The Panther Fusion SARS-CoV-2/Flu A/B/RSV assay is a fully automated multiplexed real-time polymerase chain reaction (RT-PCR) in viro diagnostic test intended for the qualitative detection and differentiation of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), influenza A virus (Flu A), influenza B virus (Flu B), and respiratory syncytial virus (RSV). Nucleic acids are isolated and purified from nasopharyngeal (NP) specimens obtained from individuals exhibiting signs and symptoms of a respiratory tract infection. Clinical signs and symptoms of respiratory viral infection due to SARS-CoV-2, influenza, and RSV can be similar. This assay is intended to aid in the differential diagnosis of SARS-CoV-2, Flu A, Flu B, and RSV infections in humans and is not intended to detect influenza C virus infections.

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

Positive results do not rule out coinfection with other organism(s) detected by the Panther Fusion SARS-CoV-2/Flu A/B/RSV assay may not be the definite cause of disease.

Negative results do not preclude SARS-CoV-2, influenza A virus, influenza B virus, or RSV infections. This assay is designed for use on the Panther Fusion system.

The Hologic RespDirect Collection Kit can be used to collect NP specimens for testing with the Panther Fusion

Device Description

The Panther Fusion SARS-CoV-2/Flu A/B/RSV assay is a multiplex real-time reverse transcriptase PCR (RT-PCR) in vitro diagnostic test developed for use on the fully automated Panther Fusion system to detect and differentiate SARS-CoV-2, influenza A, influenza B, and respiratory syncytial virus (RSV) directly from nasopharyngeal (NP) swab specimens collected into UTM/VTM or with the Hologic RespDirect Collection Kit, from individuals exhibiting signs and symptoms of a respiratory tract infection.

The Hologic RespDirect Collection Kit is intended for the collection of NP swab specimens. Each individual collection kit is comprised of a single flocked NP swab and an enhanced Direct Load Tube (eDLT) containing 2.9mL of enhanced Specimen Transport Media (eSTM) which are flow wrapped together for customer convenience.

The Panther Fusion SARS-CoV-2/Flu A/B/RSV assay involves the following steps: a) Sample lysis; b) Nucleic acid capture and elution; c) Elution transfer and multiplex RT-PCR.

The Panther Fusion system integrates Hologic's commercialized Panther instrument system with an add-on sidecar, the Panther Fusion module, which extends the functionality of the Panther system by increasing the assay processing capabilities to include real-time PCR (RT-PCR). The Panther Fusion module includes instrument hardware and can be installed on existing Panther instruments or ordered with new Panther instruments.

The Panther Fusion system employs non-specific target capture (NSTC) for the purification of RNA and DNA from the sample, followed by nucleic acid amplification and real-time fluorescent detection. The process involves sample loading and preparation (i.e. nucleic acid extraction) on the Panther instrument using similar workflow and processing steps as for other commercialized Hologic Aptima TMA assays. The extracted nucleic acid for each sample is transferred to the Panther Fusion module where PCR amplification and detection occurs.

AI/ML Overview

The information provided in the document refers to a Panther Fusion SARS-CoV-2/Flu A/B/RSV Assay, which is an in vitro diagnostic test for the qualitative detection and differentiation of SARS-CoV-2, influenza A virus (Flu A), influenza B virus (Flu B), and respiratory syncytial virus (RSV). This is a laboratory diagnostic device, not an AI/ML-enabled device for medical imaging analysis. Therefore, much of the requested information (e.g., number of experts, adjudication method, MRMC study, standalone performance for an algorithm, training set details) is not applicable to this type of device submission.

However, I can extract and structure the relevant acceptance criteria and performance data as presented for this diagnostic assay.

Acceptance Criteria and Reported Device Performance

1. A table of acceptance criteria and the reported device performance:

Since specific "acceptance criteria" for each performance metric are not explicitly stated in a single table, I will synthesize them from the descriptions of the studies. Performance is generally demonstrated by achieving high Positive Percent Agreement (PPA) and Negative Percent Agreement (NPA) with comparator methods, or by demonstrating acceptable analytical performance (LoD, cross-reactivity, precision, etc.).

Criterion (Implicit from Study Design)	Reported Device Performance
Clinical Performance (Prospective Study):
- SARS-CoV-2 PPA	Overall: 96.9% (95% CI: 94.7-98.2)
- SARS-CoV-2 NPA	Overall: 98.5% (95% CI: 97.7-99.0)
- Flu A PPA	Overall: 98.4% (95% CI: 94.3-99.6)
- Flu A NPA	Overall: 99.7% (95% CI: 99.3-99.9)
- Flu B PPA	Overall: Not calculable (0/0 positive cases)
- Flu B NPA	Overall: 99.8% (95% CI: 99.4-99.9)
- RSV PPA	Overall: 84.6% (95% CI: 57.8-95.7)
- RSV NPA	Overall: 100% (95% CI: 99.8-100)
Clinical Performance (Retrospective Study):
- Flu A PPA	93.1% (95% CI: 78.0-98.1)
- Flu B PPA	95.5% (95% CI: 78.2-99.2)
- RSV PPA	100% (95% CI: 92.4-100)
- Flu A NPA (Confirmed Negative)	100% (95% CI: 94.5-100)
- Flu B NPA (Confirmed Negative)	100% (95% CI: 95.0-100)
- RSV NPA (Confirmed Negative)	100% (95% CI: 92.6-100)
Analytical Sensitivity (LoD):	Defined as lowest concentration at which >95% of all replicates tested positive. Specific LoD values per viral strain are provided in Table 3. For example, SARS-CoV-2 USA-WA1/2020: 0.03 TCID50/mL; Flu A/Brisbane/02/18 (H1N1): 0.06 TCID50/mL.
Reproducibility:	Agreement values were 100% for all panel member components, except: True negative (Flu B): 98.9% (one false positive). Flu A low positive: 98.9% (one false negative), indicating robustness. Total signal variability overall <2.04% in the precision study for panel members.
Interfering Substances:	No cross-reactivity observed in analyte negative pools (0% positive results). No interference to analyte positive pools at 3xLoD (100% positive results).
Cross-Reactivity (Wet Testing):	All pools had expected results of no interference or cross-reactivity. For example, Adenovirus 1 at 1x10^5 TCID50/mL did not cross-react.
Analytical Reactivity (Strain Testing):	The assay demonstrates SARS-CoV-2, Flu A, Flu B, and RSV strain wet testing inclusivity for the strains tested at the illustrated concentrations (e.g., SARS-CoV-2 strains detected at 0.09-0.30 TCID50/mL, Flu A strains at 0.18-183 TCID50/mL, Flu B strains at 0.006-33 TCID50/mL, RSV strains at 0.06-0.30 TCID50/mL).
Analytical Reactivity (In Silico):	Predicted to detect all 934,493 SARS-CoV-2 sequences evaluated (10% random sampling up to 6/25/2022), including VOCs like Delta and Omicron. Predicted to detect ≥99.998% of 88,128 Flu A, ≥99.94% of 31,801 Flu B, ≥98.12% of 1,599 RSV A, and ≥98.23% of 1,240 RSV B sequences (Jan 2015 - Feb 2022).
Cross-Reactivity (In Silico):	No predicted cross-reactivity or interference with genetically closely related or commonly encountered respiratory microorganisms, with the exception of S. marcescens (which wet testing confirmed caused no interference).
Analytical Exclusivity:	The assay did not detect non-indicated influenza types C and D.
Specimen Stability:	VTM/UTM: Primary specimens stable refrigerated for up to 96 hours, or frozen. Processed specimens stable at room temp for up to 6 days, refrigerated for 3 months, or frozen (-20°C or -70°C) for 3 months. Supports minimization of freeze/thaw cycles.RespDirect: Stable at 15°C-30°C for up to 6 days. Stable refrigerated (2-8°C) or frozen (-20°C or -70°C) for up to 3 months. May undergo up to 3 freeze-thaw cycles when stored at -20°C or -70°C.
Carry-over/Cross-contamination:	No carry-over or cross-contamination observed (100% Flu A positivity for positive panels, 0% for negative panels in checkerboard runs).
Transport Media Equivalency:	Demonstrated equivalency between various VTM/UTM vendor types and between clinical NP matrix and simulated NP matrix based on comparable positivity observed at 1xLoD and 5xLoD for all targets.
Collection Device/Swab Equivalency:	Comparable positivity observed between RespDirect Collection Kit and control collection device for all concentrations and targets. Swab equivalency study demonstrated equivalency between RespDirect swab and control NP swab for absorption and elution of targets (100% positivity for all targets with both swab types at 4x LoD).
Competitive Interference/Co-Infection:	Most targets at 3xLoD maintained positivity when paired with other assay targets at 1x10^4 TCID50/mL (with specific exceptions noted for high concentrations of RSV B affecting SARS-CoV-2 and Flu A, and RSV B affecting Flu B, and SARS-CoV-2 affecting Flu A at specific concentrations). Five co-infections detected by the Panther Fusion assay were largely confirmed by comparator testing in the prospective study. Two co-infections in the retrospective study were also confirmed by comparator testing.

Study Details (Applicable Sections)

2. Sample sizes used for the test set and the data provenance (e.g. country of origin of the data, retrospective or prospective)

Clinical Performance Study:
- Prospective Study:
  - Total specimens enrolled: 1949 NP swab specimens.
  - Evaluable specimens: 1900 (after withdrawals and removals).
    - SARS-CoV-2 evaluation: 1887 samples (790 fresh, 1097 frozen).
    - Flu A, Flu B, RSV evaluation: 1837 samples (798 fresh, 1039 frozen).
  - Data Provenance: Multicenter study in the US (five participating US pediatric/adolescent, private and/or university hospitals). Data collected between January 2022-March 2022 and November 2020-March 2021. Remnant NP swab specimens.
- Retrospective Study:
  - Evaluable specimens: 95 preselected archived NP swab specimens.
  - Data Provenance: Collected between December 2019 and March 2020. Preselected based on historic qualitative positive result.
Analytical Studies:
- LoD: Minimum of 24 replicates per concentration, per reagent lot (3 lots total = 72 replicates per strain) for initial determination, confirmed with additional 24 replicates. For RespDirect, 30 replicates per concentration.
- Interfering Substances: Not specified as a total number, but pools tested with and without assay analytes.
- Cross-Reactivity (Wet Testing): Minimum of three replicates per microorganism panel.
- Analytical Reactivity (Strain Wet Testing): Not specified by exact number of replicates, but sufficient to demonstrate detection at ~3x LoD.
- Within Laboratory Precision and Repeatability: 96 measurements performed for each panel member (total from 2 operators, 2 runs/day, 3 reagent lots, 3 Panther Fusion systems, 12 days).
- Competitive Interference: 3 replicates per panel.
- Hologic RespDirect Collection Device Equivalency/Swab Equivalency: Paired NP swabs from 25 symptomatic donors for collection device equivalency. 20 swabs per swab type for swab equivalency.
- Carry-over/Cross-contamination: 100 replicates of negative panel per baseline run; 30 positive and 30 negative panels in alternating "checkerboard" runs.
- Reproducibility Study: 150 samples tested at each of 3 sites (total ~450 samples). Each panel member had 90 results across all sites.

3. Number of experts used to establish the ground truth for the test set and the qualifications of those experts (e.g. radiologist with 10 years of experience)

This is not applicable as the ground truth was established by molecular diagnostic comparator assays, not human experts in medical imaging.

4. Adjudication method (e.g. 2+1, 3+1, none) for the test set

Prospective Clinical Study (for SARS-CoV-2): A composite comparator algorithm (CCA) was used. It consisted of two highly sensitive US FDA EUA SARS-CoV-2 molecular tests and a validated PCR followed by bi-directional sequencing (PCR/BDS) assay. A final CCA result was assigned when two of the three composite comparator assays were in concordance.
Prospective Clinical Study (for Flu A, Flu B, and RSV): A US FDA-cleared molecular Flu A/B/RSV assay was the comparator method.
Retrospective Clinical Study: Confirmatory testing using a US FDA-cleared molecular Flu A/B/RSV assay.
Discordant Testing: For discordant results in both clinical studies, additional testing with a US FDA EUA SARS-CoV-2/Flu A/B/RSV molecular test was performed (volume permitting).

5. If a multi reader multi case (MRMC) comparative effectiveness study was done, If so, what was the effect size of how much human readers improve with AI vs without AI assistance

This is not applicable as the device is an in vitro diagnostic assay and does not involve human readers for interpretation, nor does it incorporate AI/ML in a way that would be assessed through an MRMC study.

6. If a standalone (i.e. algorithm only without human-in-the-loop performance) was done

This is not applicable in the context of an AI/ML algorithm. The device, an automated RT-PCR assay, inherently performs without human intervention beyond sample loading and results interpretation; its performance is thus "standalone" in that sense.

7. The type of ground truth used (expert consensus, pathology, outcomes data, etc)

Clinical Performance Studies:
- SARS-CoV-2: Composite Comparator Algorithm (CCA) consisting of two highly sensitive US FDA EUA SARS-CoV-2 molecular tests and a validated PCR followed by bi-directional sequencing (PCR/BDS) assay.
- Flu A, Flu B, RSV: US FDA-cleared molecular Flu A/B/RSV assays (for both prospective and retrospective studies), and confirmatory testing with US FDA EUA SARS-CoV-2/Flu A/B/RSV molecular test for discordant results.
Analytical Studies: Ground truth was established by spiked samples with known concentrations of viral strains, or by known negative matrices.

8. The sample size for the training set

This is not applicable as the device is an in vitro diagnostic assay and does not use a training set in the AI/ML sense. Analytical and clinical validation studies are performed to characterize its performance.

9. How the ground truth for the training set was established

This is not applicable as there is no training set in the AI/ML sense.

Summary

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Image /page/0/Picture/0 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 - USA 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.

Hologic, Inc. Vlada Rudenko Regulatory Affairs Specialist 10210 Genetic Center Drive San Diego, California 92121

Re: K222736

Trade/Device Name: Panther Fusion SARS-CoV-2/Flu A/B/RSV 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, OOI Dated: September 8, 2022 Received: September 9, 2022

Dear Vlada Rudenko:

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 (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 located 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.

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

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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 803) for devices or postmarketing safety reporting (21 CFR 4, Subpart B) for combination products (see https://www.fda.gov/combination-products/guidance-regulatory-information/postmarketing-safety-reportingcombination-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 4, Subpart A) for combination products; and, if applicable, the electronic product radiation control provisions (Sections 531-542 of the Act); 21 CFR 1000-1050.

Also, please note the regulation entitled, "Misbranding by reference to premarket notification" (21 CFR Part 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 medical devices and radiation-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 Viral Respiratory and HPV Branch Division of Microbiology Devices OHT7: Office of In Vitro Diagnostics Office of Product Evaluation and Quality Center for Devices and Radiological Health

Enclosure

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Indications for Use

510(k) Number (if known) K222736

Device Name Panther Fusion SARS-CoV-2/Flu A/B/RSV Assay

Indications for Use (Describe)

Positive results do not rule out coinfection with other organism(s) detected by the Panther Fusion SARS-CoV-2/Flu A/B/RSV assay may not be the definite cause of disease.

Negative results do not preclude SARS-CoV-2, influenza A virus, influenza B virus, or RSV infections. This assay is designed for use on the Panther Fusion system.

The Hologic RespDirect Collection Kit can be used to collect NP specimens for testing with the Panther Fusion

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

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

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510(k) SUMMARY

K222736

Panther Fusion SARS-CoV-2/Flu A/B/RSV Assay

Hologic, Inc. I. 510(k) Owner: 10210 Genetic Center Drive San Diego, CA 92121

Contact Person: Vlada Rudenko, MA Regulatory Affairs Specialist vlada.rudenko@hologic.com Phone: 858.410.7967 Fax: N/A
Date Prepared: May 15, 2023

II. DEVICE

Proprietary Name:	Panther Fusion SARS-CoV-2/Flu A/B/RSV Assay
Classification Name:	Multi-Target Respiratory Specimen Nucleic Acid Test IncludingSARS-CoV-2 And Other Microbial Agents
Regulation Number:	21 CFR 866.3981
Regulatory Class:	Class II
Product Code:	QOF
Secondary Product Code:	OOI

III. PREDICATE DEVICE

The predicate device for the Panther Fusion SARS-CoV-2/Flu A/B/RSV assay is the BioFire Respiratory Panel 2.1 (BioFire RP2.1) Assay (DEN200031; granted March 17, 2021, BioFire Diagnostics, LLC).

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IV. DEVICE DESCRIPTION

The Panther Fusion SARS-CoV-2/Flu A/B/RSV assay involves the following steps: a) Sample lysis; Prior to processing and testing on the Panther Fusion system, specimens are transferred to a tube containing specimen transport media (STM) that lyses the cells, releases target nucleic acid and protects them from degradation during storage. For specimens collected with the Hologic RespDirect Collection Kit, the swab is placed directly into the eDLT where the eSTM that lyses the cells, releases the nucleic acid and protects them from degradation during storage.

b) Nucleic acid capture and elution takes place in a single tube on the Panther Fusion system. The eluate is transferred to the Panther Fusion system reaction tube containing the assay reagents. The Internal Control-S (IC-S) is added to the Panther Fusion Capture Reagent-S (FCR-S) which gets added to each specimen. The IC-S in the reagent is used to monitor specimen processing, amplification, and detection. Magnetic particles with covalently bound oligonucleotides mediate the nucleic acid capture. Capture oligonucleotides hybridize to total nucleic acid in the test specimen. Hybridized nucleic acid is then separated from the lysed specimen in a magnetic field. Wash and aspiration steps remove extraneous components debris from the reaction tube. The elution step elutes purified nucleic acid.

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c) Elution transfer and multiplex RT-PCR; Eluted nucleic acid is transferred to a Panther Fusion reaction tube already containing oil and reconstituted master mix. A reverse transcriptase generates a DNA copy of the target sequence. Target specific forward and reverse primers and probes then amplify targets while simultaneously detecting and discriminating multiple target types via multiplex RT-PCR. The Panther Fusion system compares the fluorescence signal to a predetermined cut-off to produce a qualitative result for the presence or absence of the analyte. The positive result for each analyte will be accompanied by the cycle threshold (Ct value). The analytes and the channel used for their detection on the Panther Fusion system is summarized in the table below.

Analyte	Gene Targeted	Instrument Channel
SARS-CoV-2	ORF1ab	ROX
Influenza A Virus	Matrix	FAM
Respiratory Syncytial Virus A/B	Matrix	HEX
Influenza B Virus	Matrix	RED647
Internal Control	Not applicable	RED677

*Internal Control is a non-infectious synthetic nucleic acid sequence that is extracted and detected through targeted primers and probes.

Assay Components

The reagents required to perform the Panther Fusion SARS-CoV-2/Flu A/B/RSV assay are packaged and sold separately. There are 7 boxes containing 9 reagents which are required for sample processing. The Hologic RespDirect Collection Kit can be used to collect NP specimens for testing with the Panther Fusion SARS-CoV-2/Flu A/B/RSV Assay. Additionally, other NP swabs (not provided with the Hologic RespDirect Collection Kit) may be used to collect NP specimens in 3mL of VTM or UTM. A description of the components that are required to perform the Panther Fusion SARS-CoV-2/Flu A/B/RSV assay and the Hologic RespDirect Collection kit is detailed in Table 1. Boxes 3 - 7 were cleared by FDA as part of K171963.

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Table 1: Reagents Required to Perform the Panther Fusion SARS-CoV-2/Flu A/B/RSV Assav

Box	Components Description	Part Number
1	Panther Fusion SARS-CoV-2/Flu A/B/RSV Assay Cartridges -96 TestsPanther Fusion SARS-CoV-2/Flu A/B/RSV assay cartridge, 12tests, 8 per box	PRD-07400
2	Panther Fusion SARS-CoV-2/Flu A/B/RSV ControlsPositive Control tube, 5 per boxPanther Fusion Negative Control tube, 5 per box	PRD-07401
3	Panther Fusion Extraction Reagent-S - 960 TestsPanther Fusion Capture Reagent-S bottle, 240 tests, 4 per boxPanther Fusion Enhancer Reagent-S bottle, 240 tests, 4 per box	PRD-04331
4	Panther Fusion Internal Control-S - 960 TestsPanther Fusion Internal Control-S tube, 4 per box	PRD-04332
5	Panther Fusion Reconstitution Buffer I - 1920 TestsPanther Fusion Reconstitution Buffer I pack, 960 tests, 2 per box	PRD-04333
6	Panther Fusion Elution Buffer - 2400 TestsPanther Fusion Elution Buffer pack, 1200 tests, 2 per box	PRD-04334
7	Panther Fusion Oil Reagent - 1920 TestsPanther Fusion Oil Reagent pack, 960 tests, 2 per box	PRD-04335

Ancillary Kits:

1	Panther Fusion Specimen Lysis Tubes(100 tubes)	PRD-04339
2	Hologic RespDirect Collection KitCollection kit composed of 1 flocked NP swab and 1 tubecontaining 2.9mL eSTM), 50 per box	PRD-07788

In addition, select components can also be ordered in the following bundles:

Panther Fusion Universal Fluids Kit: (contains Panther Fusion Oil and Panther Fusion ● Elution Buffer).
Panther Fusion Assay Fluids Kit I-S: (contains Panther Fusion Extraction Reagents-S, ● Panther Fusion Internal Control-S, and Panther Fusion Reconstitution Buffer I).

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Instrumentation

The Panther Fusion SARS-CoV-2/Flu A/B/RSV assay has been designed for and validated on the Panther Fusion system. The Panther Fusion system fully automates all the steps necessary to perform the Panther Fusion SARS-CoV-2/Flu A/B/RSV assay.

V. INDICATIONS FOR USE

Panther Fusion SARS-CoV-2/Flu A/B/RSV Assay

The Panther Fusion® SARS-CoV-2/Flu A/B/RSV assay is a fully automated multiplexed realtime polymerase chain reaction (RT-PCR) in vitro diagnostic test intended for the qualitative detection and differentiation of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), influenza A virus (Flu A). influenza B virus (Flu B) and respiratory syncytial virus (RSV). Nucleic acids are isolated and purified from nasopharyngeal (NP) specimens obtained from individuals exhibiting signs and symptoms of a respiratory tract infection. Clinical signs and

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symptoms of respiratory viral infection due to SARS-CoV-2, influenza and RSV can be similar. This assay is intended to aid in the differential diagnosis of SARS-CoV-2, Flu A, Flu B and RSV infections in humans and is not intended to detect influenza C virus infections.

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

Positive results do not rule out coinfection with other organisms. The organism(s) detected by the Panther Fusion SARS-CoV-2/Flu A/B/RSV assay may not be the definite cause of disease. Negative results do not preclude SARS-CoV-2, influenza A virus, influenza B virus, or RSV infections. This assay is designed for use on the Panther Fusion system.

The Hologic® RespDirect™ Collection Kit can be used to collect NP specimens for testing with the Panther Fusion SARS-CoV-2/Flu A/B/RSV Assay. Additionally, other NP swabs (not provided with the Hologic RespDirect Collection Kit) may be used to collect NP specimens in 3mL of VTM or UTM.

Ancillary Collection Kit:

Hologic RespDirect Collection Kit

The Hologic® RespDirect™ Collection Kit is intended to be used for the collection of nasopharyngeal (NP) swab specimens (collected by a healthcare provider) for testing with the Panther Fusion SARS-CoV-2/Flu A/B/RSV Assay.

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Panther Fusion®

VI. COMPARISON OF TECHNOLOGICAL CHARACTERISTICS WITH THE PREDICATE DEVICE

A comparison of the Panther Fusion SARS-CoV-2/Flu A/B/RSV assay to the predicate BioFire Respiratory Panel 2.1 (DEN200031) is summarized in Table 2.

Table 2: Comparison of Similarities and Differences Between the Subject Device (Panther Fusion SARS-CoV-2/Flu AB/RSV Assay) and the Predicate Device (BioFire Respiratory Panel 2.1 (RP2.1))

Item	Panther FusionSARS-CoV-2/Flu A/B/RSV Assay(Subject Device)	BioFire Respiratory Panel 2.1 (RP2.1)(Predicate Device)DEN200031
PrescriptionUse Only	Yes	Same
SpecimenTypes	Nasopharyngeal (NP) swab specimens	Same
Intended User	Professional use	Same
TechnologyPrinciple ofOperation	Reverse transcriptase multiplexed polymerase chainreaction test	Same
OrganismsDetected	4 targets including:	Same, however the BioFire RP 2.1 includes additionaltargets (listed below)
	SARS-CoV-2	Adenovirus
	Flu A	Coronavirus 229E
	Flu B	Coronavirus HKU1
	RSV (A and B)	Coronavirus NL63
		Coronavirus OC43
		Severe Acute Respiratory Syndrome Coronavirus 2(SARS-CoV-2)
		Human Metapneumovirus
		Human Rhinovirus/Enterovirus
		Panther Fusion
		Influenza A, including subtypes H1, H1-2009, andH3
		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
		Mycoplasma pneumoniae
Assay Controls	Internal and external controls	Same
	Automated nucleic acid amplification platform.	Automated nucleic acid amplification platform.
Platform	Uses Panther Fusion system for all steps includingnucleic acid extraction, amplification, detection andresult processing.	Uses BioFire FilmArray 2.0 or BioFire FilmArrayTorch systems including integrated samplepreparation, amplification, detection, and analysis.
	The Panther Fusion SARS-CoV-2/Flu A/B/RSV assay isa fully automated multiplexed real-time polymerasechain reaction (RT-PCR) in vitro diagnostic testintended for the qualitative detection and differentiationof severe acute respiratory syndrome coronavirus 2(SARS-CoV-2), influenza A virus (Flu A), influenza Bvirus (Flu B), and respiratory syncytial virus (RSV).Nucleic acids are isolated and purified fromnasopharyngeal (NP) specimens obtained fromindividuals exhibiting signs and symptoms of arespiratory tract infection. Clinical signs and symptomsof respiratory viral infection due to SARS-CoV-2,influenza, and RSV can be similar. This assay isintended to aid in the differential diagnosis of SARS-CoV-2, Flu A, Flu B, and RSV infections in humans andis not intended to detect influenza C virus infections.	The BioFire Respiratory Panel 2.1 (RP2.1) is a PCR-based multiplexed nucleic acid test intended for usewith the BioFire FilmArray 2.0 or BioFire FilmArrayTorch systems for the simultaneous qualitativedetection and identification of multiple respiratoryviral and bacterial nucleic acids in nasopharyngealswabs (NPS) obtained from individuals suspected ofrespiratory tract infections, including COVID-19.The following organism types and subtypes areidentified using the BioFire RP2.1:• Adenovirus,• Coronavirus 229E,• Coronavirus HKU1,• Coronavirus NL63,• Coronavirus OC43,• Severe Acute Respiratory Syndrome Coronavirus(SARS-CoV-2)
Intended Use
Nucleic acids from the viral organisms identified bythis test are generally detectable in NP specimensduring the acute phase of infection. The detection andidentification of specific viral nucleic acids fromindividuals exhibiting signs and symptoms ofrespiratory tract infection are indicative of thepresence of the identified virus and aids in diagnosisif used in conjunction with other clinical andepidemiological information, and laboratory findings.The results of this test should not be used as the sole	• Human Metapneumovirus,• Human Rhinovirus/Enterovirus,• Influenza A, including subtypes H1, H1-2009, andH3,• Influenza B,• Parainfluenza Virus 1,• Parainfluenza Virus 2,• Parainfluenza Virus 3,• Parainfluenza Virus 4,• Respiratory Syncytial Virus,
basis for diagnosis, treatment, or other patientmanagement decisions.	• Bordetella parapertussis (IS1001),• Bordetella pertussis (ptxP),• Chlamydia pneumoniae, and
Positive results do not rule out coinfection with otherorganisms. The organism(s) detected by the PantherFusion SARS-CoV-2/Flu A/B/RSV assay may not bethe definite cause of disease.	• Mycoplasma pneumoniaeNucleic acids from the respiratory viral and bacterialorganisms identified by this test are generallydetectable in NPS specimens during the acute phase of
Negative results do not preclude SARS-CoV-2,influenza A virus, influenza B virus, or RSV infections.This assay is designed for use on the Panther Fusionsystem.	infection. The detection and identification of specificviral and bacterial nucleic acids from individualsexhibiting signs and/or symptoms of respiratoryinfection is indicative of the presence of the identified
The Hologic RespDirect Collection Kit can be used tocollect NP specimens for testing with the Panther FusionSARS-CoV-2/Flu A/B/RSV Assay. Additionally, otherNP swabs (not provided with the Hologic RespDirectCollection Kit) may be used to collect NP specimens in3mL of VTM or UTM.	microorganism and aids in the diagnosis of respiratoryinfection if used in conjunction with other clinical andepidemiological information. The results of this testshould not be used as the sole basis for diagnosis,treatment, or other patient management decisions.
Ancillary Collection Kit:	Negative results in the setting of a respiratory illnessmay be due to infection with pathogens that are not
Hologic RespDirect Collection KitThe Hologic RespDirect Collection Kit is intended to beused for the collection of nasopharyngeal (NP) swabspecimens (collected by a healthcare provider) for	detected by this test, or lower respiratory tractinfection that may not be detected by an NPSspecimen. Positive results do not rule out coinfectionwith other organisms. The agent(s) detected by theBioFire RP2.1 may not be the definite cause ofdisease. Additional laboratory testing (e.g., bacterial)
SARS-CoV-2/Flu A/B/RSV Assay Performance		Panther Fusion®
	testing with the Panther Fusion SARS-CoV-2/Flu A/B/RSV Assay.	and viral culture, immunofluorescence, and radiography) may be necessary when evaluating a patient with possible respiratory tract infection.
Time to Obtain Test Results	Approximately 2.5 hours	Approximately 45 minutes
Flu A Subtyping	No	Yes

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Panther Fusion®

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Panther Fusion®

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For Regulatory Submission Use Only

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VII. PERFORMANCE DATA

The following performance data (analytical and clinical) were provided in support of the substantial equivalence determination.

Brief Description of Analytical (Non-Clinical) Studies

The following analytical studies (non-clinical) were conducted to support the clearance of the Panther Fusion SARS-CoV-2/Flu A/B/RSV Assay on the Panther Fusion system.

Specimen Stability (VTM/UTM)

The purpose of this study was to demonstrate the stability of NP specimens collected in Viral Transport Media/Universal Transport Media (VTM/UTM) when stored under recommended temperature conditions and durations for the Panther Fusion SARS-CoV-2/Flu A/B/RSV Assay on the Panther Fusion system. Additionally, freeze/thaw stability was demonstrated. Specimen positive panels were prepared by spiking virus strains of each assay target at 3xLoD or 10xLoD into 20 negative unique clinical matrix pools and tested at 20 replicates per storage condition. This study demonstrates stability for SARS-CoV-2, Flu A, Flu B, and RSV for the following:

Primary Specimens (NP swab stored in VTM/UTM)

a) Stored refrigerated (2-8ºC) for up to 96 hours prior to transfer into a specimen lysis tube.
b) Frozen at -70°C. Freeze/thaw cycles should be minimized due to potential for sample degradation.

Processed Specimens (NP swab in VTM/UTM transferred to a specimen lysis tube)

c) Stored at room temperature (15-30℃) for up to 6 days.

d) Refrigerated (2-8ºC) for up to 3 months.

e) Frozen at-20℃ for up to 3 months. Freeze/thaw cycles should be minimized due to potential for sample degradation.

f) Frozen at -70℃ for up to 3 months. Freeze/thaw cycles should be minimized due to potential for sample degradation.

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Hologic RespDirect Specimen Stability at Room Temperature

The purpose of this study was to demonstrate the stability of specimens collected into eSTM using the RespDirect Collection Kit when stored at room temperature (15℃ to 30℃) up to 7 days and tested in the Panther Fusion SARS-CoV-2/ Flu A/B/RSV assay. The Hologic RespDirect specimen storage condition at 30°C for 7 days was tested to support 15°C to 30°C specimen stability. An unspiked negative panel was prepared from pooled negative clinical eSTM matrix. Positive panels were prepared by spiking one virus strain of each assay target at 3xLoD or 10xLoD. Testing included a minimum of 20 replicates per target concentration per panel on day 0 (baseline) and day 7. A day 0 baseline testing was set. All 20 replicates of positive panels (3x and 10xLoD) showed 100% positivity for all targets on day 0 and after storage at 30 ℃ for 7 days. All 20 replicates of negative panels tested on both Day 0 and Day 7 were negative.

This study demonstrates stability for SARS-CoV-2, Flu A, Flu B and RSV for specimens collected in the Hologic RespDirect Collection Kit stored at 15°C-30°C for up to 6 days.

Hologic RespDirect Specimen Stability at 2-8°C, -20 °C, and -70 °C

The purpose of this study was to demonstrate the stability of specimens collected in eSTM using the Hologic RespDirect Collection Kit when stored refrigerated (2-8ºC) or frozen at -20ºC or-70ºC for 3 months and tested in the Panther Fusion SARS-CoV-2/Flu A/B/RSV assay. An unspiked negative panel was prepared from pooled negative clinical eSTM matrix. Positive panels were prepared by spiking one virus strain of each assay target at 3xLoD or 10xLoD in each negative clinical eSTM matrix pool. Baseline testing was performed on day 0 with 20 replicates per panel member. At each time point (33 days and 99 days), 20 panel aliquots per temperature condition and target concentration were removed from the storage condition (2-8ºC, -20 ºC) and -70 ºC) and tested. Stability results showed 100% positivity for all assay targets (SARS-CoV-2, Flu A, Flu B, and RSV) at 3xLoD and 10xLoD for all timepoints and storage conditions tested, with the exception of SARS-CoV-2 at 3xLoD in the baseline of 2-8°C condition which was 95% positive. All 20 replicates of negative panels tested on day 0 and the various timepoints were negative.

This study demonstrates the following stability for SARS-CoV-2, Flu A, Flu B and RSV for specimens collected in the Hologic RespDirect Collection Kit and tested with the Panther Fusion SARS-CoV-2/Flu A/B/RSV Assay:

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Refrigerated (2-8°C) for up to 3 months or, .
Frozen at -20°C for up to 3 months or, .
. Frozen at -70°C for up to 3 months.

Hologic RespDirect Specimen Freeze Thaw Stability

The purpose of this study was to demonstrate that specimens collected with the RespDirect Collection Kit are stable when stored frozen at -20°C and -70°C and subjected to 4 freeze-thaw cycles. A negative panel was prepared from the pooled negative clinical eSTM matrix. Positive panels were prepared by spiking one virus strain of each assay target at 3xLoD or 10xLoD. Multiple panels were prepared and placed on stability in the appropriate test conditions (storage cycled between -20°C for 2 and 4 freeze/thaw (F/T) cycles or cycled between -70°C and 30°C for 2 and 4 freeze/thaw (F/T) cycles). At each timepoint, samples were removed from their specified storage temperature and tested using the Panther Fusion SARS-CoV-2/Flu A/B/RSV assay. Testing included a minimum of 20 replicates per target concentration per condition at each time point. Results showed 100.0% positivity for all assay targets (SARS-CoV-2, Flu A, Flu B, and RSV) at 3xLoD and 10xLoD in both storage conditions (-20°C and -70°C), with the exception of SARS-CoV-2 at 3xLoD in the 4 F/T condition which was 95.0% positive.

This study demonstrates that NP swab specimens in the Hologic RespDirect Collection Kit may undergo up to 3 freeze-thaw cycles when stored at either -20°C or -70°C.

Transport Media Equivalency

The purpose of this study was to demonstrate equivalency between viral transport media (VTM) and universal transport media (UTM) vendor types when used with the Panther Fusion SARS-CoV-2/Flu A/B/RSV assay. Multiple viral transport media (VTM) and universal transport media (UTM) types were tested and included Remel Micro Test M4RT, Remel Micro Test M5, Remel Micro Test M6, BD Universal Viral Transport Media, Copan Universal Transport Medium, and Hardy Diagnostics Viral Transport Media. To demonstrate equivalency between the viral transport medias, contrived positive samples were created by spiking one strain of SARS-CoV-2, Flu A H3N2, Flu B Victoria, and RSV B into either true negative clinical NP swab matrix or simulated NP swab matrix (i.e., transport media spiked with HeLa cells at a concentration of 2x10 cells/mL) at 3 concentrations (0.5x, 1x, and 5x LoD). Flu A, Flu B, and RSV B were co-spiked while SARS-CoV-2 was evaluated independently. Contrived positives generated with true clinical NP matrix were included to demonstrate equivalency between clinical NP matrix and simulated NP

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matrix, supporting the use of simulated viral transport media in the study. The Panther Fusion SARS-CoV-2/Flu A/B/RSV assay demonstrated equivalency between VTM/UTM vendor medias and NP clinical VTM/UTM matrix based on comparable positivity observed at the 1x LoD and 5x LoD concentrations for SARS-CoV-2, Flu A (H3N2), Flu B (Victoria lineage), and RSV B.

Analytical Sensitivity - Limit of Detection (LoD)

The analytical sensitivity (limit of detection) of the Panther Fusion SARS-CoV-2/Flu A/B/RSV assay was determined by testing dilutions of processed negative clinical NP swab VTM/UTM matrix spiked with the WHO International Standard for SARS-CoV-2, NIBSC (20/146) or viral cultures of: SARS-CoV-2 (1 strain), Influenza A (one H1N1 strain and one H3N2 strain), Influenza B (one Victoria lineage strain and one Yamagata lineage strain) , RSV (one RSV-A strain and one RSV-B strain). Dilutions of cultured virus in TCID50/mL and SARS-CoV-2 WHO standard in IU/mL were prepared in pooled negative clinical nasopharyngeal (NP) swab VTM/UTM matrix processed into STM (1:1.56 ratio). The dilutions were tested at a minimum of 24 replicates per concentration, per reagent lot, using 3 reagent lots for a total of at least 72 replicates per strain. The LoD for each target was determined by Probit analysis for each reagent lot and was confirmed with an additional 24 replicates using a single reagent lot. Analytical sensitivity is defined as the lowest concentration at which >95% of all replicates tested positive.

The results from this study demonstrated that the Panther Fusion SARS-CoV-2/Flu A/B/RSV assay has the following LoD for processed NP specimens:

Viral Strain	LoD Concentration
SARS-CoV-2 USA-WA1/2020	0.03 TCID50/mL
Influenza A/Brisbane/02/18 (H1N1)	0.06 TCID50/mL
Influenza A/Kansas/14/17 (H3N2)	0.10 TCID50/mL
Influenza B/Washington/02/19 (Victoria lineage)	0.03 TCID50/mL
Influenza B/Phuket/3073/13 (Yamagata lineage)	0.003 TCID50/mL
RSV A	0.03 TCID50/mL
RSV B	0.03 TCID50/mL
SARS-CoV-2 WHO International Standard, NIBSC (20/146)	47.20 IU/mL

Table 3: Analytical Sensitivity

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LoD testing was also performed with the Hologic RespDirect Collection Kit. Negative clinical eSTM matrix was spiked with the WHO International Standard for SARS-CoV-2 and 1 strain each for Flu A, Flu B, RSV A, and RSV B. Thirty replicates were tested with a single reagent lot. The lowest concentration that observed ≥95% detection was 98.6 IU/mL for the WHO International Standard for SARS-CoV-2, 0.11 TCID59/mL for Influenza A/Kansas/14/17 (H3N2), 0.03 TCID56/mL for Influenza B/Washington/02/19 (Victoria lineage), 0.03 TCID50/mL for RSV A and 0.05 TCID50/mL for RSV B.

Note: The stated LoDs pertain to the concentrations in the tubes loaded onto the instrument. For samples collected in VTM/UTM, this is the concentration in the processed sample in an SLT. For samples collected using the Hologic RespDirect Collection kit, this is the concentration in the Enhanced Direct Load tube.

Interfering Substances

This study evaluated the performance of the Panther Fusion SARS-CoV-2/Flu A/B/RSV Assay in the presence of potentially interfering endogenous and exogenous substances. Potentially interfering substances were prepared in processed negative clinical NP swab VTM/UTM matrix in STM with up to three substances in each pool. Pools were tested with and without the presence of assay analytes. The analyte negative pools contained the negative clinical matrix and the potentially interfering substances only. The analyte positive pools contained the negative clinical matrix, the potentially interfering substances and one representative strain of each targeted analyte spiked to a final testing concentration of 3xLoD for each analyte. No cross-reactivity was observed in analyte negative pools in the presence of interfering substances which produced 0% positive results. No interference to analyte positive pools at 3xLoD was observed in the presence of interfering substances which produced 100% positive results. All pools met the acceptance criteria for each condition tested. The results from this study demonstrate that the evaluated substances do not interfere with the Panther Fusion SARS-CoV-2/Flu A/B/RSV assay.

Substance Type	Substance Name	Active Ingredient(s)	Concentration1
Endogenous	Mucin	Purified mucin protein	60 µg/mL
	Blood (human)	N/A	2% v/v
	Neo-Synephrine®	Phenylephrine	15% v/v

Table 4: Potentially Interfering Substances

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SARS-CoV-2/Flu A/B/RSV Assay Performance			Panther Fusion®
Nasal sprays ordrops	Anefrin	Oxymetazoline	15% v/v
	Saline	Sodium chloride	15% v/v
	Ventolin HFA®2	Albuterol	45 ng/mL
	QVAR® BeconaseAQ2	Beclomethasone	15 ng/mL
	Dexacort®2	Dexamethasone	12 µg/mL
	Nasacort®	Triamcinolone	5% v/v
Nasal corticosteroids	Flonase®	Fluticasone	5% v/v
	Rhinocort®	Budesonide	5% v/v
	Nasonex®2	Mometasone	0.5 ng/mL
	AEROSPAN®2	Flunisolide	10 µg/mL
Nasal gel	Zicam® (AllergyRelief)	Luffa opperculata, Galphimia,Glauca, Histaminumhydrochloricum, Sulfur	5% v/v
Throat lozenge	Cepacol ExtraStrength	Benzocaine, Menthol	0.7 mg/mL
	Relenza®2	Zanamivir	3.3 mg/mL
Anti-viral drug	TamiFlu®2	Oseltamivir	400 ng/mL
	Virazole®2	Ribavirin	10.5 µg/mL
Antibiotic, nasalointment	Bactroban cream²	Mupirocin	1.6 µg/mL
Antibiotic, systemic	Tobramycin	Tobramycin	33.1 µg/mL

For Regulatory Submission Use Only

¹ v/v: volume by volume.

2 Active ingredients tested.

Cross-Reactivity and Microbial Interference Wet Testing

The purpose of this study was to demonstrate that the Panther Fusion SARS-CoV-2/Flu A/B/RSV assay does not cross-react with genetically closely related microorganisms or microorganisms commonly encountered in a respiratory specimen. Microorganism wet testing consisted of preparing live or inactivated organisms in processed negative clinical NP swabs VTM/UTM matrix. In total, 41 microorganisms, including bacteria, fungi, and viruses were evaluated. Bacterialfungi were tested at 10° CFU/mL and viruses were tested at 105 TCID56/mL, except where noted. In vitro transcripts (IVTs) corresponding to the ORF lab regions targeted by the assay or SARS coronavirus HKU1 were used to evaluate cross-reactivity. Pools were tested with and without the presence of assay analytes. The analyte negative

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pools contained the negative clinical matrix and the microorganisms. The analyte positive pools contained the negative clinical matrix, microorganisms and one representative strain of each targeted analyte (SARS-CoV-2, Flu A H3N2, Flu B Victoria lineage, RSV B) spiked to a final testing concentration of 3xLoD for each analyte. A minimum of three replicates of each microorganism panel were evaluated using one reagent lot and one Panther Fusion system. All pools had the expected results of no interference or cross reactivity and observed the expected SARS-CoV-2/Flu A/B/RSV results.

Microorganism	Concentration¹	Microorganism	Concentration¹
Adenovirus 1	1x105 TCID50/mL	Bordetella pertussis	1x106 CFU/mL
Adenovirus 7a	1x105 TCID50/mL	Candida albicans	1x106 CFU/mL
CMV Strain AD 169	1x104 TCID50/mL	Chlamydophila pneumoniae	1x106 IFU/mL
Human coronavirus 229E	1x104 TCID50/mL	Corynebacterium diphtheriae	1x106 CFU/mL
Human coronavirus NL63	1x104 TCID50/mL	Escherichia coli	1x106 CFU/mL
Human coronavirus OC43	1x105 TCID50/mL	Haemophilus influenzae	1x106 CFU/mL
Epstein-Barr virus (EBV)	1x106 copies/mL	Lactobacillus plantarum	1x106 CFU/mL
Enterovirus (e.g., EV68)	1x105 TCID50/mL	Legionella pneumophila	1x106 CFU/mL
Human coronavirus HKU12	1x106 copies/mL	Moraxella catarrhalis	1x105 CFU/mL
Human Metapneumovirus(hMPV)	1x105 TCID50/mL	Mycobacterium tuberculosis	1x109 rRNA copies/mL
HPIV-1	1x105 TCID50/mL	Mycoplasma pneumoniae	1x109 rRNA copies/ mL
HPIV-2	1x105 TCID50/mL	Neisseria spp	1x106 CFU/mL
HPIV-3	1x105 TCID50/mL	Neisseria meningitides	1x106 CFU/mL
HPIV-4	1x104 TCID50/mL	Neisseria mucosa	1x106 CFU/mL
Measles	1x104 TCID50/mL	Pneumocystis jirovecii	1x106 CFU/mL
MERS-Coronavirus	5x104 TCID50/ mL	Pseudomonas aeruginosa	1x106 CFU/mL
Mumps virus	1x105 TCID50/mL	Staphylococcus aureus	1x106 CFU/mL
Rhinovirus 1A	1x104 TCID50/mL	Staphylococcus epidermidis	1x106CFU/mL
SARS coronavirus 12	1x106 copies/mL	Streptococcus pneumoniae	1x106 CFU/mL
Varicella Zoster Virus	1x103 TCID50/mL	Streptococcus pyogenes	1x106 CFU/mL

Table 5: Cross Reactivity and Microbial Interference Microorganisms

1CFU = Colony Forming Units; IFU = Inclusion Forming Units; TCID50 = Median Tissue Culture Infectious Dose.

3 Cultured virus and whole genome purified nucleic acid for Human HKU1 and SARS-coronavirus are not readily available. HKU1 and SARScoronavirus in vitro transcript (IVT) corresponding to the ORF la gene regions targeted by the assay were used to evaluate cross-reactivity and microbial interference.

31x109 rRNA copies/mL is equivalent to ~2x105 CFU/mL.

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Analytical Reactivity - Strain Wet Testing

The purpose of this study was to verify that the Panther Fusion SARS-CoV-2/Flu A/B/RSV Assay on the Panther Fusion system can detect multiple strains of SARS-CoV-2, Flu A, Flu B, RSV B. Multiple strains of each assay target were tested at ~3x the assay Limit of Detection (LoD). For strains not detected at 3x LoD, additional testing at higher concentrations was performed until 100% positivity was observed. Samples consisted of 15 strains of SARS-CoV-2, 28 strains of Flu A (13 H1N1, 14 H3N2 and 1 H5N1), 17 strains of Flu B (8 Victoria, 8 Yamagata and 1 unknown), 4 strains of RSV A, and 3 strains of RSV B. The Panther Fusion SARS-CoV-2/Flu A/B/RSV Assay demonstrates SARS-CoV-2, Flu A, Flu B and RSV strain wet testing inclusivity for the strains tested at the concentrations illustrated in Table 6.

Description	Subtype	Concentration	SARS-CoV-2	Flu A	Flu B	RSV
USA-WA1/2020*	SARS-CoV-2	0.09 TCID50/mL	+	-	-	-
USA-CA1/2020	SARS-CoV-2	0.09 TCID50/mL	+	-	-	-
USA-AZ1/2020	SARS-CoV-2	0.09 TCID50/mL	+	-	-	-
USA-WI1/2020	SARS-CoV-2	0.09 TCID50/mL	+	-	-	-
USA/OR-OHSU-PHL00037/2021 B.1.1.7	SARS-CoV-2	0.09 TCID50/mL	+	-	-	-
Uganda/MUWRP-20200195568/2020/A.23.1	SARS-CoV-2	0.09 TCID50/mL	+	-	-	-
USA/PHC658/2021 B.1.617.2	SARS-CoV-2	0.09 TCID50/mL	+	-	-	-
USA/MD/HP05285/2021 B.1.617.2	SARS-CoV-2	0.09 TCID50/mL	+	-	-	-
USA/CA/VRLC009/2021 B.1.427	SARS-CoV-2	0.09 TCID50/mL	+	-	-	-
USA/CA/VRLC012/2021 P.2	SARS-CoV-2	0.30 TCID50/mL	+	-	-	-
USA/MD-HP03056/2021 B.1.525	SARS-CoV-2	0.30 TCID50/mL	+	-	-	-
USA/CA-Stanford-16_S02/2021 B.1.617.1	SARS-CoV-2	0.09 TCID50/mL	+	-	-	-
Description	Subtype	Concentration	SARS-CoV-2	Flu A	Flu B	RSV
Peru/un-CDC-2-4069945/2021 C.37	SARS-CoV-2	0.09 TCID50/mL	+	-	-	-
USA/MD-HP20874/2021 B.1.1.529	SARS-CoV-2	0.09 TCID50/mL	+	-	-	-
USA/GA-EHC-2811C/2021 B.1.1.529	SARS-CoV-2	0.09 TCID50/mL	+	-	-	-
A/Brisbane/02/18*	Flu A (H1N1)	0.18 TCID50/mL	-	+	-	-
A/Michigan/45/2015	Flu A (H1N1)	0.18 TCID50/mL	-	+	-	-
A/Christ Church/16/2010	Flu A (H1N1)	180 TCID50/mL	-	+	-	-
A/Kentucky/2/06	Flu A (H1N1)	0.60 TCID50/mL	-	+	-	-
A/Solomon Islands/03/06	Flu A (H1N1)	0.60 TCID50/mL	-	+	-	-
A/Guangdong-maonan/1536/2019	Flu A (H1N1)	180 TCID50/mL	-	+	-	-
A/Taiwan/42/2006	Flu A (H1N1)	0.60 TCID50/mL	-	+	-	-
A/Henan/8/05	Flu A (H1N1)	0.60 TCID50/mL	-	+	-	-
A/Hawaii/15/01	Flu A (H1N1)	183 TCID50/mL	-	+	-	-
A/California/07/2009	Flu A (H1N1)	0.18 TCID50/mL	-	+	-	-
A/Hawaii/66/2019	Flu A (H1N1)	180 CEID50/mL	-	+	-	-
A/Indiana/02/2020	Flu A (H1N1)	60 CEID50/mL	-	+	-	-
A/Michigan/45/2015 pdm09-likevirus	Flu A (H1N1)	0.60 TCID50/mL	-	+	-	-
A/Kansas/14/17*	Flu A (H3N2)	0.33 TCID50/mL	-	+	-	-
A/Arizona/45/2018	Flu A (H3N2)	3.3 FFU/mL	-	+	-	-
A/New York/21/2020	Flu A (H3N2)	3.3 FFU/mL	-	+	-	-
A/Hong Kong/45/2019	Flu A (H3N2)	3.3 FFU/mL	-	+	-	-
A/Singapore/INFIMH-16-0019/2016	Flu A (H3N2)	110 CEID50/mL	-	+	-	-
A/Hong Kong/2671/2019	Flu A (H3N2)	112 TCID50/mL	-	+	-	-
A/Hiroshima/52/05	Flu A (H3N2)	1.1 TCID50/mL	-	+	-	-
Description	Subtype	Concentration	SARS-CoV-2	Flu A	Flu B	RSV
A/Costa Rica/07/99	Flu A (H3N2)	113 TCID50/mL	-	+	-	-
A/Port Chalmers/1/73	Flu A (H3N2)	1.1 TCID50/mL	-	+	-	-
A/Brazil/113/99	Flu A (H3N2)	1.1 TCID50/mL	-	+	-	-
A/Perth/16/2009	Flu A (H3N2)	0.33 TCID50/mL	-	+	-	-
A/Texas/50/2012	Flu A (H3N2)	0.33 TCID50/mL	-	+	-	-
A/Hong Kong/4801/2014	Flu A (H3N2)	1.1 TCID50/mL	-	+	-	-
A/Indiana/08/2011	Flu A (H3N2)	1.1 TCID50/mL	-	+	-	-
A/Hong Kong/486/97	Flu A (H5N1)	0.01 ng/mL	-	+	-	-
B/Washington/02/2019*	Flu B (Victoria)	0.09 TCID50/mL	-	-	+	-
B/Colorado/06/2017	Flu B (Victoria)	0.09 TCID50/mL	-	-	+	-
B/Florida/78/2015	Flu B (Victoria)	0.30 TCID50/mL	-	-	+	-
B/Alabama/2/17	Flu B (Victoria)	0.09 TCID50/mL	-	-	+	-
B/Ohio/1/2005	Flu B (Victoria)	0.30 TCID50/mL	-	-	+	-
B/Michigan/09/2011	Flu B (Victoria)	33 TCID50/mL	-	-	+	-
B/Hawaii/01/2018 (NA D197N)	Flu B (Victoria)	0.901 TCID50/mL	-	-	+	-
B/Brisbane/33/08	Flu B (Victoria)	0.09 TCID50/mL	-	-	+	-
B/Phuket/3073/2013*	Flu B (Yamagata)	0.006 TCID50/mL	-	-	+	-
B/Wisconsin/1/2010	Flu B (Yamagata)	21 TCID50/mL	-	-	+	-
B/Utah/9/14	Flu B (Yamagata)	0.006 TCID50/mL	-	-	+	-
B/St. Petersburg/04/06	Flu B (Yamagata)	0.06 TCID50/mL	-	-	+	-
B/Texas/81/2016	Flu B (Yamagata)	21 TCID50/mL	-	-	+	-
B/Indiana/17/2017	Flu B (Yamagata)	0.601TCID50/mL	-	-	+	-
B/Oklahoma/10/2018	Flu B (Yamagata)	21 TCID50/mL	-	-	+	-
B/Massachusetts/02/2012	Flu B (Yamagata)	0.22 TCID50/mL	-	-	+	-
Description	Subtype	Concentration	SARS-CoV-2	Flu A	Flu B	RSV
B/Lee/40	Flu B	0.09 TCID50/mL	-	-	+	-
RSV-A/2006 Isolate*	RSVA	0.06 TCID50/mL	-	-	-	+
RSV A/4/2015 isolate #1	RSVA	0.06 TCID50/mL	-	-	-	+
RSV A/A2	RSVA	0.06 TCID50/mL	-	-	-	+
RSV A/12/2014 isolate #2	RSVA	0.06 TCID50/mL	-	-	-	+
RSV-B/CH93(18)-18*	RSVB	0.30 TCID50/mL	-	-	-	+
RSV B/3/2015 isolate #1	RSVB	0.09 TCID50/mL	-	-	-	+
RSV B/9320	RSVB	0.09 TCID50/mL	-	-	-	+

Table 6: Analytical Reactivity Summary for SARS-CoV-2, Flu A and Flu B and RSV Strains

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For Regulatory Submission Use Only

SARS-CoV-2/Flu A/B/RSV Assay Performance

Panther Fusion®

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For Regulatory Submission Use Only

SARS-CoV-2/Flu A/B/RSV Assay Performance

Panther Fusion®

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*Strain used to establish LoD.

In silico analysis showed 100% homology to amplification region. Virus stock degradation or error in TCID50/mL quantification may have impacted the concentration at 100% detection.

2 In silico analysis identified a single mismatch in the forward and reverse primers for A/Hong Kong/2671/2019 and a single mismatch in the reverse primer of B/Massachusetts/02/2012. Due to the mismatches, amplification, and detection are not expected to be impacted. Virus stock degradation or error in TCID50/mL quantification may have impacted the concentration at 100% detection. Sequence of strain in targeted amplification regions are not available in NCBI or GISAID to further evaluate sensitivity.

Analytical Reactivity - In Silico Inclusivity

The purpose of this analysis is to document SARS-CoV-2, Flu A, Flu B, RSV A, and RSV B strain and isolate inclusivity of the Panther Fusion SARS-CoV-2/Flu A/B/RSV assay. Inclusivity was assessed based on in silico analysis of the assay forward primers, reverse primers, and probe oligos compared to available sequences in the GISAID and NCBI gene databases. For SARS-CoV-2, sequences on or before 6/25/2022 were evaluated (10% of the database was randomly sampled for analysis); for Flu A, Flu B, RSV A and RSV B, sequences from between January 01, 2015 and February 15, 2022 were evaluated. Exclusion criteria removed all non-human isolates and partial sequences that do not span the amplicon regions of the assay. Sequence alignment for the analysis was performed using MAFFT (multiple alignment using fast Fourier transform) and MSAE (Hologic Multiple Sequence Alignment Editor) alignment programs. Based on in silico analysis of GISAID and NCBI sequences available up to June 25, 2022 for SARS-CoV-2 (10% random sampling), the Panther Fusion SARS-CoV-2/Flu A/B/RSV assay is predicted to detect all 934,493 SARS-CoV-2 sequences evaluated. The sequences evaluated included lineages and variants of concern (VOC) or variants

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under investigation (VUI) that may have important epidemiological, immunological, or pathogenic properties from a public health perspective, such as Delta and Omicron variants. All lineages and variants of public health interest identified as of June 25, 2022 are predicted to be detected; new sequences and variants will continue to be monitored for impacts on detection by the Panther Fusion SARS-CoV-2/Flu A/B/RSV assay.

Based on in silico analysis of all sequences available from January 01, 2015 to February 15, 2022 in GISAID and NCBI databases, the Panther Fusion SARS-CoV-2/Flu A/B/RSV assay is predicted to detect ≥99.998% of 88,128 Flu A, ≥99.94% of 31,801 Flu B, ≥98.12% of 1,599 RSV A, and ≥98.23% of 1,240 RSV B sequences evaluated.

Cross-Reactivity in the Presence of Microorganisms – In Silico

The purpose of this study was to demonstrate that the Panther Fusion SARS-CoV-2/Flu A/B/RSV assay does not cross-react with genetically closely related or commonly encountered microorganisms through in silico BLAST analysis. The 143 microorganisms were evaluated with in silico BLAST analysis using multiple representative strains from GenBank. BLAST results were analyzed for homology to the amplification primers (forward and reverse primer) and probes for the SARS-CoV-2 (Region 1 and Region 2), Flu A (Region A1 and Region A2), Flu B, RSV A, and RSV B targets included in the assay. Any microorganism sequence showing ≥80% homology to a single primer or probe was further evaluated for homology of ≥50% to the other target region primers and probes. If homology of a primer pair was detected, additional analysis included determining the proximity of the pair and evaluation the probability of generating and/or detecting amplicon from the microorganism sequences.

A total of 545 GenBank sequences for the 143 non-target microorganisms were analyzed. Homology to each of the sequences was analyzed per primer or probe oligo for the SARS-CoV-2, Flu A, Flu B, and RSV assay target regions. The in-silico BLAST analysis of the primers and probes of the Panther Fusion SARS-CoV-2/Flu A/B/RSV assay demonstrates that there is no predicted cross reactivity or interference to genetically closely related or commonly encountered respiratory microorganisms, with the exception of S. marcescens, which had a possibility of low amplification without detection. Wet testing in processed negative clinical NP swab VTM/UTM matrix of each target at 3X LoD in the presence of this organism at 106 CFU/mL demonstrated that no interference was observed.

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Analytical Exclusivity

The purpose of this study was to demonstrate the influenza analytical specificity in relation to influenza types and subtypes not intended to be detected with the Panther Fusion SARS CoV-2/Flu A/B/RSV Assay on the Panther Fusion System. Two non-indicated influenza types, influenza type C and influenza type D, were tested. The Panther Fusion SARS-CoV-2/Flu A/B/RSV Assay did not detect either.

Within Laboratory Precision and Repeatability

This study demonstrated the within laboratory precision and repeatability of results generated by the Panther Fusion SARS-CoV-2/Flu A/B/RSV Assay on the Panther Fusion system when evaluated under different conditions of variability (i.e., operators, days, reagent lots, instruments). The panels tested consisted of a negative panel, a low level (2xLoD) SARS-CoV-2 & Flu A (H3N2) positive panel, a moderate level (5x LoD) SARS-CoV-2 & Flu A positive panel, a low level (2x LoD) Flu B (Victoria lineage) & RSV B positive panel, and a moderate level (5x LoD) Flu B & RSV B positive panel. Panels were prepared by spiking viruses into processed negative nasopharyngeal (NP) swab VTM/UTM matrix processed with STM (1:1.56 ratio). Two operators tested 2 replicates of each panel in 2 runs per day for each of the 3 reagent lots across 3 Panther Fusion systems for a total of 12 days of testing. In total, 96 measurements were performed for each panel member. All low (2x) and moderate (5x) panel members were 100% positive for the spiked target analytes. The negative panel member was 0.0% positive for Flu A. Flu B. and RSV. while 1.0% positive for SARS-CoV-2 (1/96) An overall total signal variability of <2.04% was observed.

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Table 7: Signal Variability of the Panther Fusion SARS-CoV-2/Flu A/B/RSV Assay by Panel Member

Panel	Description	Analyte	Agreement/N	Agreement %	Mean CT	SD	CV(%)	Between Lots		SD	CV(%)	Between Instrument		SD	CV(%)	Between Operators		SD	CV(%)
1	Neg	InternalControl	95/96	99**	33.7	0.19	0.57	0.08	0.23	0.00	0.00	0.00	0.00	0.21	0.62	0.29	0.86	0.42	1.23
2	SARS CoV-2/Flu A	Flu A	96//96	100	35.1	0.33	0.93	0.06	0.17	0.00	0.00	0.00	0.00	0.30	0.85	0.56	1.59	0.72	2.04
	Low Pos	SARS-CoV-2	96//96	100	35.9	0.00	0.00	0.13	0.36	0.00	0.00	0.00	0.00	0.00	0.00	0.60	1.67	0.61	1.71
3	Flu B/RSV	Flu B	96//96	100	36.0	0.14	0.40	0.09	0.25	0.00	0.00	0.00	0.00	0.00	0.00	0.36	0.99	0.39	1.09
	Low Pos	RSV	96//96	100	36.1	0.12	0.33	0.28	0.77	0.00	0.00	0.00	0.00	0.37	1.04	0.53	1.46	0.71	1.97
4	SARS CoV-2/Flu A	Flu A	96//96	100	33.9	0.23	0.66	0.00	0.00	0.00	0.00	0.19	0.56	0.00	0.00	0.47	1.37	0.55	1.63
	Mod Pos	SARS-CoV-2	96//96	100	34.7	0.21	0.62	0.16	0.45	0.06	0.17	0.00	0.00	0.00	0.00	0.45	1.30	0.52	1.51
5	Flu B/RSV	Flu B	96//96	100	34.7	0.15	0.44	0.00	0.00	0.00	0.00	0.00	0.00	0.06	0.18	0.28	0.80	0.32	0.93
	Mod Pos	RSV	96//96	100	34.5	0.10	0.30	0.18	0.51	0.00	0.00	0.00	0.00	0.00	0.00	0.40	1.15	0.44	1.29

*Agreement to expected panel positivity result.

**One SARS-CoV-2 false positive result was obtained for the negative panel member.

Low Pos = Both targets are 2X LoD.

Mod Pos = Both targets are 5X LoD.

Note: Variability from some factors may be numerically the variability due to those factors is very small. When this occurs, SD=0 and CV=0%,

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Competitive Interference/Co-Infection

This study evaluated potential competitive interference for paired low/high target combinations of SARS-CoV-2, Flu A, Flu B, RSV A, and RSV B cultured virus in the Panther Fusion SARS-CoV-2/Flu A/B/RSV Assay on the Panther Fusion system. SARS-CoV-2, Flu A H3N2, Flu B Victoria, RSV A, and RSV B cultured viral targets were spiked into processed negative clinical NP VTM/UTM matrix in STM at multiple target combinations of low (3xLoD) and high concentrations (up to 1x104 TCIDsomL) and tested at 3 replicates per panel. If less than 100% positivity was observed in the low target, the high target was diluted from 1x104 TCID50/mL until 100% positivity was achieved.

SARS-CoV-2 at 3xLoD was 100% positive when paired with Flu A, Flu B, and RSV A at 1 x104 TCID50/mL and RSV B at 30 TCID50/mL. Flu A at 3xLoD was 100% positive when paired with Flu B and RSV A at 1x104 TCID50/mL, SARS-CoV-2 at 100 TCID50/mL and RSV B at 30 TCID50/mL. Flu B at 3xLoD was 100% positive when paired with SARS-CoV-2. Flu A. RSV A at 1x104 TCIDsy/mL and RSV B at 1x103 TCIDsomL. RSV A at 3xLoD was 100% positive when paired with SARS-CoV-2, Flu A, and Flu B at 1x104 TCIDs0/mL. RSV B at 3xLoD was 100% positive when combined with SARS-CoV-2, Flu A, and Flu B at 1x104 TCID50/mL.

All targets at 3xLoD maintained positivity when paired with other assay targets at 1x104 TCIDsomL with the exception of the following: SARS-CoV-2 and Flu A each in the presence of RSV B at 3x10' TCID50/mL, Flu B in the presence of RSV B at 1x103 TCID50/mL and Flu A in the presence of SARS-CoV-2 at 1x102 TCID50/mL.

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Table 8: Competitive Interference

Target 1			Target 2	SARS-CoV-2% detected	Flu A% detected	Flu B% detected	RSV% detected
Virus	3X LoDVirus(TCID50/mL)		HighConcentration(TCID50/mL)
		Flu A	1.0E+4	100%	100%	0%	0%
SARS-CoV-2	9.0E-2	Flu B	1.0E+4	100%	0%	100%	0%
		RSV A	1.0E+4	100%	0%	0%	100%
		RSV B	3.0E+1	100%	0%	0%	100%
		SARS-CoV-2	1.0E+2	100%	100%	0%	0%
Flu A	3.3E-1	Flu B	1.0E+4	0%	100%	100%	0%
		RSV A	1.0E+4	0%	100%	0%	100%
		RSV B	3.0E+1	0%	100%	0%	100%
		SARS-CoV-2	1.0E+4	100%	0%	100%	0%
Flu B	9.0E-2	Flu A	1.0E+4	0%	100%	100%	0%
		RSV A	1.0E+4	0%	0%	100%	100%
		RSV B	1.0E+3	0%	0%	100%	100%
		SARS-CoV-2	1.0E+4	100%	0%	0%	100%
RSV A	6.0E-2	Flu A	1.0E+4	0%	100%	0%	100%
		Flu B	1.0E+4	0%	0%	100%	100%
		SARS-CoV-2	1.0E+4	100%	0%	0%	100%
RSV B	9.0E-2	Flu A	1.0E+4	0%	100%	0%	100%
		Flu B	1.0E+4	0%	0%	100%	100%

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Hologic RespDirect Collection Device Equivalency/Swab Equivalency

The purpose of this study was to demonstrate equivalency between the Hologic RespDirect Collection Kit and a control collection device, consisting of a control nasopharyngeal (NP) swab in UTM. A swab equivalency study was also performed to demonstrate equivalency between the Hologic RespDirect swab and a control NP swab for absorption and elution of targets.

Paired individual nasopharyngeal swab specimens were collected with Hologic RespDirect Collection Kits and control collection kits, consisting of Copan Universal Transport Medium (UTM) and Copan NP swab. Paired individual donor NP swabs from 25 symptomatic donors each were co-spiked with SARS-CoV-2, Flu A, Flu B and RSV at 2x and 5x LoD respectively. In addition to this, paired individual donor NP swabs from 25 donors each were spiked to 2x and 5x LoD respectively for SARS-CoV-2 target only.

For the swab equivalency study, Hologic RespDirect swab and Copan NP swab were dipped into eSTM solution that was co-spiked with SARS-CoV-2, Flu A (H3N2), Flu B (Victoria lineage), and RSV B for absorption of targets. The wet swabs were then transferred into an unspiked Hologic RespDirect tube for elution of targets and tested in the Panther Fusion SARS-CoV-2/Flu A/B/RSV assay. The target concentration in the final Hologic RespDirect tube being tested after elution of targets from the swabs was at 4x LoD. A total of 20 swabs were used per swab type. The results from the swab equivalency study demonstrated equivalency between the Hologic RespDirect swab and a control NP swab for absorption and elution of respiratory virus targets and are illustrated in Table 9.

For the collection device equivalency study, comparable positivity was observed between the two collection devices for all concentrations and targets tested. The results from this study demonstrated equivalency between the Hologic RespDirect Collection Kit and a control collection device for use in the Panther Fusion SARS-CoV-2/Flu A/B/RSV assay and are illustrated in Table 10.

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Table 9: Positivity Results Summary from the Swab Device Equivalency Study

	SARS-CoV-2 (ROX)		Flu A (FAM)		Flu B (Red647)		RSV (HEX)
Condition	N Pos. / NValid	% Pos	N Pos. / NValid	% Pos	N Pos. / NValid	% Pos	N Pos. / NValid	% Pos
Copan NP Swab	20/20	100.0%	20/20	100.0%	20/20	100.0%	20/20	100.0%
RespDirect Swab	20/20	100.0%	20/20	100.0%	20/20	100.0%	20/20	100.0%

Table 10: Results of negative and contrived panels composed of paired individual donor NP clinical specimens, collected with each collection device spiked with SARS-CoV-2, Flu A, Flu B, and RSV

Analyte	SampleConcentration	N per CollectionDevice	VTM/UTM%Positive	HologicRespDirect %Positive
None(negativesample)	0	181	0	0
SARS-CoV-2	2X LoD	50	100	98
	5X LoD	50	100	100
Flu A	2X LoD	25	100	100
	5X LoD	25	100	100
Flu B	2X LoD	25	100	100
	5X LoD	25	100	100
RSV	2X LoD	25	100	100
	5X LoD	25	100	100

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Carry-over/Cross-contamination

The purpose of this study was to demonstrate that no carry-over or cross-contamination is observed from specimens collected in eSTM using the Hologic RespDirect Collection Kit when tested with the Panther Fusion SARS-CoV-2/Flu A/B/RSV assay.

Unique pools of negative clinical eSTM were assembled. Positive panels were prepared by spiking Flu A H3N2 at high titer (1x104 TCID50/mL) into one pool. Multiple identical positive and negative panels were prepared. 100 replicates of negative panel were tested per baseline run. One baseline run and five "checkerboard" runs were completed on each of two Panther Fusion instruments. Checkerboard runs were set up by alternating 30 positive and 30 negative panel in the sample racks.

Results showed 100% Flu A positivity for all positive panels, and 0% Flu A positivity in the negative panels. The Panther Fusion SARS-CoV-2/Flu A/B/RSV Assay using the Hologic RespDirect sample collection kit does not show carry-over or cross-contamination between samples on the same or adjacent sample racks.

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Panther Fusion System Assay Performance

Clinical Performance Study

Prospective Clinical Study

This study was performed to demonstrate clinical performance characteristics for the Panther Fusion SARS-CoV-2/Flu A/B/RSV assay. A prospective multicenter study was conducted using remnant NP swab specimens from male and female individuals of all ages exhibiting signs and/or symptoms of respiratory infection consistent with COVID-19, influenza, or RSV. Five participating US pediatric/adolescent, private and/or university hospitals prospectively provided remnant NP swab specimens collected during portions of the 2020-2021 and 2021-2022 respiratory infection seasons. These specimens were tested at 3 US sites with the Panther Fusion SARS-CoV-2/Flu A/B/RSV assay.

The Panther Fusion SARS-CoV-2/Flu A/B/RSV assay was evaluated for SARS-CoV-2 performance by comparing the candidate device testing results to a composite comparator algorithm (CCA) consisting of two highly sensitive US FDA EUA SARS-CoV-2 molecular tests and a validated PCR followed by bi-directional sequencing (PCR/BDS) assay. A final CCA result was assigned when two of the three composite comparator assays were in concordance. The comparator method utilized to establish performance for the Flu A, Flu B, and RSV targets was a US FDA-cleared molecular Flu A/B/RSV assay.

Of the 1949 specimens enrolled during the study, 1056 were collected between January 2022 and March 2022, while the remaining 893 were collected between November 2020 and March 2021. Forty-five (45) of these specimens were withdrawn; mishandling during transport was the most common reason for withdrawal. A total of 1905 NP swab specimens were tested in valid Panther Fusion SARS-CoV-2/Flu A/B/RSV assay runs, including 12 (0.6%) with initial invalid results. Upon retest, 8 of the 12 specimens yielded valid results and 4 yielded invalid results, for a total of 1901 (99.8%) specimens with final valid results. One specimen with valid Panther Fusion assay results was withdrawn following testing with the Panther Fusion assay upon determination that the sample had not been stored according to the Panther Fusion assay package insert instructions. The final data set consisted of 1900 evaluable NP swab specimens; not all were evaluable for all analytes. For the SARS-CoV-2 target, 13 of these 1900 NP swab specimens were excluded from analysis due to

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unknown infection status obtained from the CCA tests. A total of 1887 prospective samples were evaluated for SARS-CoV-2, including 790 (41.9%) tested fresh and 1097 (58.1%) tested after freezing. For the Flu A, Flu B, and RSV targets, 63 specimens were excluded from analysis due to obtaining an invalid result from the comparator test. A total of 1837 valid prospective specimens were evaluated for Flu A, Flu B, and RSV, including 798 (43.4%) tested fresh and 1039 (56.6%) tested after freezing. Demographic information for the 1900 evaluable prospective specimens is shown in Table 11.

		N (%)
Total		1900 (100)
Sex	Female	1049 (55.2)
	Male	850 (44.7)
	Unknown	1 (0.1)
Age Group	<5 years	388 (20.4)
	5-21 years	435 (22.9)
	22 to 40 years	372 (19.6)
	41 to 60 years	326 (17.2)
	> 60 years	379 (19.9)

Table 11: Summary of Subject Demographics for Evaluable Prospectively Collected Specimens

The performance of the Panther Fusion SARS-CoV-2/Flu A/B/RSV assay with prospective specimens is summarized in Table 12. Positive Percent Agreement (PPA) was calculated as 100% × (TP / (TP + FN)). True positive (TP) indicates that both the Panther Fusion SARS-CoV-2/Flu A/B/RSV assay and the comparator method had a positive result for the specific analyte. and false negative (FN) indicates that the Panther Fusion SARS-CoV-2/Flu A/B/RSV 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 Panther Fusion SARS-CoV-2/Flu A/B/RSV assay and the comparator method had negative results, and false positive (FP) indicates that the Panther Fusion SARS-CoV-2/Flu A/B/RSV assay was positive while the comparator result was negative. Specimens that obtained discordant results underwent additional testing with a US FDA EUA SARS-CoV-2/Flu A/B/RSV molecular test, volume permitting.

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Analyte		Positive Percent Agreement			Negative Percent Agreement
		TP/(TP+FN)	%	95% CI	TN/(TN+FP)	%	95% CI
SARS-CoV-2	Fresh	60/64	93.8	85.0-97.5	716/726	98.6	97.5-99.3
	Frozen	318/326	97.5	95.2-98.8	758/771	98.3	97.1-99.0
	Overall	378/3901	96.9	94.7-98.2	1474/14972	98.5	97.7-99.0
Flu A	Fresh	98/100	98.0	93.0-99.5	696/698	99.7	99.0-99.9
	Frozen	23/23	100	85.7-100	1013/1016	99.7	99.1-99.9
	Overall	121/1233	98.4	94.3-99.6	1709/17144	99.7	99.3-99.9
Flu B	Fresh	0/0	NC	NC	796/798	99.7	99.1-99.9
	Frozen	0/0	NC	NC	1037/1039	99.8	99.3-99.9
	Overall	0/0	NC	NC	1833/18375	99.8	99.4-99.9
RSV	Fresh	11/13	84.6	57.8-95.7	785/785	100	99.5-100
	Frozen	0/0	NC	NC	1039/1039	100	99.6-100
	Overall	11/136	84.6	57.8-95.7	1824/1824	100	99.8-100

Table 12: Panther Fusion SARS-CoV-2/Flu A/B/RSV Assay Performance with Prospectively Collected Specimens

TP - true positive; FN - false negative; TN - true negative: FP - false positive; NC - Not calculable

I Five (5) specimens with false negative SARS-CoV-2 results had sufficient volume remaining for discordant testing. All five specimens were positive for SARS-CoV-2 by a US FDA EUA SARS-CoV-2/Flu A/B/RSV molecular test.

2Eleven (11) specimens with false positive SARS-CoV-2 results had sufficient volume remaining for discordant testing. Seven of the specimens were negative for SARS-CoV-2 by a US. FDA EUA SARS-CoV-2/Flu A/B/RSV molecular test.

3No specimens with false negative Flu A results had sufficient volume remaining for discordant testing.

4Two (2) specimens with false positive Flu A results had sufficient volume remaining for discordant testing. Both specimens were negative for Flu A by a US FDA EUA SARS-CoV-2/Flu A/B/RSV molecular test.

30ne (1) specimen with false positive Flu B result had sufficient volume remaining for discordant testing. This specimen was negative for Flu B by a US FDA EUA SARS-CoV-2/Flu A/B/RSV molecular test.

9No specimens with false negative RSV results had sufficient volume remaining for discordant testing. The specimens had C values of 41.3 and 43.5 with the comparator molecular assay.

Five co-infections were detected by the Panther Fusion SARS-CoV-2/Flu A/B/RSV assay: 4 SARS-CoV-2 positive/Flu A positive and 1 SARS-CoV-2 positive/Flu B positive; 3 of the 4 SARS-CoV-2 / Flu A coinfections were also detected by comparator testing.

Retrospective Clinical Study

Flu B, and RSV were of low prevalence during the prospective clinical study and were therefore not encountered in large enough numbers to adequately demonstrate assay performance. To supplement the results of the prospective specimen population, retrospective specimen testing was performed. This study included 95 preselected archived retrospective NP swab specimens in VTM or UTM that were collected between December 2019 and March 2020. Specimens were selected for enrollment in the study based solely on the historic qualitative positive result. In addition to evaluating Flu B and RSV positive specimens, Flu A positive specimens were included in the study. All known positive specimens underwent confirmatory testing using a US FDA-cleared molecular Flu A/B/RSV assay.

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The 95 samples were distributed uniformly across all three clinical testing sites. Demographic information for the 95 evaluable retrospective specimens is provided in Table 13.

		N (%)
Total		95 (100)
	Female	51 (53.7)
Sex	Male	44 (46.3)
	<5 years	16 (16.8)
	5-21 years	12 (12.6)
Age Group	22 to 40 years	15 (15.8)
	41 to 60 years	16 (16.8)
	> 60 years	36 (37.9)

Table 13: Summary of Subject Demographics for Evaluable Retrospective Specimens

The PPA and NPA of the Panther Fusion SARS-CoV-2/Flu A/B/RSV assay with retrospective were calculated against results from confirmatory testing. Specimens that obtained discordant results underwent additional testing with a US FDA EUA SARS-CoV-2/Flu A/B/RSV molecular test, volume permitting. Table 14 shows the PPA for specimens that were confirmed positive for at least one target analyte.

Table 14: Panther Fusion SARS-CoV-2/ Flu A/B/RSV Assay Clinical Performance in Retrospective Specimens

Analyte	Positive Percent AgreementTP/(TP+FN)	%	95% CI
Flu A	27/291	93.1	78.0-98.1
Flu B	21/222	95.5	78.2-99.2
RSV	47/47	100	92.4-100

TP = true positive; FN = false negative; TN = true negative; FP = false positive

10ne (1) specimen with a false negative Flu A result tested positive for Flu A with a US FDA EUA SARS-CoV-2/Flu AB/RSV molecular test, while one ( ) specimen with a false negative Flu A result tested negative for Flu A with a US FDA EUA SARS-COV-2/Flu A/B/RSV molecular test.

20ne (1) specimen with a false negative Flu B result tested positive for Flu B with a US. FDA EUA SARS-CoV-2/Flu AB/RSV molecular test.

Table 15 shows the NPA for specimens that had a negative result on the comparator assay although they were confirmed positive for one of the other target analytes.

Table 15: Panther Fusion SARS-CoV-2/ Flu A/B/RSV Assay Clinical Performance with Confirmed Negative Specimens

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Analyte	Negative Percent Agreement
	TN/(TN+FP)	%	95% CI
Flu A	66/66	100	94.5-100
Flu B	73/73	100	95.0-100
RSV	48/48	100	92.6-100

FP = false positive; TN = true negative.

Two co-infections were detected by the Panther Fusion SARS-CoV-2/Flu A/B/RSV assay were also detected by comparator testing: 1 Flu A positive and 1 Flu A positive and 1 Flu A positive/RSV positive.

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Reproducibility Study

Panther Fusion SARS-CoV-2/Flu A/B/RSV assay reproducibility was evaluated at three US sites using five panel members. At each of the three sites, two operators at each site completed 5 valid runs each, for a total of 10 valid runs per site, using one reagent kit lot. Samples were tested over approximately 5 days at each site.

Overall, 150 samples were tested at each of the 3 sites, for a total of approximately 450 samples tested. Each panel member had 90 results per panel member component across all sites.

One (1) panel member was negative for SARS-CoV-2, Flu A, Flu B, and RSV (i.e., NP specimen in UTM VTM processed into STM matrix only), 2 panel members were dual-positive for Flu A and SARS-CoV-2and 2 panel members were dual-positive for Flu B and RSV. Each dual positive panel was generated at 2 concentrations, a low positive (1-2x LoD) and a moderate positive (3-5x LoD). The positive panel members were prepared by spiking viral strain stocks into a negative matrix of pooled negative clinical NP specimens in VTM/UTM processed into STM. The negative panel member contained matrix only.

Agreement values (see Table 16) were 100% for all panel member components except the following:

. True negative: 98.9% - One negative panel member had a positive Flu B result.
Flu A low positive: 98.9% One low positive panel member had a negative Flu A result. ●

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Table 16: Reproducibility Study: Overall Agreement of Panther Fusion SARS-CoV-2/Flu A/B/RSV Assay Results with Expected Results

PanelMember	Description	ExpectedResult	Agreed/n	Agreement (%)95% CI
1	True Negative*	Negative	89/90	98.9 (94.0-99.8)
2	SARS-CoV-2 Low Positive	Positive	90/90	100 (95.9-100)
4	SARS-CoV-2 Moderate Positive	Positive	90/90	100 (95.9-100)
2	Flu A Low Positive	Positive	89/90	98.9 (94.0-99.8)
4	Flu A Moderate Positive	Positive	90/90	100 (95.9-100)
3	Flu B Low Positive	Positive	90/90	100 (95.9-100)
5	Flu B Moderate Positive	Positive	90/90	100 (95.9-100)
3	RSV Low Positive	Positive	90/90	100 (95.9-100)
5	RSV Moderate Positive	Positive	90/90	100 (95.9-100)

*There was one false positive Flu B result on the negative panel.

All moderate positive panel member components had total %CV less than or equal to 1.53 and total SD values less than or equal to 0.51. Low positive panel member components for SARS-CoV-2, Flu B, and RSV had total %CV less than or equal to 1.82 and total SD values less than or equal to 0.65. The total %CV and SD for the Flu A low positive panel member component were 10.92% and 3.77, respectively, due to the false negative result for 1 sample. For all positive panel member components, the within-run factor (i.e., random error) was the largest contributor to total variability (see Table 17).

Target Virus	PanelMember	Description	N	Mean Ct	Contribution to Total VarianceSD (%CV)
					BetweenSites	BetweenOperators/Runs1	BetweenDays	WithinRuns	Total VarianceSD (%CV)
SARS-CoV-2	2	Low Pos	90	35.53	0.24(0.68)	0.18(0.50)	0.19(0.52)	0.49(1.38)	0.60(1.70)
SARS-CoV-2	4	Mod Pos	90	34.15	0.11(0.32)	0.00(0.00)	0.00(0.00)	0.40(1.16)	0.41(1.20)
Flu A	2	Low Pos	90	34.55	0.57(1.66)	0.62(1.81)	0.00(0.00)	3.68(10.64)	3.77(10.92)
Flu A	4	Mod Pos	90	33.55	0.09(0.27)	0.03(0.10)	0.17(0.49)	0.48(1.42)	0.51(1.53)
Flu B	3	Low Pos	90	35.80	0.12(0.35)	0.00(0.00)	0.22(0.60)	0.39(1.10)	0.47(1.30)

Table 17: Reproducibility Study: Signal Variability of the Panther Fusion SARS-CoV-2/Flu A/B/RSV Assay by Positive Panel Component

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5		Mod Pos	90	34.56	0.00(0.00)	0.10(0.29)	0.00(0.00)	0.29(0.83)	0.30(0.88)
RSV	3	Low Pos	90	35.78	0.07(0.20)	0.23(0.65)	0.14(0.39)	0.59(1.64)	0.65(1.82)
5		Mod Pos	90	34.41	0.05(0.14)	0.00(0.00)	0.00(0.00)	0.43(1.25)	0.43(1.26)

Ct = cycle threshold; CV = coefficient of variation; Mod = moderate; Pos = positive.

Note: Variability from some factors may be numerically negative; this can occur if the variability due to those factors is very small. In these cases, SD and %CV are displayed as 0.

'Between-Operator may be confounded with Between-Run; therefore, Between-Run estimates are combined in Between-Operator/Run.

These results indicate that the repeatability and reproducibility of the PF SARS/Flu/RSV assay on the Panther Fusion system are robust in NP samples. These findings support the proposed intended use.

VIII. CONCLUSIONS

The analytical and clinical study results demonstrate that the Panther Fusion SARS-CoV-2/Flu A/B/RSV assay on the Panther Fusion system is substantially equivalent to the predicate device that is currently marketed for the same intended use. Hardware and software verification and validation demonstrate that the Panther Fusion SARS-CoV-2/Flu A/B/RSV assay on the Panther Fusion system will perform as intended.

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.