Predicate Devices

BioFire COVID-19 Test 2 (K211079)

Reference Devices

Not Found

AI/ML (Artificial Intelligence / Machine Learning)

No
The device description and intended use focus on standard real-time RT-PCR technology and automated data interpretation based on predefined rules (CT cutoff values and calling rules). There is no mention of AI or ML in the document.

Therapeutic

No

Explanation: This device is an in vitro diagnostic (IVD) test intended for the qualitative detection of SARS-CoV-2 nucleic acid, aiding in the diagnosis of COVID-19. It does not provide any therapeutic effect or treat any condition.

Diagnostic

Yes

The "Intended Use / Indications for Use" section explicitly states that the device is "intended for use as an aid in the diagnosis of COVID-19."

SaMD (Software as a Medical Device)

No

The device is a kit that includes physical components (assays, controls) and requires specific hardware (real-time PCR instruments) for its primary function of detecting SARS-CoV-2 RNA. While it includes interpretive software, it is not solely software.

IVD (In Vitro Diagnostic)

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

Here's why:

Intended Use: The document explicitly states the device is "intended for the qualitative detection of nucleic acid from SARS-CoV-2 in nasopharyngeal and anterior nasal specimens from individuals with signs and symptoms of respiratory tract infection." This is a diagnostic purpose, aiming to detect the presence of a pathogen in a sample taken from the human body.
Aid in Diagnosis: The intended use also states it is "intended for use as an aid in the diagnosis of COVID-19 if used in conjunction with other clinical observations, epidemiological information and laboratory findings." This clearly positions the device as a tool used in the diagnostic process.
In Vitro Diagnostic Procedures: The intended user is described as "qualified clinical laboratory personnel specifically instructed and trained in the techniques of real-time PCR and in vitro diagnostic procedures." This directly references the use of the device within the context of in vitro diagnostic procedures.
Device Description: The description details a test that analyzes biological specimens (nasopharyngeal and anterior nasal specimens) outside of the body to detect the presence of SARS-CoV-2 RNA. This is the core function of an in vitro diagnostic device.
Clinical Laboratory Use: The intended user and care setting specify use by "qualified clinical laboratory personnel" in "laboratories that are certified under the Clinical Laboratory Improvement Amendments of 1988 (CLIA)... that meet requirements to perform high complexity tests." This is the typical environment for performing IVD tests.
Performance Studies: The document includes detailed performance studies (Limit of Detection, Reproducibility, Clinical Validation, etc.) which are standard requirements for demonstrating the analytical and clinical performance of an IVD.
Predicate Device: The mention of a "Predicate Device(s)" with K numbers (K211079; BioFire COVID-19 Test 2) indicates that this device is being compared to a previously cleared IVD, which is a common part of the regulatory process for new IVDs.

All these points strongly indicate that the TaqPath™ COVID-19 Diagnostic PCR Kit is an In Vitro Diagnostic device.

PCCP (Predetermined Change Control Plan) Authorized

N/A

Overview

Intended Use / Indications for Use

The TaqPath™ COVID-19 Diagnostic PCR Kit is a real-time reverse transcription polymerase chain reaction (RT-PCR) test intended for the qualitative detection of nucleic acid from SARS-CoV-2 in nasopharyngeal and anterior nasal specimens from individuals with signs and symptoms of respiratory tract infection.

The TaqPath™ COVID-19 Diagnostic PCR Kit is intended for use as an aid in the diagnosis of COVID-19 if used in conjunction with other clinical observations, epidemiological information and laboratory findings. The SARS-CoV-2 RNA is generally detectable in upper respiratory (anterior nasal and nasopharyngeal swabs) specimens during the acute phase of infection. Positive results are indicative of the presence of SARS-CoV-2 RNA. Positive results do not rule out bacterial infection or co-infection with other pathogens. The agent detected may not be the definite cause of disease.

Negative results do not preclude SARS-CoV-2 infection and should not be used as the sole basis for patient management decisions.

The TaqPath™ COVID-19 Diagnostic PCR Kit is intended for use by qualified clinical laboratory personnel specifically instructed and trained in the techniques of real-time PCR and in vitro diagnostic procedures.

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

QQX

Device Description

The Applied BioSystems™ TaqPath™ COVID-19 Diagnostic PCR Kit (TaqPath™ COVID-19 Diagnostic PCR Kit) includes the assays and controls for a multiplex real-time RT-PCR test for the qualitative detection of RNA from SARS-CoV-2 in nasopharyngeal and anterior nasal specimens from individuals with signs and symptoms of respiratory tract infection.

Each kit includes the following components:

Multiplexed assays that contain three primer/probe sets specific to different SARS-CoV-2 . genomic regions and primers/probes for bacteriophage MS2
MS2 Phage Control as an internal process control for nucleic acid extraction ●
TaqPath™ COVID-19 Diagnostic PCR Control as a positive RNA control that contains . targets specific to the SARS-CoV-2 genomic regions targeted by the assays.

The workflow begins with nucleic acid extraction from upper respiratory specimens (nasopharyngeal and anterior nasal swabs) that arrive in the testing site in transport media. Nucleic acids are isolated and purified from the specimens using the MagMAX™ Viral/Pathogen II Nucleic Acid Isolation Kit. Nucleic acid isolation is performed via an automated process using the KingFisher™ Flex Purification System. The nucleic acid is reverse transcribed into cDNA and amplified using the TaqPath™ COVID-19 Diagnostic PCR Kit and one of the following real-time PCR instruments:

Applied Biosystems™ 7500 Fast Dx Real-Time PCR Instrument ●
Applied Biosystems™ QuantStudio™ 5 Dx Real-Time PCR Instrument .

In the process, the probes anneal to three (3) specific SARS-CoV-2 target sequences located between three (3) unique forward and reverse primers for the following genes:

ORF1ab .
N gene ●
. S gene

During the extension phase of the PCR cycle, the 5' nuclease activity of Taq polymerase degrades the probe, causing the reporter dye to separate from the quencher dye, generating a fluorescent signal. With each PCR cycle, additional reporter dye molecules are cleaved from their respective probes, increasing the fluorescent intensity, which is measured at each cycle by the real-time PCR instrument.

Following RT-PCR, the data from the instrument's data collection software are imported into COVID-19 Interpretive Software IVD Edition for analysis and interpretation. After data import, the software analyzes the run data, performs quality check (QC) analysis, and calculates the interpretive results for each sample and control. The imported data and interpretive results for each run are saved as a batch in the software. Results can be exported as CSV files and reports can be generated in PDF format.

Validation of the results is performed automatically by the COVID-19 Interpretive Software based on performance of the positive and negative controls. The following results are automatically generated using the calling rules, plate validity and the CT cutoff values for assay targets:

ORF1ab	N gene	S gene	MS2	Status	Result	Action
NEG	NEG	NEG	NEG	INVALID	NA	RETEST Repeat test by re-extracting the original sample and repeating the RT-PCR. If the repeat result remains invalid, consider collecting a new specimen.
NEG	NEG	NEG	POS	VALID	SARS-CoV-2 Not Detected	REPORT Report the results to the healthcare provider.

ORF1ab	N gene	S gene	MS2	Status	Result	Action
	Only one SARS-CoV-2 target= POS		POS or NEG	VALID	SARS-CoV-2 Inconclusive	RETEST/REPORT 1. Repeat the test by re-extracting the original sample and repeating the RT-PCR. IMPORTANT! Samples with a result of SARS-CoV-2 Inconclusive shall be retested one time. 2. After retesting one time, report the results to the healthcare provider. 3. If the repeat result remains inconclusive, the healthcare provider should conduct additional confirmation testing with a new specimen, if clinically indicated.
	Two or more SARS-CoV-2 targets= POS		POS or NEG	VALID	Positive SARS-CoV-2	REPORT Report the results to the healthcare provider.

A minimum of one negative control and one positive control must be present for each run. Additional negative control wells shall be run for each extraction that is represented on a realtime RT-PCR plate. All control wells must pass for the real-time RT-PCR plate to be considered valid. Recommended actions of retest or report are also provided depending on the results generated.

Mentions image processing

Not Found

Mentions AI, DNN, or ML

Not Found

Input Imaging Modality

Not Found

Anatomical Site

Nasopharyngeal and anterior nasal specimens.

Indicated Patient Age Range

The mean age of subjects in the clinical validation study was 39.3 years (SD: 17.0) and the median age was 37 years (range: 2–87 years). The majority of the subjects were between 5 and 80 years (98.6%); 7 subjects (0.7%) were ≥81 years and 8 subjects (0.8%) were ≤ 5 years.

Intended User / Care Setting

Qualified clinical laboratory personnel specifically instructed and trained in the techniques of real-time PCR and in vitro diagnostic procedures. The use is limited to laboratories that are certified under the Clinical Laboratory Improvement Amendments of 1988 (CLIA), 42 U.S.C. § 263a, that meet requirements to perform high complexity tests.

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

Not Found

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

Not Found

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

a) Limit of Detection:
The LoD for both QuantStudio™ 5 Dx and 7500 Fast Dx instruments was determined to be 50 GCE/mL, as the lowest concentration tested that resulted in ≥ 95% positivity (19/20 replicates).
b) Reproducibility and Within-Laboratory Precision:
Testing was performed at three sites (two external and one internal) using samples prepared at three (3) concentrations: negative (0x LoD), a low positive (1.5x LoD), and a moderate positive (5x LoD). Testing included three (3) sites x two (2) operators x three (3) replicates x three (3) lots of TaqPath™ COVID-19 Diagnostic PCR Kits x five (5) non-consecutive days, for a total of 270 replicates per sample at each test level (0x LoD, 1.5x LoD, 5x LoD) on each PCR instrument model (7500 Fast Dx and QS5 Dx). The kit met acceptance criteria and generated 100% PPA for 5x LoD, 100% PPA for 1.5x LoD and 100% NPA for 0x LoD for each lot tested across all three sites for both instruments.
c) In Use Stability:
The study supports the in-use stability of TaqPath™ COVID-19 Diagnostic PCR Assay Multiplex for up to 10 Freeze-thaw cycles (-30°C to -10°C), TaqPath™ COVID-19 Diagnostic PCR Control Stock (Not Diluted) for up to 48 hours (2°C to 8°C), and TaqPath™ COVID-19 Diagnostic PCR Control Working Stock (Diluted) for up to 48 hours (2°C to 8°C).
d) Transport Simulation:
Demonstrated that the shipping configurations provide adequate thermal and physical protection, maintaining product integrity and performance.
e) Interfering Substances:
27 interfering substances were evaluated. All were considered non-interfering at the tested concentrations, with 100% PPA and 100% NPA for most substances, except for Oxymetazoline (Afrin - No drip, extra moisturizing) and Triamcinolone, where interference was observed at concentrations >5% (v/v), but not at 5% (v/v) (100% PPA, 100% NPA).
f) Cross-Reactivity and Microbial Interference:
46 potentially cross-reactive organisms were tested. No cross-reactivity or microbial interference was observed for all organisms tested.
g) In silico Cross-Reactivity:
In silico analysis (BLAST) was performed against 31,011 virus isolates, 9,996 bacterial isolates, and 14 fungal isolates. None of the isolates matched to two or more individual components of any TaqPath™ COVID-19 Diagnostic PCR Kit assay, indicating no predicted cross-reactivity.
h) Carry-Over Cross-Contamination:
Evaluated using 10 extraction runs of 47 positive and 47 negative replicates. Out of 470 negative samples, 4 false positive calls were observed, resulting in a carry-over cross-contamination rate of 0.85% (4/470).
i) Analytical Reactivity (Inclusivity):
Confirmed by testing 15 variants of SARS-CoV-2. All 15 strains generated 100% analytical inclusivity.
j) In silico Reactivity-Inclusivity:
In silico analysis of SARS-CoV-2 strains from GISAID and GenBank databases through April 2024. Analysis indicates that >99% of the sequences are reactive based on 100% homology or Tm greater than annealing temperature to at least two of the three assay gene targets.
k) Specimen Storage Stability:
The study demonstrated 100% PPA and 100% NPA for all timepoints tested, establishing storage stability: up to 4 hours at ambient temperature (15-30°C), up to 3 days (72 hours) refrigerated (2-8°C), up to 30 days frozen (-30 to -10°C) and up to 30 days frozen at ≤-70°C.
l) Fresh vs Frozen Equivalency Study:
All test conditions produced expected results with 100% NPA at 0x LoD and 100% PPA at 1.5x LoD and 5x LoD. Demonstrated that freeze-thaw up to 2 cycles when stored at -10°C to -30°C and up to 4 cycles when stored at ≤ -70°C does not impact performance.
m) LoD Determination with WHO Standard:
The LoD for the 7500 Fast Dx instrument was 150 IU/mL, and for the QuantStudio™ 5 Dx, it was 50 IU/mL.
n) Clinical Validation Summary:
Multi-site prospective study in nasopharyngeal swabs (NP) and anterior nasal swabs (AN) from 1076 subjects with signs and symptoms of respiratory tract infection.
1053 evaluable NP specimens and 1049 (7500 Fast Dx) / 1052 (QS5 Dx) evaluable AN specimens.
NP Specimens on QuantStudio™ 5 Dx:
PPA: 98.9% (88/89) (95% CI: 93.9% to 99.8%)
NPA: 98.7% (951/964) (95% CI: 97.7% to 99.2%)
NP Specimens on 7500 Fast Dx:
PPA: 98.9% (88/89) (95% CI: 93.9% to 99.8%)
NPA: 98.4% (949/964) (95% CI: 97.4% to 99.1%)
AN Specimens on QuantStudio™ 5 Dx:
PPA: 98.8% (84/85) (95% CI: 93.6% to 99.8%)
NPA: 98.0% (948/967) (95% CI: 97.0% to 98.7%)
AN Specimens on 7500 Fast Dx:
PPA: 98.8% (84/85) (95% CI: 93.6% to 99.8%)
NPA: 97.8% (943/964) (95% CI: 96.7% to 98.6%)

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

Limit of Detection (LoD): 50 GCE/mL (for QuantStudio™ 5 Dx and 7500 Fast Dx with inactivated virus), 50 IU/mL (QuantStudio™ 5 Dx with WHO standard), 150 IU/mL (7500 Fast Dx with WHO standard).

Reproducibility:
PPA Across all Sites: 100% (98.6%, 100%) for 1.5x LoD and 5x LoD on both 7500 Fast Dx and QuantStudio™ 5 Dx.
NPA Across all Sites: 100% (98.6%, 100%) for 0x LoD on both 7500 Fast Dx and QuantStudio™ 5 Dx.

Within-lab Precision (SD and %CV for Ct values):
7500 Fast Dx: Reproducibility SD for ORF1ab 1.5x LoD 0.81, 5x LoD 0.75. N gene 1.5x LoD 0.75, 5x LoD 0.61. S gene 1.5x LoD 0.84, 5x LoD 0.47.
QuantStudio™ 5 Dx: Reproducibility SD for ORF1ab 1.5x LoD 0.56, 5x LoD 0.35. N gene 1.5x LoD 0.67, 5x LoD 0.39. S gene 1.5x LoD 0.87, 5x LoD 0.36.

Interfering Substances:
PPA and NPA are generally 100% (3/3) at tested concentrations.
Oxymetazoline (Afrin - No drip, extra moisturizing) showed 83% (5/6) NPA at 10% (v/v), but 100% (3/3) at 5% (v/v).
Triamcinolone showed 17% (1/6) NPA at 10% (v/v), but 100% (3/3) at 5% (v/v).

Analytical Reactivity (Inclusivity): 100% PPA (3/3) for all 15 SARS-CoV-2 variants tested.

Clinical Performance:
NP Specimens on QuantStudio™ 5 Dx:
PPA: 98.9% (88/89) (95% CI: 93.9% to 99.8%)
NPA: 98.7% (951/964) (95% CI: 97.7% to 99.2%)
NP Specimens on 7500 Fast Dx:
PPA: 98.9% (88/89) (95% CI: 93.9% to 99.8%)
NPA: 98.4% (949/964) (95% CI: 97.4% to 99.1%)
AN Specimens on QuantStudio™ 5 Dx:
PPA: 98.8% (84/85) (95% CI: 93.6% to 99.8%)
NPA: 98.0% (948/967) (95% CI: 97.0% to 98.7%)
AN Specimens on 7500 Fast Dx:
PPA: 98.8% (84/85) (95% CI: 93.6% to 99.8%)
NPA: 97.8% (943/964) (95% CI: 96.7% to 98.6%)

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

BioFire COVID-19 Test 2 (K211079)

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

Not Found

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

Not Found

Regulation Number and Section

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

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

Summary

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

Life Technologies Corporation Ian Mcgill Regulatory Affairs Manager 6055 Sunol Boulevard Pleasanton, California 94566

Re: K233453

Trade/Device Name: Applied Biosystems TaqPath COVID-19 Diagnostic PCR Kit 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: QQX Dated: October 20, 2023 Received: October 20, 2023

Dear Ian Mcgill:

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

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

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

1

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

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

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

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

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

Sincerely.

Image /page/1/Picture/12 description: The image shows the FDA logo in a light blue color. The letters "FDA" are large and bold, and they are the main focus of the image. The logo is simple and recognizable, and it is often used to represent the Food and Drug Administration.

Stefanie Akselrod -S 2024.07.10 13:37:01 -04'00'

For

Himani Bisht. Ph.D. Assistant Director 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

2

Enclosure

3

Indications for Use

510(k) Number (if known) K233453

Device Name

Applied BioSystems™ TaqPath™ COVID-19 Diagnostic PCR Kit

Indications for Use (Describe)

The TaqPath™ COVID-19 Diagnostic PCR Kit is a real-time reverse transcription polymerase chain reaction (RT-PCR) test intended for the qualitative detection of nucleic acid from SARS-CoV-2 in nasopharyngeal and anterior nasal specimens from individuals with signs and symptoms of respiratory tract infection.

The TaqPath™ COVID-19 Diagnostic PCR Kit is intended for use as an aid in the diagnosis of COVID-19 if used in conjunction with other clinical observations, epidemiological information and laboratory findings. The SARS-CoV-2 RNA is generally detectable in upper respiratory (anterior nasal and nasopharyngeal swabs) specimens during the acute phase of infection. Positive results are indicative of the presence of SARS-CoV-2 RNA. Positive results do not rule out bacterial infection or co-infection with other pathogens. The agent detected may not be the definite cause of disease.

Negative results do not preclude SARS-CoV-2 infection and should not be used as the sole basis for patient management decisions.

The TaqPath™ COVID-19 Diagnostic PCR Kit is intended for use by qualified clinical laboratory personnel specifically instructed and trained in the techniques of real-time PCR and in vitro diagnostic procedures.

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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4

510(k) Summary

Submitter:	Life Technologies Corporation (a legal entity of Thermo Fisher Scientific
Address:	6055 Sunol Blvd Building C, Pleasanton, CA 94566
Phone:	(1)760-603-7200
Fax:	N/A
Contact Person:	Ian McGill
Contact email:	Ian.mcgill@thermofisher.com
Contact Phone Number:	1 (480) 536-4400
Date:	July 08, 2024
Proprietary Name:	Applied BioSystems™ TaqPath™ COVID-19 Diagnostic PCR Kit
Measurand:	SARS-CoV-2
Regulation:	866.3981
Classification:	Class II
Product Code:	QQX
Panel:	MI - Microbiology
Predicate Device:	BioFire COVID-19 Test 2 (K211079)

Life Technologies Corporation is a legal entity of Thermo Fisher Scientific. The Applied BioSystems™ TaqPath™ COVID-19 Diagnostic PCR Kit (TaqPath™ COVID-19 Diagnostic PCR Kit) includes the assays and controls for a multiplex real-time RT-PCR test for the qualitative detection of RNA from SARS-CoV-2 in nasopharyngeal and anterior nasal specimens from individuals with signs and symptoms of respiratory tract infection.

Each kit includes the following components:

Multiplexed assays that contain three primer/probe sets specific to different SARS-CoV-2 . genomic regions and primers/probes for bacteriophage MS2
MS2 Phage Control as an internal process control for nucleic acid extraction ●
TaqPath™ COVID-19 Diagnostic PCR Control as a positive RNA control that contains . targets specific to the SARS-CoV-2 genomic regions targeted by the assays.

5

Assay workflow

Extract RNA from a patient sample

Set up the 7500 Fast Dx Real-Time PCR Instrument OR Set up the QuantStudio™ 5 Dx Real-Time PCR Instrument

Prepare and run the RT-PCR reaction

Import the data and review the results in COVID-19 Interpretive Software IVD Edition

The workflow begins with nucleic acid extraction from upper respiratory specimens (nasopharyngeal and anterior nasal swabs) that arrive in the testing site in transport media. Nucleic acids are isolated and purified from the specimens using the MagMAX™ Viral/Pathogen II Nucleic Acid Isolation Kit. Nucleic acid isolation is performed via an automated process using the KingFisher™ Flex Purification System. The nucleic acid is reverse transcribed into cDNA and amplified using the TaqPath™ COVID-19 Diagnostic PCR Kit and one of the following real-time PCR instruments:

Applied Biosystems™ 7500 Fast Dx Real-Time PCR Instrument ●
Applied Biosystems™ QuantStudio™ 5 Dx Real-Time PCR Instrument .

In the process, the probes anneal to three (3) specific SARS-CoV-2 target sequences located between three (3) unique forward and reverse primers for the following genes:

ORF1ab .
N gene ●
. S gene

During the extension phase of the PCR cycle, the 5' nuclease activity of Taq polymerase degrades the probe, causing the reporter dye to separate from the quencher dye, generating a fluorescent signal. With each PCR cycle, additional reporter dye molecules are cleaved from their respective

6

probes, increasing the fluorescent intensity, which is measured at each cycle by the real-time PCR instrument.

Following RT-PCR, the data from the instrument's data collection software are imported into COVID-19 Interpretive Software IVD Edition for analysis and interpretation. After data import, the software analyzes the run data, performs quality check (QC) analysis, and calculates the interpretive results for each sample and control. The imported data and interpretive results for each run are saved as a batch in the software. Results can be exported as CSV files and reports can be generated in PDF format.

Validation of the results is performed automatically by the COVID-19 Interpretive Software based on performance of the positive and negative controls. The following results are automatically generated using the calling rules, plate validity and the CT cutoff values for assay targets:

ORF1ab	N gene	S gene	MS2	Status	Result	Action
NEG	NEG	NEG	NEG	INVALID	NA	RETEST
Repeat test by re-extracting the original
sample and repeating the RT-PCR. If the
repeat result remains invalid, consider
collecting a new specimen.
NEG	NEG	NEG	POS	VALID	SARS-CoV-2
Not Detected	REPORT
Report the results to the healthcare
provider.

Table 1: Patient Samples

7

ORF1ab	N gene	S gene
	Only one SARS-CoV-2 target
= POS		POS or
NEG	VALID	SARS-CoV-2
Inconclusive	RETEST/REPORT

Repeat the test by re-extracting the
original sample and repeating the
RT-PCR.
IMPORTANT! Samples with a
result of SARS-CoV-2 Inconclusive
shall be retested one time.
After retesting one time, report the
results to the healthcare provider.
If the repeat result remains
inconclusive, the healthcare
provider should conduct additional
confirmation testing with a new
specimen, if clinically indicated. |
| | Two or more SARS-CoV-2 targets
= POS | | POS or
NEG | VALID | Positive SARS-
CoV-2 | REPORT
Report the results to the healthcare
provider. |

A minimum of one negative control and one positive control must be present for each run. Additional negative control wells shall be run for each extraction that is represented on a realtime RT-PCR plate. All control wells must pass for the real-time RT-PCR plate to be considered valid. Recommended actions of retest or report are also provided depending on the results generated.

Table 2: Materials provided in each TaqPath™ COVID-19 Diagnostic PCR Kit, 1000 reactions:

Component	Quantity	Amount
TaqPath COVID-19 Diagnostic PCR Assay
Kit	1 Tube Multiplex Assay	1,500 µL
	10 Tubes MS2 Phage	$10 \times 1,000$ µL
TaqPath COVID-19 Diagnostic PCR Control	5 Boxes, 2 Tubes per Box	$2 \times 10$ µL per box;
5 boxes per kit
TaqPath COVID-19 Diagnostic PCR Control Dilution Buffer	5 Boxes, 2 Tubes per Box	$2 \times 250$ µL per box;
5 boxes per kit

Materials required but not provided:

MagMAX™ Viral/Pathogen II Nucleic Acid Isolation Kit .
KingFisher™ Flex Purification System .

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. TaqPath™ 1-Step Multiplex Master Mix (No ROX™)
Applied Biosystems™ 7500 Fast Dx Real-Time PCR Instrument .
Applied Biosystems™ QuantStudio™ 5 Dx Real-Time PCR Instrument .
. Additional reagents and materials required are listed in the instructions for use document.

The use of the TaqPath™ COVID-19 Diagnostic PCR Kit assay as an In vitro diagnostic is limited to laboratories that are certified under the Clinical Laboratory Improvement Amendments of 1988 (CLIA), 42 U.S.C. § 263a, that meet requirements to perform high complexity tests.

Intended Use

The TaqPath™ COVID-19 Diagnostic PCR Kit is a real-time reverse transcription polymerase chain reaction (RT-PCR) test intended for the qualitative detection of nucleic acid from SARS-CoV-2 in nasopharyngeal and anterior nasal specimens from individuals with signs and symptoms of respiratory tract infection.

The TaqPath™ COVID-19 Diagnostic PCR Kit is intended for use as an aid in the diagnosis of COVID-19 if used in conjunction with other clinical observations, epidemiological information and laboratory findings. The SARS-CoV-2 RNA is generally detectable in upper respiratory (anterior nasal and nasopharyngeal swabs) specimens during the acute phase of infection. Positive results are indicative of the presence of SARS-CoV-2 RNA. Positive results do not rule out bacterial infection or co-infection with other pathogens. The agent detected may not be the definite cause of disease.

Negative results do not preclude SARS-CoV-2 infection and should not be used as the sole basis for patient management decisions.

The TaqPath™ COVID-19 Diagnostic PCR Kit is intended for use by qualified clinical laboratory personnel specifically instructed and trained in the techniques of real-time PCR and in vitro diagnostic procedures.

9

Table 3: Comparison of the TagPath™ COVID-19 Diagnostic PCR Kit and the predicate
device
Device & Predicate Device	New Device	K211079
Device Trade Name	TaqPath™ COVID-19
Diagnostic PCR Kit	BioFire COVID-19 Test 2
General Device
Characteristic Similarities
Organisms Detected	SARS-CoV-2	SARS-CoV-2
Analyte	RNA	RNA
Technological Principles	RT-PCR	RT-PCR
Test Interpretation	Automated test interpretation	Automated test interpretation
General Device
Characteristic Differences	New Device	K211079
Intended Use/ Indications For
Use	The TaqPath™ COVID-19
Diagnostic PCR Kit is a real-
time reverse transcription
polymerase chain reaction
(RT-PCR) test intended for the
qualitative detection of nucleic
acid from SARS-CoV-2 in
nasopharyngeal and anterior
nasal specimens from
individuals with signs and
symptoms of respiratory tract
infection.
The TaqPath™ COVID-19
Diagnostic PCR Kit is
intended for use as an aid in
the diagnosis of COVID-19 if
used in conjunction with other
clinical observations,
epidemiological information
and laboratory findings. The
SARS-CoV-2 RNA is
generally detectable in upper
respiratory (anterior nasal and
nasopharyngeal swabs)
specimens during the acute
phase of infection. Positive
results are indicative of the
presence of SARS-CoV-2
RNA. Positive results do not
rule out bacterial infection or
co-infection with other
pathogens.	The BioFire COVID-19 Test 2 is a
qualitative nested multiplexed RT-
PCR in vitro diagnostic. test
intended for use with the BioFire
FilmArray 2.0 and BioFire
FilmArray Torch Systems. The
BioFire COVID-19 Test 2 detects
nucleic acids from severe acute
respiratory syndrome coronavirus 2
(SARS- CoV-2) in nasopharyngeal
swabs (NPS) from individuals
suspected of COVID-19 by their
healthcare provider.
Results are for the identification of
SARS- CoV-2 RNA. The SARS-
CoV-2 RNA is generally detectable
in NPS specimens during the acute
phase of infection. Positive results
are indicative of the presence of
SARS-CoV- 2 RNA; clinical
correlation with patient history and
other diagnostic information is
necessary to determine patient
infection status. Positive results do
not rule out co-infection with other
pathogens.
Results are meant to be used in
conjunction with other clinical,
epidemiologic, and laboratory data,

	may not be the definite cause
of disease.
Negative results do not
preclude SARS-CoV-2
infection and should not be
used as the sole basis for
patient management decisions.
The TaqPath™ COVID-19
Diagnostic PCR Kit is
intended for use by qualified
clinical laboratory personnel
specifically instructed and
trained in the techniques of
real-time PCR and in vitro
diagnostic procedures.	provided by the relevant public
health authorities.
The BioFire COVID-19 Test 2 is
intended for use by trained medical
and laboratory professionals in a
laboratory setting or under the
supervision of a trained laboratory
professional.
Specimen Types	Anterior Nasal Swabs and
Nasopharyngeal swabs	Nasopharyngeal swabs only
Instrumentation	7500 Fast Dx, QuantStudio™
5 Dx	FilmArray 2.0 or FilmArray Torch
Controls	One negative control and one
positive control are run for
each assay.	Two internal controls are included in
each reagent pouch for quality
control of sample processing and
both PCR stages and melt analysis
Target Genes	ORF1ab, S gene, N Gene	ORF1ab, S gene, N Gene, ORF8

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Performance Characteristics:

a) Limit of Detection

The limit of detection (LoD) for the TaqPath™ COVID-19 Diagnostic PCR Kit was determined using inactivated SARS-CoV-2 virus (USA-WA1/2020) in pooled negative NP (Nasopharyngeal swab) specimen. Samples were randomized and blinded prior to testing by the operator. A preliminary LoD was determined by testing serial dilutions of NP spiked with inactivated virus. Samples were tested on two (2) PCR instruments: QuantStudio™ 5 Dx (QS5 Dx) and 7500 Fast Dx.

The preliminary LoD was confirmed by testing twenty (20) replicates of contrived positive samples formulated at three concentration levels based around the preliminary LoD in Phase I: 3x (150 GCE/mL), 1x (50 GCE/mL), 0.33x (17 GCE/mL). Confirmatory testing results determined the LoD for both QS5 Dx and 7500 Fast Dx to be 50 GCE/mL as the lowest concentration tested

11

for each instrument that resulted in ≥ 95% positivity (19/20 replicates). The results of Phase II testing are summarized in Table 4 below.

| Viral Target | Concentration
(GCE/mL) | QuantStudio™ 5 Dx | 7500 Fast Dx |
|------------------------------------|---------------------------|-------------------|--------------|
| SARS-CoV-2 virus
(USA-WA1/2020) | 150 | 100% (20/20) | 100% (20/20) |
| | 50 | 100% (20/20) | 100% (20/20) |
| | 17 | 85% (17/20) | 70% (14/20) |
| | 0 | 0% (0/20) | 0% (0/20) |

Table 4: LoD Phase II Summary

b) Reproducibility and Within-Laboratory Precision

The reproducibility of the TaqPath™ COVID-19 Diagnostic PCR kit was assessed separately for each PCR instrument (7500 Fast Dx and QS5 Dx). Testing was performed at three sites (two external and one internal) using samples prepared at three (3) concentrations; negative (0x LoD), a low positive (1.5x LoD), and a moderate positive (5x LoD). Positive samples were spiked with inactivated SARS-CoV-2 virus in pooled negative NP specimen matrix. All samples were blinded and randomized before testing.

Testing included three (3) sites x two (2) operators x three (3) replicates x three (3) lots of TaqPath™ COVID-19 Diagnostic PCR Kits x five (5) non-consecutive days, for a total of 270 replicates per sample at each test level (0x LoD, 1.5x LoD, 5x LoD) on each PCR instrument model (7500 Fast Dx and QS5 Dx) to account for potential sources of variation.

The TaqPath™ COVID-19 Diagnostic PCR kit met acceptance criteria and generated 100% PPA for 5x LoD, 100% PPA for 1.5x LoD and 100% NPA for 0x LoD for each lot tested across all three sites. Qualitative study results are summarized below in Table 6 below. Overall reproducibility results for each instrument are summarized in Table 8. Precision and reproducibility results for each instrument by site are summarized in Table 10. The

12

number of positive target detections (n/N), mean Ct, standard deviation (SD) and coefficient of variation (CV) are shown by instrument and site for all assay targets.

| Instrument
Type | LoD | Site | Positive SARS-CoV-2 | | PPA Across all Sites
(%, Low CI %, High CI %) |
|-----------------------------|------|------|---------------------|---------|--------------------------------------------------|
| | | | N | Percent | |
| 7500 Fast Dx
System | 1.5x | 1 | 90/90 | 100% | |
| | | 2 | 90/90 | 100% | 100% (98.6%, 100%) |
| | | 3 | 90/90 | 100% | |
| | 5x | 1 | 90/90 | 100% | |
| | | 2 | 90/90 | 100% | 100% (98.6%, 100%) |
| | | 3 | 90/90 | 100% | |
| QuantStudio™ 5 Dx
System | 1.5x | 1 | 90/90 | 100% | |
| | | 2 | 90/90 | 100% | 100% (98.6%, 100%) |
| | | 3 | 90/90 | 100% | |
| | 5x | 1 | 90/90 | 100% | |
| | | 2 | 90/90 | 100% | 100% (98.6%, 100%) |
| | | 3 | 90/90 | 100% | |

Table 5: Positive Percent Agreement (PPA) Across All Sites

Table 6: Negative Percent Agreement (NPA) Across All Sites

| Instrument
Type | LoD | Site | SARS-CoV-2 Not
Detected | | NPA Across all Sites
(%, Low CI %, High CI %) |
|------------------------------|-----|------|----------------------------|---------|--------------------------------------------------|
| | | | N | Percent | |
| 7500 Fast Dx
System | 0x | 1 | 90/90 | 100% | 100% (98.6%, 100%) |
| 7500 Fast Dx
System | 0x | 2 | 90/90 | 100% | |
| 7500 Fast Dx
System | 0x | 3 | 90/90 | 100% | |
| QuantStudioTM 5 Dx
System | 0x | 1 | 90/90 | 100% | 100% (98.6%, 100%) |
| QuantStudioTM 5 Dx
System | 0x | 2 | 90/90 | 100% | |
| QuantStudioTM 5 Dx
System | 0x | 3 | 90/90 | 100% | |

13

| Panel
member | Target | Detected
(n/N) | mean
Ct | Between-lots | | Between-
sites | | Between-
days | | Between-
operators/runs | | Repeatability
(Within-Run) | | Reproducibility | |
|-----------------|--------|-------------------|------------|--------------|------|-------------------|------|------------------|------|----------------------------|------|-------------------------------|------|-----------------|------|
| | | | | SD | %CV | SD | %CV | SD | %CV | SD | %CV | SD | %CV | SD | %CV |
| 1.5x
LoD | ORF1ab | 270/270 | 31.82 | 0.02 | 0.07 | 0.56 | 1.76 | 0.14 | 0.43 | 0.15 | 0.48 | 0.55 | 1.72 | 0.81 | 2.54 |
| 1.5x
LoD | N gene | 270/270 | 32.27 | 0.02 | 0.06 | 0.43 | 1.34 | 0.00 | 0.00 | 0.19 | 0.60 | 0.58 | 1.81 | 0.75 | 2.33 |
| | S gene | 264/270 | 32.27 | 0.07 | 0.20 | 0.05 | 0.15 | 0.00 | 0.00 | 0.26 | 0.82 | 0.80 | 2.47 | 0.84 | 2.61 |
| 1.5x
LoD | MS2 | 270/270 | 24.48 | 0.14 | 0.58 | 0.26 | 1.06 | 0.28 | 1.14 | 0.23 | 0.92 | 0.18 | 0.73 | 0.50 | 2.04 |
| 5x LoD | ORF1ab | 270/270 | 30.14 | 0.06 | 0.21 | 0.62 | 2.07 | 0.06 | 0.21 | 0.14 | 0.45 | 0.39 | 1.31 | 0.75 | 2.50 |
| 5x LoD | N gene | 270/270 | 30.57 | 0.01 | 0.04 | 0.48 | 1.58 | 0.16 | 0.53 | 0.00 | 0.00 | 0.34 | 1.12 | 0.61 | 2.01 |
| 5x LoD | S gene | 270/270 | 30.41 | 0.08 | 0.25 | 0.26 | 0.87 | 0.08 | 0.28 | 0.05 | 0.17 | 0.36 | 1.19 | 0.47 | 1.53 |
| | 5x LoD | MS2 | 270/270 | 24.46 | 0.13 | 0.52 | 0.33 | 1.33 | 0.33 | 1.33 | 0.27 | 1.09 | 0.22 | 0.92 | 0.59 |
| 0x LoD | MS2 | 270/270 | 24.50 | 0.11 | 0.46 | 0.18 | 0.75 | 0.27 | 1.09 | 0.30 | 1.21 | 0.17 | 0.69 | 0.48 | 1.97 |

Table 7: Overall Reproducibility - 7500 Fast Dx Real-Time PCR Instrument

Table 8: Overall Reproducibility - QuantStudio™ 5 Dx Real-Time PCR Instrument

| Panel
member | Target | Detected
(n/N) | mean
Ct | Between-lots | | Between-
sites | | Between-
days | | Between-
operators/runs | | Repeatability
(Within-Run) | | Reproducibility | |
|-----------------|--------|-------------------|------------|--------------|------|-------------------|------|------------------|------|----------------------------|------|-------------------------------|------|-----------------|------|
| SD | %CV | SD | %CV | SD | %CV | SD | %CV | SD | %CV | SD | %CV | | | | |
| 1.5x
LoD | ORF1ab | 270/270 | 32.56 | 0.11 | 0.35 | 0.10 | 0.29 | 0.00 | 0.00 | 0.17 | 0.53 | 0.51 | 1.56 | 0.56 | 1.71 |
| | N gene | 269/270 | 33.07 | 0.00 | 0.00 | 0.15 | 0.44 | 0.23 | 0.70 | 0.10 | 0.30 | 0.61 | 1.84 | 0.67 | 2.04 |
| | S gene | 264/270 | 33.35 | 0.09 | 0.26 | 0.10 | 0.31 | 0.00 | 0.00 | 0.37 | 1.12 | 0.77 | 2.31 | 0.87 | 2.60 |
| | MS2 | 270/270 | 25.55 | 0.05 | 0.19 | 0.12 | 0.46 | 0.30 | 1.17 | 0.40 | 1.58 | 0.17 | 0.65 | 0.54 | 2.13 |
| 5x LoD | ORF1ab | 270/270 | 30.94 | 0.11 | 0.34 | 0.00 | 0.00 | 0.00 | 0.00 | 0.13 | 0.43 | 0.30 | 0.99 | 0.35 | 1.13 |
| | N gene | 270/270 | 31.34 | 0.10 | 0.31 | 0.05 | 0.15 | 0.07 | 0.22 | 0.14 | 0.44 | 0.35 | 1.10 | 0.39 | 1.25 |
| | S gene | 270/270 | 31.35 | 0.11 | 0.34 | 0.11 | 0.35 | 0.00 | 0.00 | 0.19 | 0.59 | 0.27 | 0.87 | 0.36 | 1.16 |
| | MS2 | 270/270 | 25.54 | 0.06 | 0.22 | 0.17 | 0.67 | 0.32 | 1.25 | 0.32 | 1.24 | 0.23 | 0.90 | 0.54 | 2.10 |

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| Panel
member | Target | Detected
(n/N) | mean
Ct | Between-lots | | Between-sites | | Between-days | | Between-
operators/runs | | Repeatability
(Within-Run) | | Reproducibility | |
|-----------------|--------|-------------------|------------|--------------|------|---------------|------|--------------|------|----------------------------|------|-------------------------------|------|-----------------|------|
| | | | | SD | %CV | SD | %CV | SD | %CV | SD | %CV | SD | %CV | SD | %CV |
| 0x LoD | MS2 | 270/270 | 25.52 | 0.00 | 0.00 | 0.25 | 0.97 | 0.31 | 1.23 | 0.29 | 1.13 | 0.14 | 0.56 | 0.51 | 2.01 |

Table 9: Summary of Within-lab Precision Results by Site -7500 Fast Dx Real-Time PCR Instrument

| Site | Panel
member | Target | Detected
(n/N) | Mean
Ct | Between-
lots | | Between-
days | | Between-
operators/runs | | Repeatability
(Within-run) | | Within-lab
Precision | |
|--------|-----------------|--------|-------------------|------------|------------------|------|------------------|------|----------------------------|------|-------------------------------|------|-------------------------|------|
| | | | | | SD | %CV | SD | %CV | SD | %CV | SD | %CV | SD | %CV |
| Site 1 | 1.5x LoD | ORF1ab | 90/90 | 32.19 | 0.00 | 0.00 | 0.14 | 0.42 | 0.14 | 0.44 | 0.54 | 1.69 | 0.58 | 1.80 |
| | | N gene | 90/90 | 32.56 | 0.00 | 0.00 | 0.00 | 0.00 | 0.19 | 0.58 | 0.49 | 1.50 | 0.52 | 1.61 |
| | | S gene | 89/90 | 32.38 | 0.00 | 0.00 | 0.14 | 0.43 | 0.00 | 0.00 | 0.57 | 1.76 | 0.59 | 1.81 |
| | | MS2 | 90/90 | 24.64 | 0.42 | 1.70 | 0.22 | 0.89 | 0.12 | 0.50 | 0.18 | 0.75 | 0.52 | 2.12 |
| Site 1 | 5x LoD | ORF1ab | 90/90 | 30.39 | 0.06 | 0.18 | 0.12 | 0.41 | 0.00 | 0.00 | 0.38 | 1.24 | 0.40 | 1.32 |
| | | N gene | 90/90 | 30.90 | 0.15 | 0.48 | 0.11 | 0.35 | 0.00 | 0.00 | 0.34 | 1.11 | 0.39 | 1.26 |
| | | S gene | 90/90 | 30.65 | 0.17 | 0.55 | 0.08 | 0.27 | 0.00 | 0.00 | 0.34 | 1.10 | 0.38 | 1.26 |
| | | MS2 | 90/90 | 24.55 | 0.48 | 1.94 | 0.08 | 0.32 | 0.18 | 0.74 | 0.20 | 0.83 | 0.55 | 2.26 |
| Site 1 | 0x LoD | MS2 | 90/90 | 24.48 | 0.51 | 2.08 | 0.11 | 0.46 | 0.19 | 0.78 | 0.21 | 0.87 | 0.60 | 2.43 |
| Site 2 | 1.5x LoD | ORF1ab | 90/90 | 31.17 | 0.00 | 0.00 | 0.19 | 0.60 | 0.06 | 0.18 | 0.53 | 1.71 | 0.57 | 1.82 |
| | | N gene | 90/90 | 31.76 | 0.00 | 0.00 | 0.00 | 0.00 | 0.22 | 0.69 | 0.58 | 1.83 | 0.62 | 1.95 |
| | | S gene | 86/90 | 32.18 | 0.00 | 0.00 | 0.00 | 0.00 | 0.00 | 0.00 | 0.74 | 2.30 | 0.74 | 2.30 |
| | | MS2 | 90/90 | 24.17 | 0.10 | 0.40 | 0.26 | 1.06 | 0.22 | 0.93 | 0.14 | 0.59 | 0.38 | 1.58 |
| Site 2 | 5x LoD | ORF1ab | 90/90 | 29.42 | 0.05 | 0.16 | 0.12 | 0.41 | 0.18 | 0.60 | 0.44 | 1.48 | 0.49 | 1.66 |
| | | N gene | 90/90 | 30.02 | 0.08 | 0.26 | 0.11 | 0.36 | 0.00 | 0.00 | 0.36 | 1.21 | 0.39 | 1.29 |
| | | S gene | 90/90 | 30.12 | 0.04 | 0.12 | 0.10 | 0.34 | 0.00 | 0.00 | 0.36 | 1.21 | 0.38 | 1.26 |
| | | MS2 | 90/90 | 24.09 | 0.15 | 0.61 | 0.17 | 0.69 | 0.27 | 1.12 | 0.27 | 1.12 | 0.44 | 1.83 |
| Site 2 | 0x LoD | MS2 | 90/90 | 24.30 | 0.23 | 0.94 | 0.00 | 0.00 | 0.32 | 1.34 | 0.13 | 0.52 | 0.42 | 1.71 |
| Site 3 | 1.5x LoD | ORF1ab | 90/90 | 32.11 | 0.00 | 0.00 | 0.12 | 0.39 | 0.22 | 0.67 | 0.56 | 1.75 | 0.62 | 1.92 |
| | | N gene | 90/90 | 32.47 | 0.18 | 0.57 | 0.00 | 0.00 | 0.13 | 0.39 | 0.67 | 2.07 | 0.71 | 2.18 |
| | | S gene | 89/90 | 32.27 | 0.00 | 0.00 | 0.00 | 0.00 | 0.54 | 1.66 | 1.01 | 3.13 | 1.14 | 3.55 |
| | | MS2 | 90/90 | 24.64 | 0.17 | 0.71 | 0.12 | 0.47 | 0.30 | 1.20 | 0.20 | 0.83 | 0.42 | 1.69 |

15

| Site | Panel
member | Target | Detected
(n/N) | Mean
Ct | Between-
lots | | Between-
days | | Between-
operators/runs | | Repeatability
(Within-run) | | Within-lab
Precision | |
|------|-----------------|--------|-------------------|------------|------------------|------|------------------|------|----------------------------|------|-------------------------------|------|-------------------------|------|
| | | | | | SD | %CV | SD | %CV | SD | %CV | SD | %CV | SD | %CV |
| | 5x LoD | ORF1ab | 90/90 | 30.60 | 0.11 | 0.38 | 0.00 | 0.00 | 0.23 | 0.74 | 0.36 | 1.19 | 0.44 | 1.45 |
| | 5x LoD | N gene | 90/90 | 30.80 | 0.14 | 0.46 | 0.15 | 0.49 | 0.00 | 0.00 | 0.32 | 1.03 | 0.38 | 1.23 |
| | 5x LoD | S gene | 90/90 | 30.44 | 0.10 | 0.32 | 0.00 | 0.00 | 0.15 | 0.49 | 0.38 | 1.26 | 0.42 | 1.39 |
| | 5x LoD | MS2 | 90/90 | 24.75 | 0.23 | 0.91 | 0.32 | 1.30 | 0.33 | 1.33 | 0.19 | 0.77 | 0.55 | 2.21 |
| | 0x LoD | MS2 | 90/90 | 24.71 | 0.10 | 0.42 | 0.15 | 0.61 | 0.35 | 1.42 | 0.16 | 0.64 | 0.43 | 1.72 |

Table 10: Summary of Within-lab Precision Results by Site - QuantStudio™ 5 Dx Real-Time PCR Instrument

| Site | Panel
member | Target | Detected
(n/N) | Mean
Ct | Between-lots | | Between-days | | Between-
operators/runs | | Repeatability
(Within-run) | | Within-lab
Precision | |
|--------|-----------------|--------|-------------------|------------|--------------|------|--------------|------|----------------------------|------|-------------------------------|------|-------------------------|------|
| | | | | | SD | %CV | SD | %CV | SD | %CV | SD | %CV | SD | %CV |
| Site 1 | 1.5x LoD | ORF1ab | 90/90 | 32.50 | 0.14 | 0.42 | 0.00 | 0.00 | 0.20 | 0.62 | 0.51 | 1.58 | 0.57 | 1.75 |
| | | N gene | 90/90 | 33.11 | 0.00 | 0.00 | 0.27 | 0.82 | 0.00 | 0.00 | 0.56 | 1.69 | 0.62 | 1.88 |
| | | S gene | 90/90 | 33.20 | 0.11 | 0.33 | 0.23 | 0.71 | 0.00 | 0.00 | 0.53 | 1.59 | 0.59 | 1.77 |
| | | MS2 | 90/90 | 25.36 | 0.41 | 1.62 | 0.19 | 0.75 | 0.18 | 0.71 | 0.14 | 0.55 | 0.51 | 2.00 |
| Site 1 | 5x LoD | ORF1ab | 90/90 | 30.93 | 0.10 | 0.33 | 0.00 | 0.00 | 0.07 | 0.22 | 0.30 | 0.97 | 0.33 | 1.05 |
| | | N gene | 90/90 | 31.41 | 0.12 | 0.38 | 0.11 | 0.34 | 0.00 | 0.00 | 0.35 | 1.10 | 0.38 | 1.21 |
| | | S gene | 90/90 | 31.47 | 0.15 | 0.47 | 0.00 | 0.00 | 0.09 | 0.29 | 0.27 | 0.87 | 0.32 | 1.03 |
| | | MS2 | 90/90 | 25.31 | 0.45 | 1.77 | 0.09 | 0.37 | 0.16 | 0.63 | 0.14 | 0.56 | 0.51 | 2.00 |
| Site 1 | 0x LoD | MS2 | 90/90 | 25.21 | 0.46 | 1.81 | 0.10 | 0.39 | 0.22 | 0.89 | 0.12 | 0.47 | 0.53 | 2.11 |
| Site 2 | 1.5x LoD | ORF1ab | 90/90 | 32.69 | 0.06 | 0.17 | 0.00 | 0.00 | 0.32 | 0.98 | 0.42 | 1.27 | 0.53 | 1.61 |
| | | N gene | 89/90 | 33.22 | 0.00 | 0.00 | 0.22 | 0.67 | 0.06 | 0.19 | 0.61 | 1.84 | 0.65 | 1.97 |
| | | S gene | 90/90 | 33.43 | 0.26 | 0.77 | 0.00 | 0.00 | 0.34 | 1.02 | 0.85 | 2.55 | 0.95 | 2.85 |
| | | MS2 | 90/90 | 25.63 | 0.06 | 0.22 | 0.18 | 0.70 | 0.36 | 1.40 | 0.12 | 0.46 | 0.42 | 1.65 |
| Site 2 | 5x LoD | ORF1ab | 90/90 | 30.98 | 0.09 | 0.28 | 0.00 | 0.00 | 0.00 | 0.00 | 0.31 | 1.01 | 0.32 | 1.05 |
| | | N gene | 90/90 | 31.33 | 0.07 | 0.23 | 0.00 | 0.00 | 0.00 | 0.00 | 0.36 | 1.14 | 0.37 | 1.17 |
| | | S gene | 90/90 | 31.24 | 0.10 | 0.31 | 0.00 | 0.00 | 0.17 | 0.56 | 0.27 | 0.86 | 0.34 | 1.07 |
| | | MS2 | 90/90 | 25.58 | 0.13 | 0.52 | 0.05 | 0.21 | 0.27 | 1.05 | 0.28 | 1.08 | 0.41 | 1.61 |

16

| Site | Panel
member | Target | Detected
(n/N) | Mean
Ct | Between-lots | | Between-days | | Between-
operators/runs | | Repeatability
(Within-run) | | Within-lab
Precision | |
|--------|-----------------|--------|-------------------|------------|--------------|------|--------------|------|----------------------------|------|-------------------------------|------|-------------------------|------|
| | | | | | SD | %CV | SD | %CV | SD | %CV | SD | %CV | SD | %CV |
| | 0x LoD | MS2 | 90/90 | 25.70 | 0.17 | 0.68 | 0.06 | 0.25 | 0.27 | 1.06 | 0.13 | 0.51 | 0.35 | 1.38 |
| Site 3 | 1.5x LoD | ORF1ab | 90/90 | 32.48 | 0.16 | 0.49 | 0.21 | 0.65 | 0.00 | 0.00 | 0.58 | 1.78 | 0.64 | 1.96 |
| | | N gene | 90/90 | 32.89 | 0.12 | 0.36 | 0.24 | 0.72 | 0.27 | 0.83 | 0.65 | 1.97 | 0.75 | 2.28 |
| | | S gene | 84/90 | 33.44 | 0.00 | 0.00 | 0.00 | 0.00 | 0.61 | 1.82 | 0.89 | 2.65 | 1.08 | 3.22 |
| | | MS2 | 90/90 | 25.65 | 0.25 | 0.98 | 0.15 | 0.58 | 0.58 | 2.27 | 0.22 | 0.86 | 0.69 | 2.68 |
| | 5x LoD | ORF1ab | 90/90 | 30.91 | 0.16 | 0.52 | 0.03 | 0.09 | 0.23 | 0.74 | 0.30 | 0.97 | 0.41 | 1.33 |
| | | N gene | 90/90 | 31.28 | 0.21 | 0.68 | 0.00 | 0.00 | 0.25 | 0.79 | 0.33 | 1.06 | 0.47 | 1.49 |
| | | S gene | 90/90 | 31.34 | 0.07 | 0.22 | 0.00 | 0.00 | 0.26 | 0.82 | 0.28 | 0.88 | 0.38 | 1.22 |
| | | MS2 | 90/90 | 25.72 | 0.35 | 1.37 | 0.25 | 0.96 | 0.45 | 1.74 | 0.25 | 0.96 | 0.67 | 2.60 |
| | 0x LoD | MS2 | 90/90 | 25.66 | 0.29 | 1.11 | 0.22 | 0.86 | 0.36 | 1.39 | 0.17 | 0.67 | 0.54 | 2.09 |

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c) In Use Stability

The In-use stability of the TaqPath™ COVID-19 Diagnostic PCR Kit reagents under expected use conditions was evaluated.

In-use stability of the TaqPath™ COVID-19 Diagnostic PCR Control was evaluated at undiluted (1 x 104) and working concentrations (25 copies/ µL) under refrigeration at 25 and 50 hours. Additionally, the working stability of the TaqPath™ COVID-19 Diagnostic PCR Assay Multiplex reagents under freeze thaw conditions was also evaluated. Freeze thaw stability of the TagPath™ COVID-19 Diagnostic PCR Control, TaqPath™ COVID-19 Diagnostic PCR Control Dilution Buffer, and MS2 Phage Control were not evaluated as multiple tubes are provided in with the TaqPath™ COVID-19 Diagnostic PCR Kit and freeze-thaw cycles are not recommended. Likewise, the refrigerated stability of the TaqPath™ COVID-19 Diagnostic PCR Assay Multiplex was not evaluated as storage under refrigeration is not recommended.

The study data supports the in-use stability under the conditions listed in Table 11.

| Component Tested | Storage Holding
Condition | In-Use Stability claim |
|------------------------------------------------------------------------------------------|------------------------------|---------------------------------------|
| TaqPath™ COVID-19
Diagnostic PCR Assay
Multiplex (Freeze-Thaw) | -30°C to -10°C | Stable up to 10 Freeze-thaw
cycles |
| TaqPath™ COVID-19
Diagnostic PCR Control
Stock – Not Diluted
(Refrigerated) | 2°C to 8°C | Stable up to 48 hours |
| TaqPath™ COVID-19
Diagnostic PCR Control
Working Stock - Diluted
(Refrigerated) | 2°C to 8°C | Stable up to 48 hours |

Table 11: TaqPath™ COVID-19 Diagnostic PCR Kit In-use Stability

d) Transport Simulation

The purpose of the transport simulation study was to demonstrate that the shipping configurations developed for the TaqPath™ COVID-19 Diagnostic PCR Kit reagents provide adequate thermal

18

and physical protection as packages are transported to customer sites. Three (3) lots of TaqPath™ COVID-19 Diagnostic PCR Kit, components of the kit were exposed to thermal challenges and vibration in their final shipping configuration followed by functional testing. Simulated transport challenges were performed according to the American Society for Testing and Materials (ASTM) D4169, International Safe Transit Association (ISTA) 7E requirements by a third-party testing organization followed by functional testing at Thermo Fisher Scientific.

This study demonstrated that primary and distribution packaging integrity including that of labels has been maintained, temperature ranges for materials distributed are maintained within allowable limits and all reagents perform as expected according to requirements and specifications upon exposure to the simulated distribution environment conditions.

e) Interfering Substances

The potential impact of endogenous and exogenous interferents found in respiratory specimens on the performance of the TaqPath™ COVID-19 Diagnostic PCR Kit were evaluated. A total of 27 interfering substances were spiked in contrived negative matrix prepared using negative pooled nasopharyngeal swab samples (0x LoD) and contrived positive samples using inactivated SARS-CoV-2 virus at a concentration of 3x LoD, to evaluate potential false positive interference and false negative interference respectively.

All interfering substances are considered non-interfering at the tested concentrations as shown below.

Results are summarized in Table 12 below.

Substance	Concentration	PPA	NPA
Leukocytes (human)	1% (v/v)	100% (3/3)	100% (3/3)
Mucin: bovine submaxillary gland, type
I-S	0.1 mg/mL	100% (3/3)	100% (3/3)
Blood (human)	1% (v/v)	100% (3/3)	100% (3/3)
Substance	Concentration	PPA	NPA
Throat lozenges: Benzocaine (7.5 mg),
Dextromethorphan HBr (5 mg)	1% (w/v)	100% (3/3)	100% (3/3)
Throat lozenges: Menthol (5.4 mg)	2.2 mg/mL	100% (3/3)	100% (3/3)
Nasal sprays or drops: Phenylephrine	10% (v/v)	100% (3/3)	100% (3/3)
Nasal sprays or drops: Oxymetazoline
(Afrin - Allergy Sinus)	10% (v/v)	100% (3/3)	100% (3/3)
Nasal sprays or drops: Oxymetazoline	10% (v/v)	100% (6/6)	83% (5/6)
(Afrin - No drip, extra moisturizing)[1]	5% (v/v)	100% (3/3)	100% (3/3)
Nasal sprays or drops: Sodium chloride
with preservatives	10% (v/v)	100% (3/3)	100% (3/3)
Bronchodilator: Albuterol	0.83 mg/mL	100% (3/3)	100% (3/3)
Nasal corticosteroids: Beclomethasone	2 mg/mL	100% (3/3)	100% (3/3)
Nasal corticosteroids: Dexamethasone	1.5 mg/mL	100% (3/3)	100% (3/3)
Nasal corticosteroids: Flunisolide	2 mg/mL	100% (3/3)	100% (3/3)
Nasal corticosteroids: Budesonide	1% (v/v)	100% (3/3)	100% (3/3)
Nasal corticosteroids: Mometasone	1 mg/mL	100% (3/3)	100% (3/3)
Nasal corticosteroid: Triamcinolone[1]	10% (v/v)	83% (5/6)	17% (1/6)
	5% (v/v)	100% (3/3)	100% (3/3)
Nasal corticosteroid: Fluticasone	5 ug/mL	100% (3/3)	100% (3/3)
Nasal gel: Luffa opperculata, sulfur	1% (v/v)	100% (3/3)	100% (3/3)
Zinc Acetate	7.5 mg/mL	100% (3/3)	100% (3/3)
Anti-viral drug: Remdesivir	6.7 µg/mL	100% (3/3)	100% (3/3)
Anti-viral drug: Zanamivir	5.5 mg/mL	100% (3/3)	100% (3/3)
Anti-viral drug: Oseltamivir	33 ug/mL	100% (3/3)	100% (3/3)
Antibiotic, nasal ointment: Mupirocin	5 mg/mL	100% (3/3)	100% (3/3)
Antibacterial, systemic: Tobramycin	4 µg/mL	100% (3/3)	100% (3/3)
Homeopathic allergy medicine:
Galphimia glauca, Histaminum
hydrochloricum	10% (w/v)	100% (3/3)	100% (3/3)
Tobacco product	1% w/v	100% (3/3)	100% (3/3)
Analgesic (e.g., ibuprofen)	21.9 mg/dL	100% (3/3)	100% (3/3)

Table 12: Interfering substances

19

1 Interference was observed at concentrations >5% (v/v).

20

f) Cross-Reactivity and Microbial Interference

These studies evaluated the potential for cross-reactivity and microbial interference for the TaqPath™ COVID-19 Diagnostic PCR Kit assay with microorganisms found in respiratory samples.

In the cross-reactivity study, a total of 46 potentially cross-reactive organisms were spiked into pooled negative NP specimen matrix at a high, clinically relevant concentration (106 CFU/mL or higher for bacteria/fungi and $10^5$ PFU/mL or TCID50/mL or higher for viruses). In the microbial interference study, the same 46 organisms were spiked into contrived positive NP specimen matrix formulated at a concentration of 3x LoD with inactivated SARS-CoV-2 virus. Additionally, testing was also performed with nasal wash solution contrived with 3x LoD (150 GCE/mL) with SARS-CoV-2 virus. Organisms were obtained from vendors at the highest commercially available concentration. Organisms that did not meet the target testing concentration or did not come in equivalent quantitative units (i.e., copies/mL or Ct values) were tested at the highest concentration possible. Three (3) replicates of all microorganisms were tested using one lot of TagPath™ COVID-19 Diagnostic PCR Kit on one (1) QuantStudio™ 5 Dx Real-Time PCR instrument for each segment of the study. The microorganisms and concentrations tested and a summary of the cross-reactivity and microbial interference results is provided in Table 13 below. For all organisms tested, no cross-reactivity or microbial interference was observed.

Type	Organism	Test concentration	Units
Virus	Human coronavirus 229E[1]	$2.82\times10^4$	TCID50/mL
Virus	Human coronavirus OC43	$1.00\times10^5$	TCID50/mL
Virus	Human coronavirus NL63[1]	$2.34\times10^4$	TCID50/mL
Virus	MERS-CoV	29.4	Ct
Virus	Coronavirus-SARS	28.8	Ct
Virus	Adenovirus Type 1	$1.00\times10^5$	TCID50/mL
Virus	Mastadenovirus B Type 7	$1.00\times10^5$	TCID50/mL
Virus	Human Metapneumovirus (hMPV)	$1.00\times10^5$	TCID50/mL
Type	Organism	Test concentration	Units
Virus	Parainfluenza virus 1	$1.00×10^5$	TCID50/mL
Virus	Parainfluenza virus 2	$1.00×10^5$	TCID50/mL
Virus	Parainfluenza virus 3	$1.00×10^5$	TCID50/mL
Virus	Parainfluenza virus 4	$1.00×10^5$	TCID50/mL
Virus	Influenza A[1]	$8.34×10^4$	TCID50/mL
Virus	Influenza B[1]	$8.34×10^4$	TCID50/mL
Virus	Respiratory syncytial virus A	$1.00×10^5$	TCID50/mL
Virus	Respiratory syncytial virus B	$1.00×10^5$	TCID50/mL
Virus	Rhinovirus[1]	$2.82×10^4$	TCID50/mL
Virus	Measles	$1.00×10^5$	TCID50/mL
Virus	Mumps	$1.00×10^5$	TCID50/mL
Virus	Cytomegalovirus[1]	$3.20×10^4$	TCID50/mL
Virus	Epstein-Barr virus	$1.57×10^7$ (cross-reactivity)
$5.88×10^6$ (microbial interference)	copies/mL
Virus	Human Immunodeficiency Virus type 1[1]	$1.00×10^4$	IU/mL
Virus	Human coronovirus (HKU1)[1] [2]	$9.99 × 10^4$ copies/mL	copies/mL
Virus	Enterovirus (EV-D68)[1] [2]	$9.88 × 10^4$ copies/mL	copies/mL
Bacteria	Chlamydia pneumoniae	$1.00×10^6$	IFU/mL
Bacteria	Haemophilus influenzae	$1.00×10^6$	CFU/mL
Bacteria	Legionella pneumophila	$1.00×10^6$	CFU/mL
Bacteria	Mycobacterium tuberculosis	$5.40×10^7$ (cross-reactivity)
$8.60×10^7$ (microbial interference)	GCE/mL
Bacteria	Streptococcus pneumoniae	$1.00×10^6$	CFU/mL
Bacteria	Fusobacterium necrophorum	$1.00×10^6$	CFU/mL
Bacteria	Pseudomonas aeruginosa	$1.00×10^6$	CFU/mL
Bacteria	Staphylococcus epidermis, MRSE	$1.00×10^6$	CFU/mL
Bacteria	Streptococcus salivarius	$1.00×10^6$	CFU/mL
Bacteria	Corynebacterium diphtheriae	$1.00×10^6$	CFU/mL
Bacteria	Escherichia coli	$1.00×10^6$	CFU/mL
Type	Organism	Test concentration	Units
Bacteria	Lactobacillus acidophilus	1.00×106	CFU/mL
Bacteria	Moraxella catarrhalis	1.00×106	CFU/mL
Bacteria	Neisseria meningitidis	1.00×106	CFU/mL
Bacteria	Neisseria elongata	1.00×106	CFU/mL
Bacteria	Staphylococcus aureus, MRSA	1.00×106	CFU/mL
Bacteria	Streptococcus pyogenes	1.00×106	CFU/mL
Bacteria	Bordetella pertussis	1.00×106	CFU/mL
Bacteria	Bordetella parapertussis	1.00×106	CFU/mL
Bacteria	Mycoplasma pneumoniae	1.00×106	CCU/mL
Bacteria	Mycoplasma genitalium[1]	4.00×105	Bacteria/mL
Fungi	Candida albicans	1.00×106	CFU/mL
Fungi	Aspergillus flavus	1.00×106	CFU/mL
Fungi	Pneumocystis carinii	1.00×106	Cells/mL
Biofluid	Pooled human nasal wash[3]	Neat	N/A

Table 13: Microorganisms and Concentrations Tested

21

22

l ] ] A lower concentration was tested due to inability to obtain stock material with high titer.

[2] Microbial interference was not evaluated

[3] Used to represent diverse microbial flora in the human respiratory tract.

g) In silico Cross-Reactivity

In silico analysis was performed to demonstrate that the TaqPath™ COVID-19 Diagnostic PCR kit probes and primers do not cross-react with common microorganisms found in respiratory clinical specimen.

Genome sequences of respiratory pathogens including 31,011 virus isolates, 9,996 bacterial isolates and 14 fungal isolates from GenBank were analyzed using BLAST to compare against the TaqPath™ COVID-19 Diagnostic PCR kit assay forward primers, reverse primers, and probes. Complete genomes for all isolates were available and downloaded except for Candida albicans and Pneumocystis jirovecii which did not possess any complete genomes on NCBI (National Center for Biotechnology Information). Instead, the RefSeq reference sequence was tested. Rhinovirus Type 1A did not have any complete genomes or a RefSeq, so all 18 available partial

23

genomes downloaded from NCBI were tested. Atypical genomes were excluded. Atypical genomes are genomes with one or more problems that have been identified by NCBI relating to quality, unusual size, or other flaws in the genome assembly.

A total of 101 sequences aligned with exactly one individual component (primer or probe) of the TaqPath™ COVID-19 Diagnostic PCR Kit (COVID-19 Kit) with greater than or equal to 80% homology. None of the isolates matched to two or more individual components of any TaqPath™ COVID-19 Diagnostic PCR Kit assay. An amplicon requires all three individual assay components to be generated and detected by qPCR, therefore, the isolates evaluated are predicated to be negative for cross-reactivity.

h) Carry-Over Cross-Contamination

The carry-over cross-contamination rate of the TaqPath™ COVID-19 Diagnostic PCR Kit was evaluated in a study using contrived SARS-CoV-2 samples with high viral titers (formulated at approximately $1\times10^5$ PFU/mL in negative NP matrix) plated in an alternating checkerboard pattern with negative samples. This study tested a total of ten (10) extraction runs across two (2) KingFisher 130 Flex Purification Instruments, two (2) operators, and one (1) of each real time PCR instruments - 7500 Fast Dx Real-Time PCR Instrument and QuantStudio™ 5 Dx Real-Time PCR Instrument. Each extraction run tested forty-seven (47) replicates of contrived positive samples and forty-seven (47) replicates of negative NP matrix samples. Out of a total of four hundred and seventy (470) negative samples tested, a total of four (4) false positive SARS-CoV-2 calls (2 false positive calls on 7500 Fast Dx Real-Time PCR Instrument and 2 false positive calls on QuantStudio™ 5 Dx Real-Time PCR Instrument) were observed. The carry-over cross-contamination rate of the TaqPath™ COVID-19 Diagnostic PCR Kit was determined to be 0.85% (4/470).

i) Analytical Reactivity (Inclusivity)

The inclusivity of the TaqPath™ Diagnostic PCR Kit for detection of SARS-CoV-2 was confirmed by testing fifteen variants of SARS-CoV-2. Positive samples were contrived by spiking inactivated SARS-CoV-2 virus into negative pooled NP specimen at a concentration of 3x LoD. Each SARS-

24

CoV-2 strain below was tested in triplicate along with negative a sample (0x LoD) in a blinded and randomized fashion. All fifteen (15) strains of SARS-CoV-2 tested met the acceptance criteria and generated 100% analytical inclusivity. This study demonstrated that the TaqPath™ COVID-19 Diagnostic PCR Kit has an analytical inclusivity of 100% for the common SARS-CoV-2 variants tested. Results are summarized in Table 14.

| Variant | Isolate ID | Concentration
(GCE/mL) | %PPA |
|-----------------------|-------------------------------------------------|---------------------------|------------|
| N/A | USA-WA1/2020 | 150 | 100% (3/3) |
| N/A | Italy-INMI1 | 150 | 100% (3/3) |
| N/A | Hong Kong/VM20001061/2020 | 150 | 100% (3/3) |
| Alpha; B.1.1.7 | England/204820464/2020 | 150 | 100% (3/3) |
| Beta; B.1.351 | South Africa/KRISP-K005325/2020 | 150 | 100% (3/3) |
| Gamma; P.1 | Japan/TY7-503/2021 | 150 | 100% (3/3) |
| Delta;
B.1.617.2 | USA/PHC658/2021 | 150 | 100% (3/3) |
| Lambda; C.37 | Peru/UN-CDC-2-4069945/2021 | 150 | 100% (3/3) |
| Kappa;
B.1.617.1 | USA/CA-Stanford-15 S02/2021 | 150 | 100% (3/3) |
| Omicron;
B.1.1.529 | USA/MD-HP20874/2021 | 150 | 100% (3/3) |
| Iota; B.1.526 | USA/NY-WADSWORTH-21025952-
01/2021 Isolate 1 | 150 | 100% (3/3) |
| B.1 | USA/NY-Wadsworth-103677-01/2020 | 150 | 100% (3/3) |
| B.1.595 | USA/NY-Wadsworth-33126-01/2020 | 150 | 100% (3/3) |
| Zeta; P2 | USA/NY-Wadsworth-21006055-
01/2021 | 150 | 100% (3/3) |
| Omicron; BA
2.3 | USA/MD-HP24556/2022 | 150 | 100% (3/3) |

Table 14: Inclusivity Study Results

j) In silico Reactivity-Inclusivity

In silico analysis was performed to determine inclusivity (reactivity) of the TaqPath™ COVID-19 Diagnostic PCR Kit primer/probe sequences with all known strains/isolates of SARS-CoV-2 from GISAID and GenBank databases through April 2024. Sequences from major SARS-CoV-2 variants were evaluated and found to be reactive with the assay include but are not limited to Alpha, Beta, Delta, Epsilon, Eta, Gamma, Iota, Kappa, Lambda, Mu, Omicron, Theta, and

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Zeta. Specific Omicron pango lineages evaluated and found to be reactive with the assay include but are not limited to BA.2.86, XBB.1.9.1, XBB.1.9.2, XBB.1.16, XBB.1.5, CH.1.1, JN.1, and KP.2. Mismatch and melting temperature analyses were performed and genomes with 100% identity and/or melting temperature (Tm) greater than the annealing temperature were considered reactive. Analysis indicates that for >99% of the sequences are reactive based on 100% homology or Tm greater than annealing temperature to at least two of the three assay gene targets. Evaluation of assay components that do not match 100% to target sequences indicates that most primer or probe mismatches are unlikely to affect assay function, thus the TaqPath™ COVID-19 Diagnostic PCR Kit is predicted to be highly inclusive for SARS-CoV-2.

k) Specimen Storage Stability

Specimen storage stability testing was performed to characterize the stability of SARS-COV-2 in nasopharyngeal swabs (NP) after storage at ambient temperature (15-30°C), refrigerated (2°C to 8°C), frozen (-30°C to -10°C), and frozen at ≤-70°C. The stability of SARS-CoV-2 in nasopharyngeal (NP) swabs in viral transport media (VTM) was evaluated at several time points (T₀ – Tn) with contrived positive samples and negative samples (0x LoD) for each storage condition and tested with the TaqPath™ COVID-19 Diagnostic PCR Kit.

The study produced 100% PPA and 100% NPA for all timepoints tested for all storage conditions and established the specimen storage stability: up to 4 hours at ambient temperature (15°C to 30°C), up to 3 days (72 hours) refrigerated (2°C to 8°C), up to 30 days frozen (-30°C to -10°C) and up to 30 days frozen at ≤-70°C.

l) Fresh vs Frozen Equivalency Study

A fresh vs frozen study was performed to demonstrate equivalency between fresh and frozen SARS-CoV-2 nasopharyngeal (NP) samples in viral transport media (VTM). Contrived samples were prepared by spiking inactivated SARS-CoV-2 virus in negative pooled NP samples at a concentration of 0x LoD, 1.5x LoD and 5x LoD. Samples were tested at baseline and subsequently after 1x and 2x freeze-thaw cycles when stored at -10°C to -30°C, and after 1x, 2x, 3x and 4x freeze-thaw cycles when stored at ≤ -70°C. Ten replicates were performed for 0x and 5x LoD and 40 replicates were tested for 1.5x LoD. All test conditions produced expected results with 100% NPA at 0x LoD and 100% PPA at 1.5x LoD and 5x LoD. The results demonstrated that freeze

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thaw up to 2 cycles when stored at -10°C to -30°C and freeze-thaw up to 4 cycles when stored at