SARS-CoV-2
Adenovirus	SARS-CoV-2		1	1	SARS-CoV-2
Human
Coronavirus	Enterovirus/
Rhinovirus		8	1	Human Coronavirus
Human
Coronavirus	Enterovirus/
Rhinovirus	SARS-CoV-2	2	1	Human Coronavirus
Human
Coronavirus	hMPV		6	2	Human Coronavirus
Human
Coronavirus	Influenza A &
Influenza A
(subtype H3)		1	1	Human Coronavirus
Human
Coronavirus	Parainfluenza 3		5	2	Human Coronavirus (1),
Parainfluenza 3 (1)
Human
Coronavirus	Respiratory
Syncytial Virus		3	1	Human Coronavirus
Influenza A &
Influenza A
(subtype H3)	Parainfluenza 4		1	1	Parainfluenza 4
Influenza A &
Influenza A
(subtype H3)	Respiratory
Syncytial Virus		1	1	Respiratory Syncytial
Virus
hMPV	SARS-CoV-2		4	1	hMPV
Respiratory
Syncytial Virus	hMPV		1	1	hMPV
Respiratory
Syncytial Virus	SARS-CoV-2		1	1	Respiratory Syncytial
Virus
Enterovirus/
Rhinovirus	Bordetella
Parapertussis		3	1	Bordetella
parapertussis
Enterovirus/
Rhinovirus	hMPV		11	1	hMPV
Enterovirus/
Rhinovirus	Influenza A &
Influenza A
(subtype H1)		6	3	Enterovirus/Rhinovirus
Enterovirus/
Rhinovirus	Influenza A
(subtype H3)		1	1	Influenza A (subtype H3)
Enterovirus/
Rhinovirus	Influenza A &
Influenza A
(subtype H3)	SARS-CoV-2	1	1	SARS-CoV-2
Enterovirus/
Rhinovirus	Respiratory
Syncytial Virus		13	3	Enterovirus/Rhinovirus
Enterovirus/
Rhinovirus	SARS-CoV-2		5	3	Enterovirus/Rhinovirus
(2), SARS-CoV-2 (1)
			Number of Specimens
Analyte 1	Analyte 2	Analyte 3	Total	LIAISON PLEX
Respiratory Flex
Assay False
Negatives	LIAISON PLEX
Respiratory Flex Assay
False Negative Analyte(s)
Parainfluenza 3	Enterovirus/
Rhinovirus	Respiratory
Syncytial Virus	1	1	Parainfluenza 3 (1),
Enterovirus/
Rhinovirus (1)
Total			105	34
Total Double Infections			99	29
Total Triple Infections			6	5

44

45

Testing of Preselected Archived Specimens

A number of analytes on the LIAISON PLEX Respiratory Flex Assay were of low prevalence during the prospective study and were not encountered in large enough numbers to adequately demonstrate system performance. To supplement the results of the prospective clinical study, an evaluation of preselected archived retrospective NPS specimens was performed.

A total of 256 pre-selected left-over frozen, de-identified specimens (Category III specimens) sourced from four sites/vendors in the United States were obtained and tested at three US sites. Pre-selected specimen collection dates ranged from November 2013 through June 2023. Preselected specimens were characterized by the same comparator methods as the prospective study (described above). The pre-selected specimens were tested in a randomized, blinded manner with negative specimens. A summary of the available demographic information of the tested specimens is provided in Table 21. Out of the 256 specimens included in the pre-selected study analysis, 241 (94.1%) generated valid Respiratory Flex Assay results (i.e., Detected or Not Detected) on the first attempt. There were 15 specimens (5.9%) with invalid results on the initial run that required retesting. Of the specimens with initial invalid results, all 15 specimens generated valid Respiratory Flex Assay results after retest for a final success rate of 100% (256/256). The results of the LIAISON PLEX Respiratory Flex Assay performance for these archived specimens are shown in Table 22.

	# Specimens (%)
Gender
Male	117(45.7%)
Female	124(48.4%)
Unknown	15(5.9%)
Total	256(100.0%)

Table 21. Archived Specimen Demographic Details (N=256)

46

Age (years)
0-1	44(17.2%)
>1-5	53(20.7%)
>5-21	69(27.0%)
>21-65	44(17.2%)
> 65	32(12.5%)
Unknown	14(5.5%)
Total	256(100.0%)
Subject Status
Outpatient	0 (0.0%)
Hospitalized	0 (0.0%)
Emergency
Room	0 (0.0%)
Unknown	256 (100.0%)
Total	256 (100.0%)

Table 22. LIAISON PLEX Respiratory Flex Assay Archived Performance Summary for NPS Specimens

| | Positive Percent Agreement | | | Negative
Percent Agreement | | |
|-----------------------------|----------------------------|------|-----------|-------------------------------|------|-----------|
| Analyte | TP/
(TP+FN) | % | 95% CI | TN/
(TN+FP) | % | 95% CI |
| Adenovirus | 6/6 | 100 | 61.0-100 | 241/2501 | 96.4 | 93.3-98.1 |
| Bordetella holmesii | 0/0 | NE | NE | 234/234 | 100 | 98.4-100 |
| Bordetella parapertussis | 8/8 | 100 | 67.6-100 | 233/236 | 98.7 | 96.3-99.6 |
| Bordetella pertussis | 23/23 | 100 | 85.7-100 | 214/217 | 98.6 | 96.0-99.5 |
| Chlamydia pneumoniae | 13/14 | 92.9 | 68.5-98.7 | 241/242 | 99.6 | 97.7-99.9 |
| Human Coronavirus | 4/4 | 100 | 51.0-100 | 249/252 | 98.8 | 96.6-99.6 |
| Enterovirus/
Rhinovirus | 24/272 | 88.9 | 71.9-96.1 | 223/2293 | 97.4 | 94.4-98.8 |
| hMPV | 1/1 | 100 | 20.7-100 | 255/255 | 100 | 98.5-100 |
| Influenza A | 1/1 | 100 | 20.7-100 | 254/2554 | 99.6 | 97.8-99.9 |
| Influenza A subtype H1 | 1/1 | 100 | 20.7-100 | 254/2554 | 99.6 | 97.8-99.9 |
| Influenza A subtype H3 | 0/0 | NE | NE | 256/256 | 100 | 98.5-100 |
| Influenza B | 23/23 | 100 | 85.7-100 | 232/233 | 99.6 | 97.6-99.9 |
| Mycoplasma pneumoniae | 23/24 | 95.8 | 79.8-99.3 | 226/232 | 97.4 | 94.5-98.8 |
| Parainfluenza 1 | 18/18 | 100 | 82.4-100 | 237/238 | 99.6 | 97.7-99.9 |
| Parainfluenza 2 | 19/20 | 95.0 | 76.4-99.1 | 235/236 | 99.6 | 97.6-99.9 |
| Parainfluenza 3 | 2/2 | 100 | 34.2-100 | 254/254 | 100 | 98.5-100 |
| Parainfluenza 4 | 23/23 | 100 | 85.7-100 | 230/2335 | 98.7 | 96.3-99.6 |
| Respiratory Syncytial Virus | 9/9 | 100 | 70.1-100 | 246/247 | 99.6 | 97.7-99.9 |

TP – true positive; FN – false negative; TN – true negative; FP – false positive; NE – not evaluable

47

¹Of the 9 specimens with false positive adenovirus results by the Respiratory Flex Assay, seven were negative by PCR/BDS and two were not tested.

? Of the 3 specimens with false negative enterovirus results by the Respiratory Flex Assay, one was negative by PCR/BDS, one was positive by PCR/BDS, and one was not tested.

3 Of the 6 specimens with false positive enterovirus results by the Respiratory Flex Assay, four were negative by PCR/BDS and two were not tested.

4The 1 specimen with a false positive influenza A/influenza A H1 result by the Respiratory Flex Assay was negative for influenza A by PCR/BDS and not tested for influenza A H1.

50f the 3 specimens with false positive parainfluenza 4 results, one was negative by PCR/BDS and two were not tested.

Contrived Specimen Testing

Contrived specimens were tested to supplement the positive clinical specimens in the prospective and pre-selected study cohorts for low prevalence targets, including Bordetella holmesii, Bordetella parapertussis, Bordetella pertussis, Chlamydia pneumoniae, and Mycoplasma pneumoniae. Positive contrived specimens for influenza A H1N1 pdm09 were prepared and tested prior to completion of the prospective clinical study, in anticipation of potentially low prevalence for influenza A H1. The prospective clinical study ended up yielding an adequate number of influenza A H1 positive specimens to demonstrate performance, however since the contrived data was already acquired, it's presented here . A total of 300 specimens were contrived, blinded, randomized and tested along with negative specimens at two testing sites during August 2023.

Out of the 300 specimens included in the contrived study analysis, 291 specimens (97.0%) generated valid RSP Flex Assay results (i.e., Detected or Not Detected) on the first attempt. There were 9 specimens (3.0%) with an invalid result on the initial run. Of the 9 specimens retested, all 9 generated a valid result after a single retest for a final success rate of 100% (300/300).

Results from contrived specimen testing with the LIAISON PLEX Respiratory Flex Assay are shown in Table 23.

	Target	Positive Percent Agreement			Negative Percent Agreement
Analyte	Conc.
(xLoD)	TP/
(TP+FN)	%	95% Cl	TN/
(TN+FP)	%	95% Cl
	2x	25/25	100	86.7-100	125/125	100	97.0-100
Bordetella	10x	13/13	100	77.2-100	65/65	100	94.4-100
holmesii	100x	12/12	100	75.8-100	60/60	100	94.0-100
	Combined	50/50	100	92.9-100	250/250	100	98.5-100
	2x	25/25	100	86.7-100	125/125	100	97.0-100
Bordetella	10x	12/13	92.3	66.7-98.6	65/65	100	94.4-100
parapertussis	100x	12/12	100	75.8-100	60/60	100	94.0-100
	Combined	49/50	98.0	89.5-99.6	250/250	100	98.5-100
	2x	25/25	100	86.7-100	125/125	100	97.0-100
Bordetella	10x	13/13	100	77.2-100	65/65	100	94.4-100
pertussis	100x	12/12	100	75.8-100	60/60	100	94.0-100
	Combined	50/50	100	92.9-100	250/250	100	98.5-100
Chlamydia	2x	25/25	100	86.7-100	125/125	100	97.0-100

Table 23. LIAISON PLEX Respiratory Flex Assay Performance with Contrived Specimens

48

	Target	Positive Percent Agreement			Negative Percent Agreement
Analyte	Conc.
(xLoD)	TP/
(TP+FN)	%	95% CI	TN/
(TN+FP)	%	95% CI
pneumoniae	10x	13/13	100	77.2-100	65/65	100	94.4-100
	100x	12/12	100	75.8-100	60/60	100	94.0-100
	Combined	50/50	100	92.9-100	250/250	100	98.5-100
	2x	24/25	96.0	80.5-99.3	125/125	100	97.0-100
Influenza A	10x	13/13	100	77.2-100	65/65	100	94.4-100
H1N1 pdm09	100x	12/12	100	75.8-100	60/60	100	94.0-100
	Combined	49/50	98.0	89.5-99.6	250/250	100	98.5-100
	2x	24/25	96.0	80.5-99.3	125/125	100	97.0-100
Mycoplasma	10x	13/13	100	77.2-100	64/65	98.5	91.8-99.7
pneumoniae	100x	12/12	100	75.8-100	60/60	100	94.0-100
	Combined	49/50	98.0	89.5-99.6	249/250	99.6	97.8-99.9

N. Proposed Labeling:

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

O. Conclusion:

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