Applied Biosystems™ TaqPath™ COVID-19, Flu A, Flu B, RSV Select Panel
Device Facts
| Record ID | K241806 |
|---|---|
| Device Name | Applied Biosystems™ TaqPath™ COVID-19, Flu A, Flu B, RSV Select Panel |
| Applicant | Life Technologies Corporation |
| Product Code | QOF · Microbiology |
| Decision Date | Jan 8, 2025 |
| Decision | SESE |
| Submission Type | Traditional |
| Regulation | 21 CFR 866.3981 |
| Device Class | Class 2 |
Indications for Use
The Applied Biosystems™ TaqPath™ COVID-19, Flu A, Flu B, RSV Select Panel is a multiplex, real-time reverse transcription polymerase chain reaction (RT-PCR) in vitro diagnostic test for the qualitative detection and differentiation of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), influenza A virus, influenza B virus, and respiratory syncytial virus (RSV). Nucleic acids are isolated and purified from nasopharyngeal (NP) swab and anterior nasal (AN) swab 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 test is intended to aid in the differential diagnosis of SARS-CoV-2, influenza A, influenza B, and RSV A/B (undifferentiated) 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 and AN swab specimens during the acute phase of infection 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 Applied Biosystems™ TaqPath™ COVID-19, Flu A, Flu B, RSV Select Panel 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. The Applied Biosystems™ TaqPath™ COVID-19, Flu A, Flu B, RSV Select Panel is intended for use by qualified and trained clinical laboratory personnel specifically instructed and trained in the techniques of real-time PCR and in vitro diagnostic procedures.
Device Story
Multiplex real-time RT-PCR assay; detects SARS-CoV-2, influenza A, influenza B, and RSV RNA in nasopharyngeal/anterior nasal swabs. Workflow: nucleic acid extraction via MagMAX Dx Viral/Pathogen Kit on KingFisher Apex Dx system; amplification/detection on QuantStudio 5 Dx instrument. Diomni software performs automated results interpretation based on Cq thresholds. Used in clinical laboratories by trained personnel. Output aids differential diagnosis of respiratory infections; results used alongside clinical/epidemiological data. Benefits patient by enabling rapid, accurate identification of viral pathogens to guide management.
Clinical Evidence
Prospective clinical study (N=1,840) compared subject device to FDA-cleared molecular comparator. Evaluated 1,620 NP and 1,541 AN swabs. PPA for SARS-CoV-2 was 96.6% (NP) and 95.3% (AN); NPA was 96.47% (NP) and 96.46% (AN). Flu A/B and RSV performance showed high agreement. Enrichment study (N=69) supplemented data for Flu A/B and RSV. Bench testing confirmed LoD, inclusivity, interference, and stability.
Technological Characteristics
Multiplex real-time RT-PCR; targets SARS-CoV-2 (Orf1a, Orf1b, N), Flu A (PB1, M), Flu B (M, NS), RSV (NP, M, L), and RNase P (internal control). Uses magnetic-bead extraction (MagMAX Dx) and thermal cycling (QuantStudio 5 Dx). Software-based automated interpretation. Prescription use only.
Indications for Use
Indicated for qualitative detection and differentiation of SARS-CoV-2, influenza A, influenza B, and RSV in nasopharyngeal and anterior nasal swab specimens from symptomatic individuals. Intended to aid in differential diagnosis of respiratory infections. Not intended to detect influenza C.
Regulatory Classification
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.
Special Controls
*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.
Predicate Devices
- BioFire® Respiratory Panel 2.1 (DEN200031)
Submission Summary (Full Text)
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**FDA** U.S. FOOD & DRUG
ADMINISTRATION
# **510(k) SUBSTANTIAL EQUIVALENCE DETERMINATION
DECISION SUMMARY
ASSAY AND INSTRUMENT**
# **I Background Information:**
# **A 510(k) Number**
K241806
# **B Applicant**
Life Technologies Corporation
# **C Proprietary and Established Names**
Applied Biosystems TaqPath COVID-19, Flu A, Flu B, RSV Select Panel
# **D Regulatory Information**
| Product Code(s) | Classification | Regulation Section | Panel |
| --- | --- | --- | --- |
| QOF | Class II | 21 CFR 866.3981 - Device To Detect And Identify Nucleic Acid Targets In Respiratory Specimens From Microbial Agents That Cause The Sars-Cov-2 Respiratory Infection And Other Microbial Agents When In A Multi-Target Test | MI - Microbiology |
# **II Submission/Device Overview:**
# **A Purpose for Submission:**
The purpose of this submission is to obtain market clearance for the Applied Biosystems TaqPath COVID-19, Flu A, Flu B, RSV Select Panel (referred to as TaqPath COVID-19, Flu A, Flu B, RSV Select Panel below).
# **B Measurand:**
The TaqPath COVID-19, Flu A, Flu B, RSV Select Panel detects and identifies RNA from: SARS-CoV-2, influenza A virus, influenza B virus, and respiratory syncytial virus (RSV).
Food and Drug Administration
10903 New Hampshire Avenue
Silver Spring, MD 20993-0002
www.fda.gov
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# **C Type of Test:**
The TaqPath COVID-19, Flu A, Flu B, RSV Select Panel is a multiplex, real-time reverse transcription polymerase chain reaction (RT-PCR) test.
# **III Intended Use/Indications for Use:**
# **A Intended Use(s):**
See Indications for Use below.
# **B Indication(s) for Use:**
The Applied Biosystems TaqPath COVID-19, Flu A, Flu B, RSV Select Panel is a multiplex, real-time reverse transcription polymerase chain reaction (RT-PCR) *in vitro* diagnostic test for the qualitative detection and differentiation of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), influenza A virus, influenza B virus, and respiratory syncytial virus (RSV). Nucleic acids are isolated and purified from nasopharyngeal (NP) swab and anterior nasal (AN) swab 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 test is intended to aid in the differential diagnosis of SARS-CoV-2, influenza A, influenza B, and RSV A/B (undifferentiated) 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 and AN swab 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 Applied Biosystems TaqPath COVID-19, Flu A, Flu B, RSV Select Panel 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.
The Applied Biosystems TaqPath COVID-19, Flu A, Flu B, RSV Select Panel is intended for use by qualified and trained clinical laboratory personnel specifically instructed and trained in the techniques of real-time PCR and *in vitro* diagnostic procedures.
# **C Special Conditions for Use Statement(s):**
Rx - For Prescription Use Only
IVD - For *In Vitro* Diagnostic Use Only
# **D Special Instrument Requirements:**
This assay is to be used with the KingFisher Apex Dx Purification System and tested on the Applied Biosystems QuantStudio 5 Dx Real-Time PCR Instrument only.
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# IV Device/System Characteristics:
# A Device Description:
The TaqPath COVID-19, Flu A, Flu B, RSV Select Panel is a multiplex, real-time reverse transcription polymerase chain reaction (RT-PCR) test. The severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), influenza A, influenza B, respiratory syncytial virus (RSV) A/B and RNase P primer and probe sets are designed to detect viral RNA in nasopharyngeal (NP) and anterior nasal (AN) swab specimens from individuals exhibiting signs and symptoms of a respiratory tract infection. Each TaqPath COVID-19, Flu A, Flu B, RSV Select Panel includes the following components:
- TaqPath COVID-19, Flu A, Flu B, RSV Select Assay—Multiplex assays that contain primer and probe sets specific to the following targets:
- Three SARS-CoV-2 targets (Orf1a, Orf1b, and N genes);
- Two influenza A virus targets (PB1 and M genes);
- Two influenza B virus targets (M and NS genes);
- Three RSV targets (NP, M, and L protein genes);
- RNase P (internal human sample collection control).
- TaqPath COVID-19, Flu A, Flu B, RSV Select Positive Control—Inactivated viral control that contains SARS-CoV-2, influenza A, influenza B, and RSV.
- TaqPath COVID-19, Flu A, Flu B, RSV Select Negative Control—MS2 packaged RNA control that contains targets specific to RNase P genomic regions targeted by the assay.
- TaqPath 1-Step Select Master Mix (No ROX)—Ready-to-use PCR mix, including reverse transcriptase, polymerase, deoxyribonucleotide triphosphates (dNTPs), salts, and buffer.
In addition to the SARS-CoV-2, influenza A, influenza B and RSV viral assay targets, the assay portion of the panel includes RNase P, which serves as an endogenous internal process control to monitor extraction and amplification of each clinical sample. The TaqPath COVID-19, Flu A, Flu B, RSV Select Panel also contains external process positive and negative controls. The positive control (PC) component included is an inactivated viral control that contains SARS-CoV-2, influenza A, influenza B, and RSV viruses. The PC monitors extraction and real-time RT-PCR by demonstrating that each of the four viruses can be detected when present and that RNase P is not detected when absent. The negative control (NC) component included is an MS2 packaged RNA control that contains targets specific to RNase P genomic regions targeted by the assay. The NC also monitors extraction and real-time RT-PCR by demonstrating RNase P can be detected when present and that the four viruses are not detected when absent. The TaqPath 1-Step Select Master Mix (No ROX) included as a component of the kit is a ready-to-use PCR mix which contains a deoxyribonucleotide triphosphate mix (dNTPs), enzymes, and other components to permit reverse transcription and amplification of the assay targets. The TaqPath 1-Step Select Master Mix (No ROX) also contains ribonuclease (RNase) inhibitors as well as deoxyuridine triphosphate (dUTP) and uracil N-glycosylase (known as UNG or UDG).
The TaqPath COVID-19, Flu A, Flu B, RSV Select Panel workflow begins with nucleic acid extraction from nasopharyngeal and anterior nasal swab specimens stored in viral transport media (VTM) or universal transport medium (UTM). Nucleic acids are isolated and purified from the specimens using the MagMAX Dx Viral/Pathogen NA Isolation Kit with the
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KingFisher Apex Dx Purification System. In this process, nucleic acids from the patient samples are recovered using magnetic-bead technology.
The purified nucleic acid is reverse transcribed into cDNA and amplified using the TaqPath COVID-19, Flu A, Flu B, RSV Select Panel and the Applied Biosystems QuantStudio 5 Dx Real-Time PCR instrument. The Applied Biosystems Diomni Software is used for the TaqPath COVID-19, Flu A, Flu B, RSV Select Panel results interpretation and is supplied with the QuantStudio 5 Dx Real-Time PCR instrument.
# **B Principle of Operation:**
Once the reaction plate is loaded onto the real-time PCR instrument, RNA is reverse transcribed into cDNA. After heat inactivation of the reverse transcriptase enzyme, thermal cycling for PCR amplification commences.
During the anneal/extension phase of a PCR cycle, primers and probes specific to the target genes bind to corresponding DNA sequences in the indicated targets. 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 cycle, additional reporter dye molecules are cleaved from their respective probes, increasing the overall fluorescence intensity. Fluorescence intensity is monitored at each PCR cycle by the real-time PCR instrument.
The Diomni Software implements the thermal cycling program, secondary analysis and clinical interpretation of the results using assay-specific parameters from the Assay Definition File (ADF) that is installed in the software. When the software analyzes the run data, validation is performed automatically via quality check (QC) analysis. For a plate run, QC is based on the performance of the positive and negative controls; results interpretation of controls is described in **Table 1**. Individual samples are also subject to QC checks according to the rules provided in **Table 2**. A Positive call for RNase P is only required for samples that are negative for all four viral targets; samples with no amplification are called Invalid. The data and interpretive results for each run are saved as a batch in the software.
**Table 1.** Interpretation of Positive and Negative Controls
| Negative Control (NC) | | | | | Result (Well Call) | Status |
| --- | --- | --- | --- | --- | --- | --- |
| C19 (SARS-CoV-2) | Flu A | Flu B | RSV AB | RNase P | | |
| NEG | NEG | NEG | NEG | POS | Negative | Passed |
| All other scenarios | | | | | Invalid | Failed |
| Positive Control (PC) | | | | | Result (Well Call) | Status |
| C19 (SARS-CoV-2) | Flu A | Flu B | RSV AB | RNase P | | |
| POS | POS | POS | POS | NEG | Positive | Passed |
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| All other scenarios | Invalid | Failed |
| --- | --- | --- |
POS-Positive, NEG-Negative
**Table 2.** Result Interpretation for Patient Samples
| Target | | | | | Call | Result |
| --- | --- | --- | --- | --- | --- | --- |
| C19 (SARS-CoV-2) | Flu A | Flu B | RSV AB | RNase P | | |
| POS | POS | POS | POS | POS OR NEG | Positive | SARS-CoV-2, Flu A, Flu B, RSVAB Detected |
| POS | POS | POS | NEG | POS OR NEG | Positive | SARS-CoV-2, Flu A, Flu B Detected |
| POS | POS | NEG | POS | POS OR NEG | Positive | SARS-CoV-2, Flu A, RSVAB Detected |
| POS | POS | NEG | NEG | POS OR NEG | Positive | SARS-CoV-2, Flu A Detected |
| POS | NEG | POS | POS | POS OR NEG | Positive | SARS-CoV-2, Flu B, RSVAB Detected |
| POS | NEG | POS | NEG | POS OR NEG | Positive | SARS-CoV-2, Flu B Detected |
| POS | NEG | NEG | POS | POS OR NEG | Positive | SARS-CoV-2, RSVAB Detected |
| POS | NEG | NEG | NEG | POS OR NEG | Positive | SARS-CoV-2 Detected |
| NEG | POS | POS | POS | POS OR NEG | Positive | Flu A, Flu B, RSVAB Detected |
| NEG | POS | POS | NEG | POS OR NEG | Positive | Flu A, Flu B Detected |
| NEG | POS | NEG | POS | POS OR NEG | Positive | Flu A, RSVAB Detected |
| NEG | POS | NEG | NEG | POS OR NEG | Positive | Flu A Detected |
| NEG | NEG | POS | POS | POS OR NEG | Positive | Flu B, RSVAB Detected |
| NEG | NEG | POS | NEG | POS OR NEG | Positive | Flu B Detected |
| NEG | NEG | NEG | POS | POS OR NEG | Positive | RSVAB Detected |
| NEG | NEG | NEG | NEG | POS | Negative | SARS-CoV-2, Flu A, Flu B, RSVAB Not Detected |
| NEG | NEG | NEG | NEG | NEG | Invalid | RETEST^{[1]} |
$^{[1]}$ Retesting must be performed by re-extracting the original sample and repeating the real-time PCR. If the repeat result remains invalid, collection of a new specimen should be considered.
POS-Positive, NEG-Negative
## C Instrument Description Information:
### 1. Instrument Name:
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KingFisher Apex Dx Purification System
Applied Biosystems QuantStudio 5 Dx Real-Time PCR Instrument
2. Specimen Identification:
Specimen identification can be configured in an automated fashion or entered manually.
3. Specimen Sampling and Handling:
Nasopharyngeal and anterior nasal swab specimens eluted in viral transport media (VTM) or universal transport medium (UTM).
4. Quality Control:
Following controls are included in the TaqPath COVID-19, Flu A, Flu B, RSV Select Panel:
a) RNase P, which serves as an endogenous internal process control to monitor extraction and amplification of each clinical sample.
b) External Positive Control (PC) is an inactivated viral control that contains SARS-CoV-2, influenza A, influenza B, and RSV viruses. The PC is required for each plate run and monitors extraction and real-time RT-PCR by demonstrating that each of the four viruses can be detected when present and that RNase P is not detected when absent.
c) A Negative Control (NC) is an MS2 packaged RNA control that contains targets specific to RNase P genomic regions targeted by the assay. The NC is required for each plate run and also monitors extraction and real-time RT-PCR by demonstrating RNase P can be detected when present and that the four viruses are not detected when absent.
V Substantial Equivalence Information:
A Predicate Device Name(s):
BioFire Respiratory Panel 2.1 (RP2.1)
B Predicate 510(k) Number(s):
DEN200031
C Comparison with Predicate(s):
| Device & Predicate Device | K241806 | DEN200031 |
| --- | --- | --- |
| Device Trade Name | TaqPath COVID-19, Flu A, Flu B, RSV Select Panel | BioFire Respiratory Panel 2.1 (RP2.1) |
| Regulation Number | 21 CFR 866.3981 | Same |
| 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 | Same |
| Product Code | QOF | Same |
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| General Device Characteristics | | |
| --- | --- | --- |
| Intended Use/Indications For Use | The Applied Biosystems TaqPath COVID-19, Flu A, Flu B, RSV Select Panel is a multiplex, real-time reverse transcription polymerase chain reaction (RT-PCR) *in vitro* diagnostic test for the qualitative detection and differentiation of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), influenza A virus, influenza B virus, and respiratory syncytial virus (RSV). Nucleic acids are isolated and purified from nasopharyngeal (NP) swab and anterior nasal (AN) swab 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 test is intended to aid in the differential diagnosis of SARS-CoV-2, influenza A, influenza B, and RSV A/B (undifferentiated) 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 and AN swab 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 Applied Biosystems TaqPath COVID-19, Flu A, Flu B, RSV Select Panel may not be the definite cause of disease. Negative results do not preclude | The BioFire Respiratory Panel 2.1 (RP2.1) is a PCR-based multiplexed nucleic acid test intended for use with the BioFire FilmArray 2.0 or BioFire FilmArray Torch systems for the simultaneous qualitative detection and identification of multiple respiratory viral and bacterial nucleic acids in nasopharyngeal swabs (NPS) obtained from individuals suspected of respiratory tract infections, including COVID-19. The following organism types and subtypes are identified using the BioFire RP2.1: • Adenovirus, • Coronavirus 229E, • Coronavirus NL63, • Coronavirus OC43, • Coronavirus HKU1, • Severe Acute Respiratory Syndrome Coronavirus (SARS-CoV-2), • Human Metapneumovirus, • Human Rhinovirus/Enterovirus, • Influenza A, including subtypes H1, H1-2009, and H3, • Influenza B, • Parainfluenza Virus 1, • Parainfluenza Virus 2, • Parainfluenza Virus 3, • Parainfluenza Virus 4, • Respiratory Syncytial Virus, • *Bordetella parapertussis* (IS1001), • *Bordetella pertussis* (ptxP), • *Chlamydia pneumoniae*, and • *Mycoplasma pneumoniae* Nucleic acids from the respiratory viral and bacterial organisms identified by this test are generally detectable in NPS specimens during the acute phase of infection. The detection and identification of specific viral and bacterial nucleic acids from individuals exhibiting signs and/or symptoms of respiratory infection is indicative of the |
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| | SARS-CoV-2, influenza A virus, influenza B virus, or RSV infections. The Applied Biosystems TaqPath COVID-19, Flu A, Flu B, RSV Select Panel is intended for use by qualified and trained clinical laboratory personnel specifically instructed and trained in the techniques of real-time PCR and in vitro diagnostic procedures. | presence of the identified microorganism and aids in the diagnosis of respiratory infection if used in conjunction with other clinical and epidemiological information. The results of this test should not be used as the sole basis for diagnosis, treatment, or other patient management decisions. Negative results in the setting of a respiratory illness may be due to infection with pathogens that are not detected by this test, or lower respiratory tract infection that may not be detected by an NPS specimen. Positive results do not rule out coinfection with other organisms. The agent(s) detected by the BioFire RP2.1 may not be the definite cause of disease. Additional laboratory testing (e.g. bacterial and viral culture, immunofluorescence, and radiography) may be necessary when evaluating a patient with possible respiratory tract infection. |
| --- | --- | --- |
| Organisms Detected | SARS-CoV-2, Influenza A, Influenza B, Respiratory Syncytial Virus | Same |
| Specimen Types | Anterior Nasal Swabs and Nasopharyngeal swabs | Nasopharyngeal swabs |
| Transport Media | Universal Transport Medium (UTM) / Viral Transport Medium (VTM) | Same |
| Analyte | RNA | Same |
| Instrumentation | Sample Prep/Nucleic Acid Purification: • MagMAX Dx Viral/Pathogen NA Isolation Kit • KingFisher Apex Dx Purification System Real-time PCR instrument: • QuantStudio 5 Dx | Sample Prep/Nucleic Acid Purification: • Magnetic bead-based chemistry is contained within the BioFire RP2.1 pouch Instruments: • FilmArray 2.0 • FilmArray Torch |
| Technological Principles | Real-time Reverse Transcription PCR (RT-PCR) | Reverse Transcription PCR (RT-PCR) |
| Controls | One negative control and one positive control are run for each plate (external process controls). The assay detects endogenous RNase P present in the | Two internal controls are included in each reagent pouch for quality control of sample processing and both PCR stages and melt analysis. |
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| | specimen as an internal positive/process control. | |
| --- | --- | --- |
| Test Interpretation | Automated test interpretation | Same |
## VI Standards/Guidance Documents Referenced:
- • CLSI EP37. Supplemental Tables for Interference Testing in Clinical Chemistry; First Edition.
- • CLSI EP05-A3. Evaluation of Precision of Quantitative Measurement Procedures; Approved Guideline – Third Edition.
- • CLSI EP07. Interference Testing in Clinical Chemistry; Third Edition.
- • CLSI EP24-A2. Assessment of the Diagnostic Accuracy of Laboratory Testing Using Receiver Operating Characteristic Curves; Approved Guideline – Second Edition.
- • CLSI EP25-A. Evaluation of Stability of *In Vitro* Diagnostic Reagents; Approved Guideline.
- • CLSI EP35. Assessment of Equivalence or Suitability of Specimen Types for Medical Laboratory Measurement Procedures- First edition
- • CLSI MM06-A2. Quantitative Molecular Methods for Infectious Diseases; Approved Guideline - Second Edition
- • CLSI MM17. Verification and Validation of Multiplex Nucleic Acid Assays- Second Edition
- • ISO 23640:2011. In vitro diagnostic medical devices. Evaluation of stability of in vitro diagnostic reagents
- • ASTM D4169-22. Standard Practice for Performance Testing of Shipping Containers and Systems
- • ASTM F1980-21. Standard Guide for Accelerated Aging of Sterile Barrier Systems for Medical Devices
- • ISTA 7E. Testing Standard for Thermal Transport Packaging Used in Parcel Delivery System Shipment
- • AAMI TIR62366-2:2016. Medical devices-Part 2: Guidance on the application of usability engineering to medical devices
- • ISO 20916:2019. In vitro diagnostic medical devices. Clinical performance studies using specimens from human subjects. Good study practices.
### Special Controls
Class II Special Controls as per 21 CFR 866.3981
## VII Performance Characteristics (if/when applicable):
### A Analytical Performance:
#### 1. Precision/Reproducibility:
##### a. Within-laboratory precision
Within-laboratory precision was evaluated at a single site using the TaqPath COVID-19, Flu A, Flu B, RSV Select Panel on the QuantStudio 5 Dx (QS5Dx) instrument. A total of nine contrived samples containing known either a low (~1x LoD) or moderate (~3x LoD) concentrations of each target analyte (**Table 3**) and a negative sample were prepared in
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pooled negative NP specimens in VTM/UTM and extracted using the ThermoFisher MagMAX Dx Viral/Pathogen Nucleic Acid Isolation and the ThermoFisher KingFisher Apex Purification System. The study was conducted with two operators and one reagent lot over 12 non-consecutive days. Each panel member was tested in duplicate by each operator twice per day generating a total of 96 replicates per panel member (1 Site x 2 Operators x 1 Lot x 12 Days x 2 runs/day x 2 replicates/run). An extracted positive and negative control was included in each run and there were no invalid runs during precision studies.
**Table 3.** Precision Study Sample Panel
| Panel ID | Panel Member | Strain | Concentration |
| --- | --- | --- | --- |
| 1 | Negative | N/A | N/A |
| 2 | SARS-CoV-2 | USA-WA1/2020 | ~1x LoD |
| 3 | SARS-CoV-2 | USA-WA1/2020 | ~3x LoD |
| 4 | Influenza A | H3N2 Texas/50/12 | ~1x LoD |
| 5 | Influenza A | H3N2 Texas/50/12 | ~3x LoD |
| 6 | Influenza B | Victoria/504/00 | ~1x LoD |
| 7 | Influenza B | Victoria/504/00 | ~3x LoD |
| 8 | RSV | B CH93(18)-18 | ~1x LoD |
| 9 | RSV | B CH93(18)-18 | ~3x LoD |
The percent agreement with expected results and Cq variability analysis results from the study are shown in **Table 4** and **Table 5**, respectively.
All 96 replicates of each target analyte were positive 100% of the time for both low and moderate concentrations. The negative panel member generated negative results, as expected, for all samples (**Table 4**).
**Table 4.** Within-laboratory Precision Study Results
| Panel ID | Panel Member | Concentration | % Positive (pos n/valid n) | % Agreement with Expected Results (95% CI) |
| --- | --- | --- | --- | --- |
| 1 | Negative | N/A | 0% (0/96) | 100% (96.15-100%) |
| 2 | SARS-CoV-2 | ~1x LoD | 100% (96/96) | 100% (96.15-100%) |
| 3 | SARS-CoV-2 | ~3x LoD | 100% (96/96) | 100% (96.15-100%) |
| 4 | Influenza A | ~1x LoD | 100% (96/96) | 100% (96.15-100%) |
| 5 | Influenza A | ~3x LoD | 100% (96/96) | 100% (96.15-100%) |
| 6 | Influenza B | ~1x LoD | 100% (96/96) | 100% (96.15-100%) |
| 7 | Influenza B | ~3x LoD | 100% (96/96) | 100% (96.15-100%) |
| 8 | RSV | ~1x LoD | 100% (96/96) | 100% (96.15-100%) |
| 9 | RSV | ~3x LoD | 100% (96/96) | 100% (96.15-100%) |
The mean and variability analysis between operators, days, runs, within test, and overall are shown in **Table 5**.
**Table 5.** Within-laboratory Precision Study - Cq Analysis Results
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| Panel Member | LoD Conc. | N | Mean Cq | Between Operators | | Between Days | | Between Run | | Within Test | | Overall | |
| --- | --- | --- | --- | --- | --- | --- | --- | --- | --- | --- | --- | --- | --- |
| | | | | SD | % CV | SD | % CV | SD | % CV | SD | % CV | SD | % CV |
| Negative | N/A | 96 | 25.16 | 0.05 | 0.18 | 0.08 | 0.31 | 0.17 | 0.68 | 0.10 | 0.40 | 0.22 | 0.87 |
| SARS-CoV-2 | ~ 1x | 96 | 35.52 | 0.06 | 0.18 | 0.21 | 0.59 | 0.00 | 0.00 | 0.52 | 1.47 | 0.56 | 1.59 |
| SARS-CoV-2 | ~3x | 96 | 34.11 | 0.04 | 0.13 | 0.00 | 0.00 | 0.18 | 0.54 | 0.31 | 0.92 | 0.37 | 1.08 |
| Influenza A | ~ 1x | 96 | 33.05 | 0.00 | 0.00 | 0.10 | 0.30 | 0.07 | 0.20 | 0.24 | 0.73 | 0.27 | 0.82 |
| Influenza A | ~3x | 96 | 32.48 | 0.04 | 0.13 | 0.05 | 0.17 | 0.00 | 0.00 | 0.21 | 0.66 | 0.22 | 0.69 |
| Influenza B | ~ 1x | 96 | 34.03 | 0.04 | 0.10 | 0.00 | 0.00 | 0.13 | 0.38 | 0.25 | 0.73 | 0.28 | 0.83 |
| Influenza B | ~3x | 96 | 32.35 | 0.00 | 0.00 | 0.09 | 0.27 | 0.08 | 0.23 | 0.15 | 0.47 | 0.19 | 0.59 |
| RSV | ~ 1x | 96 | 34.28 | 0.00 | 0.00 | 0.00 | 0.00 | 0.15 | 0.45 | 0.42 | 1.22 | 0.44 | 1.30 |
| RSV | ~3x | 96 | 32.30 | 0.00 | 0.00 | 0.25 | 0.78 | 0.17 | 0.52 | 0.28 | 0.87 | 0.41 | 1.28 |
# **b. Reproducibility**
A blinded, multi-site reproducibility study was conducted to assess the total variability of the TaqPath COVID-19, Flu A, Flu B, RSV Select Panel across operators, study sites, testing days, runs, instruments, and reagent lots.
The same nine contrived panels evaluated in the Within-laboratory Precision Study (Table 3) were also tested in the Reproducibility Study. The study was performed across three independent sites, each with two operators per site across five non-consecutive days using the QS5Dx instrument. Each panel member was tested with each of the three (3) real-time PCR reagent lots in triplicate, for a total of nine (9) replicates per panel member in every run. At least one (1) Negative Control (NC) and one (1) Positive Control (PC) was included per run. For each panel member, a total of 270 replicates were generated per instrument (3 replicates/lot x 3 lots x 2 operators x 3 sites x 5 days).
The percent agreement with expected results and Cq variability analysis results from the study are summarized in Table 6 and Table 7.
**Table 6. Reproducibility Study Results**
| Analyte | Concentration | Agreement/Expected (% Agreement) | | | Overall Agreement (95% Score CI) |
| --- | --- | --- | --- | --- | --- |
| | | Site 1 | Site 2 | Site 3 | |
| Negative | Negative | 90/90 (100%) | 90/90 (100%) | 89/90 (98.89%)^{[1]} | 99.63% (97.93%, 99.93%) |
| SARS-CoV-2 | ~1x LoD | 85/90 (94.44%) | 82/90 (91.11%) | 83/90 (92.22%) | 92.59% (88.84%, 95.15%) |
| | ~3x LoD | 90/90 (100%) | 90/90 (100%) | 90/90 (100%) | 100% (98.60%, 100%) |
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| Influenza A | ~1x LoD | 88/90 (97.78%) | 90/90 (100%) | 89/90 (98.89%) | 98.89% (96.78%, 99.62%) |
| --- | --- | --- | --- | --- | --- |
| | ~3x LoD | 90/90 (100%) | 90/90 (100%) | 90/90 (100%) | 100% (98.60%, 100%) |
| Influenza B | ~1x LoD | 86/90 (95.56%) | 89/90 (98.89%) | 81/90 (90.00%) | 94.81% (91.49%, 96.89%) |
| | ~3x LoD | 90/90 (100%) | 90/90 (100%) | 90/90 (100%) | 100% (98.60%, 100%) |
| RSV | ~1x LoD | 87/90 (96.67%) | 87/90 (96.67%) | 90/90 (100%) | 97.78% (95.24%, 98.98%) |
| | ~3x LoD | 90/90 (100%) | 90/90 (100%) | 90/90 (100%) | 100% (98.60%, 100%) |
[1] One replicate was called positive for Flu A.
The mean and variability analyte between sites, operators, lots, days, runs, within-runs, and overall, for Ct values is shown in Table 7. A total of 270 replicates were performed for each panel member and variability was calculated based on samples that generated positive results.
Table 7. Reproducibility Study - Cq Analysis Results
| Analyte | Test level | Mean Cq | Between sites | | Between lots | | Between days | | Between Instruments/operators | | Repeatability | | Within-lab precision | | Reproducibility (total) | |
| --- | --- | --- | --- | --- | --- | --- | --- | --- | --- | --- | --- | --- | --- | --- | --- | --- |
| | | | SD | %CV | SD | %CV | SD | %CV | SD | %CV | SD | %CV | SD | %CV | SD | %CV |
| Negative | N/A | 25.35[1] | 0.28 | 1.11 | 0.06 | 0.24 | 0.00 | 0.00 | 0.50 | 1.99 | 0.10 | 0.39 | 0.52 | 2.04 | 0.59 | 2.32 |
| SARS-CoV-2 | 1x LoD | 36.09 | 0.29 | 0.79 | 0.12 | 0.33 | 0.00 | 0.00 | 0.39 | 1.09 | 0.66 | 1.82 | 0.77 | 2.14 | 0.82 | 2.29 |
| | 3x LoD | 34.50 | 0.26 | 0.76 | 0.08 | 0.23 | 0.26 | 0.75 | 0.23 | 0.67 | 0.61 | 1.77 | 0.71 | 2.05 | 0.75 | 2.18 |
| Influenza A | 1x LoD | 34.75 | 0.10 | 0.28 | 0.00 | 0.00 | 0.00 | 0.00 | 0.51 | 1.46 | 0.78 | 2.25 | 0.93 | 2.69 | 0.94 | 2.70 |
| | 3x LoD | 33.05 | 0.05 | 0.16 | 0.00 | 0.00 | 0.00 | 0.00 | 0.50 | 1.51 | 0.54 | 1.65 | 0.74 | 2.23 | 0.74 | 2.24 |
| Influenza B | 1x LoD | 35.01 | 0.33 | 0.96 | 0.00 | 0.00 | 0.22 | 0.63 | 0.20 | 0.56 | 0.88 | 2.51 | 0.93 | 2.65 | 0.98 | 2.81 |
| | 3x LoD | 33.50 | 0.47 | 1.40 | 0.00 | 0.00 | 0.00 | 0.00 | 0.50 | 1.50 | 0.70 | 2.08 | 0.86 | 2.56 | 0.98 | 2.92 |
| | 1x LoD | 35.43 | 0.30 | 0.86 | 0.00 | 0.00 | 0.00 | 0.00 | 0.49 | 1.37 | 1.24 | 3.50 | 1.33 | 3.76 | 1.37 | 3.85 |
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| RSV | 3x LoD | 33.81 | 0.04 | 0.11 | 0.14 | 0.42 | 0.00 | 0.00 | 0.65 | 1.93 | 1.13 | 3.34 | 1.31 | 3.88 | 1.31 | 3.88 |
| --- | --- | --- | --- | --- | --- | --- | --- | --- | --- | --- | --- | --- | --- | --- | --- | --- |
[1] Mean Cq, SD, and %CV calculated for RNase P only.
## 2. Linearity:
Linearity is not applicable, as the TaqPath COVID-19, Flu A, Flu B, RSV Select Panel is a qualitative test.
## 3. Analytical Specificity/Interference:
### a. Interfering Substances
Potential interference of the TaqPath COVID-19, Flu A, Flu B, RSV Select Panel was assessed by testing contrived positive samples consisting of one strain each of SARS-CoV-2 (USA-WA1/2020), influenza A (H1N1pdm, Michigan/45/15), influenza B (Yamagata, Florida/04/06) and RSV (RSVB/CH93(18)-18) at 3x their respective limits of detection (LoDs) as well as negative samples in the presence or absence of the twenty-six (26) potentially interfering substances, two (2) solvents, one (1) no interferent control, and five (5) viral transport media (VTM). Additionally, potential interference due to extraction reagent carryover was assessed with four (4) carryover conditions and one (1) no-carryover control. For all potential interferents other than VTM, contrived samples were prepared in pooled negative NP swab specimens in VTM/UTM. VTM testing used pooled negative AN swab specimens that had been collected in the VTM. Each substance was tested in triplicate (N=3) for each positive and negative sample through the full workflow with one (1) lot of reagents on the QS5Dx instrument.
Detection of both Flu A and Flu B in all samples tested with FluMist at 0.05%, 0.01% and 0.005% v/v was expected since it is a live-attenuated virus vaccine. The presence of FluMist did not result in false positive or false negative results for the other targets at any of the three concentrations tested. None of the other exogenous or endogenous substances, transport media or extraction kit reagents tested interfered with the TaqPath COVID-19, Flu A, Flu B, RSV Select Panel at the concentrations tested. The results from the study are presented in **Table 8**.
**Table 8.** Summary of Interfering Substances Study Results
| Substance Description | Conc. | NEG | Influenza A | | Influenza B | | SARS-CoV-2 | | RSV | |
| --- | --- | --- | --- | --- | --- | --- | --- | --- | --- | --- |
| | | % Positive (nPositive / Total) | % Positive (nPositive/ Total) | Mean Cq | % Positive (nPositive/ Total) | Mean Cq | % Positive (nPositive/ Total) | Mean Cq | % Positive (nPositive/ Total) | Mean Cq |
| No interferent control (NP) | N/A | 0% (0/9) | 100% (6/6) | 33.20 | 100% (9/9) | 33.32 | 100% (6/6) | 34.20 | 100% (6/6) | 33.42 |
| Mucin | 5 mg/mL | 0% (0/3) | 100% (3/3) | 33.94 | 100% (3/3) | 33.23 | 100% (3/3) | 35.20 | 100% (3/3) | 33.23 |
| Whole blood | 5% (v/v) | 0% (0/3) | 100% (3/3) | 32.76 | 100% (3/3) | 33.08 | 100% (3/3) | 34.09 | 100% (3/3) | 32.78 |
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| Jurkat cell gDNA | 7 μg/mL | 0% (0/3) | 100% (3/3) | 33.65 | 100% (3/3) | 33.09 | 100% (3/3) | 34.02 | 100% (3/3) | 33.15 |
| --- | --- | --- | --- | --- | --- | --- | --- | --- | --- | --- |
| Nasal decongestant – Phenylephrine (Equate) | 10% (v/v) | 0% (0/3) | 100% (3/3) | 32.23 | 100% (3/3) | 33.30 | 100% (3/3) | 34.72 | 100% (3/3) | 33.60 |
| Nasal decongestant – Oxymetazoline (Afrin) | 10% (v/v) | 0% (0/3) | 100% (3/3) | 30.50 | 100% (3/3) | 31.20 | 100% (3/3) | 34.42 | 100% (3/3) | 32.83 |
| Saline nasal spray (Equate) | 10% (v/v) | 0% (0/3) | 100% (3/3) | 32.69 | 100% (3/3) | 33.04 | 100% (3/3) | 34.04 | 100% (3/3) | 33.29 |
| Bronchodilator (Ventolin Evohaler) | 0.83 mg/mL | 0% (0/3) | 100% (3/3) | 33.67 | 100% (3/3) | 33.35 | 100% (3/3) | 34.26 | 100% (3/3) | 33.51 |
| Nasal corticosteroids – Dexamethasone | 1.5 mg/mL | 0% (0/3) | 100% (3/3) | 33.60 | 100% (3/3) | 32.87 | 100% (3/3) | 34.44 | 100% (3/3) | 33.48 |
| Nasal corticosteroids – Flunisolide | 2 mg/mL | 0% (0/3) | 100% (3/3) | 34.16 | 100% (3/3) | 32.93 | 100% (3/3) | 34.78 | 100% (3/3) | 31.27 |
| Nasal corticosteroids – Budesonide (Apotex) | 1% (v/v) | 0% (0/3) | 100% (3/3) | 33.71 | 100% (3/3) | 33.00 | 100% (3/3) | 34.10 | 100% (3/3) | 32.73 |
| Nasal corticosteroids – Fluticasone (Equate) | 1% (v/v) | 0% (0/3) | 100% (3/3) | 33.17 | 100% (3/3) | 33.57 | 100% (3/3) | 33.74 | 100% (3/3) | 33.11 |
| Nasal gel (NeilMed Nasogel) | 1% (w/v) | 0% (0/3) | 100% (3/3) | 33.38 | 100% (3/3) | 33.27 | 100% (3/3) | 34.60 | 100% (3/3) | 33.29 |
| Homeopathic allergy relief medicine (Zicam) | 1% (w/v) | 0% (0/3) | 100% (3/3) | 33.36 | 100% (3/3) | 33.84 | 100% (3/3) | 34.74 | 100% (3/3) | 33.12 |
| Throat lozenges (Chloraseptic Max) | 2.2 mg/mL methanol; 3.3 mg/mL benzocaine | 0% (0/3) | 100% (3/3) | 33.71 | 100% (3/3) | 33.46 | 100% (3/3) | 34.33 | 100% (3/3) | 33.25 |
| Zinc lozenges (Life Extension) | 7.5 mg/mL | 0% (0/3) | 100% (3/3) | 33.69 | 100% (3/3) | 33.17 | 100% (3/3) | 34.06 | 100% (3/3) | 33.20 |
| Antiviral – Zanamivir | 5.5 mg/mL | 0% (0/6) | 100% (4/4) | 33.46 | 100% (6/6) | 32.95 | 100% (3/3) | 34.33 | 100% (3/3) | 33.39 |
| Antiviral - Oseltamivir phosphate | 33 μg/mL | 0% (0/3) | 100% (3/3) | 33.25 | 100% (3/3) | 31.93 | 100% (3/3) | 34.52 | 100% (3/3) | 33.38 |
| Antiviral – Remdesivir | 6.7 μg/mL | 0% (0/3) | 100% (3/3) | 33.45 | 100% (3/3) | 33.48 | 100% (3/3) | 33.88 | 100% (3/3) | 34.03 |
| Topical antibiotic (Pseudomonic Acid) | 3.3 mg/mL | 0% (0/3) | 100% (3/3) | 32.09 | 100% (3/3) | 33.06 | 100% (3/3) | 34.56 | 100% (3/3) | 32.43 |
| Systemic antibiotic (Tobramycin) | 4 μg/mL | 0% (0/3) | 100% (3/3) | 33.33 | 100% (3/3) | 33.24 | 100% (3/3) | 34.40 | 100% (3/3) | 32.92 |
| | 0.05% (v/v) | 100% (3/3) [1] | 100% (3/3) | 21.59 | 100% (3/3) | 20.85 | 100% (3/3) | 33.19 | 100% (3/3) | 32.82 |
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| Flu vaccine: FluMist | 0.01% (v/v) | 100% (3/3) [1] | 100% (3/3) | 23.89 | 100% (3/3) | 23.36 | 100% (3/3) | 32.85 | 100% (3/3) | 32.99 |
| --- | --- | --- | --- | --- | --- | --- | --- | --- | --- | --- |
| | 0.005% (v/v) | 100% (3/3) [1] | 100% (3/3) | 24.86 | 100% (3/3) | 24.24 | 100% (3/3) | 33.37 | 100% (3/3) | 32.88 |
| Analgesic (Ibuprofen) | 220 μg/mL | 0% (0/3) | 100% (3/3) | 33.30 | 100% (3/3) | 32.69 | 100% (3/3) | 34.00 | 100% (3/3) | 33.48 |
| Tobacco (Camel Snus) | 1% (w/v) | 0% (0/3) | 100% (3/3) | 32.93 | 100% (3/3) | 32.99 | 100% (3/3) | 33.85 | 100% (3/3) | 32.82 |
| Solvent DMSO | 20% (v/v) | 0% (0/6) | 100% (3/3) | 33.53 | 100% (6/6) | 32.74 | 100% (3/3) | 34.69 | 100% (3/3) | 31.27 |
| Solvent Ethanol | 20% (v/v) | 0% (0/3) | 100% (3/3) | 32.34 | 100% (3/3) | 32.79 | 100% (3/3) | 34.01 | 100% (3/3) | 33.60 |
| VTM Remel M4RT | Neat[2] | 0% (0/3) | 100% (3/3) | 33.35 | 100% (3/3) | 33.37 | 100% (3/3) | 34.73 | 100% (3/3) | 32.10 |
| VTM Remel M5 | Neat[2] | 0% (0/3) | 100% (3/3) | 33.00 | 100% (3/3) | 33.19 | 100% (3/3) | 34.76 | 100% (3/3) | 33.06 |
| VTM Remel M6 | Neat[2] | 0% (0/3) | 100% (3/3) | 33.31 | 100% (3/3) | 33.05 | 100% (3/3) | 35.42 | 100% (3/3) | 33.15 |
| VTM Copan UTM- RT | Neat[2] | 0% (0/3) | 100% (3/3) | 32.88 | 100% (3/3) | 31.92 | 100% (3/3) | 34.57 | 100% (3/3) | 32.76 |
| VTM BD UVT | Neat[2] | 0% (0/3) | 100% (3/3) | 31.77 | 100% (3/3) | 33.40 | 100% (3/3) | 34.47 | 100% (3/3) | 32.96 |
| No Extraction Reagent Carryover Control | N/A | 0% (0/3) | 100% (3/3) | 33.43 | 100% (3/3) | 32.97 | 100% (3/3) | 33.94 | 100% (3/3) | 33.26 |
| Extraction Reagent Carryover - Sample Plate | 1% (v/v)[3] | 0% (0/3) | 100% (3/3) | 33.47 | 100% (3/3) | 33.06 | 100% (3/3) | 34.59 | 100% (3/3) | 33.52 |
| Extraction Reagent Carryover - Wash 1 | 1% (v/v)[3] | 0% (0/3) | 100% (3/3) | 33.67 | 100% (3/3) | 33.44 | 100% (3/3) | 34.25 | 100% (3/3) | 33.33 |
| Extraction Reagent Carryover - Wash 2 | 6% (v/v)[3] | 0% (0/3) | 100% (3/3) | 33.90 | 100% (3/3) | 33.13 | 100% (3/3) | 34.02 | 100% (3/3) | 33.54 |
| | 3% (v/v)[3] | 0% (0/3) | 100% (3/3) | 33.32 | 100% (3/3) | 32.79 | 100% (3/3) | 34.41 | 100% (3/3) | 33.33 |
[1] Negative samples containing FluMist were positive for Flu A and Flu B and negative for SARS-CoV-2 and RSV
[2] Neat = undiluted; AN samples originally collected in indicated VTM
[3] Interferent added to eluates (post-extraction), and concentrations are per volume of eluate
### b. Competitive Interference
In this study, the ability to detect a co-infection when one targeted virus is present at a high concentration (10⁵ TCID₅₀/mL) and another targeted virus is present at a low concentration (3x LoD) was assessed using contrived NP samples in VTM/UTM containing all possible pairings of influenza A, influenza B, SARS-CoV-2 and RSV. In parallel, viruses were tested at both levels individually to serve as controls and serve to demonstrate within-panel specificity of each of the target channels. Sample extraction was performed using the KingFisher Apex, and real-time RT-PCR was performed on the QS5Dx with one (1) lot of TaqPath COVID-19, Flu A, Flu B, RSV Select Panel reagents.
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No off-target false-positive results were observed in these test conditions, which demonstrates within-panel specificity for each of the target channels. Negative, pooled NP specimens were tested alone (with no virus) as a negative control condition and yielded negative test results. As shown in **Table 9**, no competitive interference was observed at the concentrations tested.
**Table 9.** Summary of Competitive Interference Study Results
| Viral Test Condition Combination | Low Target | | High Target | |
| --- | --- | --- | --- | --- |
| | Virus (Concentration) | nPositive/ Total (% Positivity) | Virus (Concentration) | nPositive/ Total (% Positivity) |
| SARS-CoV-2 (low), Influenza A (high) | SARS-CoV-2 (3x LoD) | 3 / 3 (100%) | Influenza A (1.0 × 10^{5} TCID_{50}/mL) | 3 / 3 (100%) |
| SARS-CoV-2 (low), Influenza B (high) | SARS-CoV-2 (3x LoD) | 3 / 3 (100%) | Influenza B (1.0 × 10^{5} TCID_{50}/mL) | 3 / 3 (100%) |
| SARS-CoV-2 (low), RSV (high) | SARS-CoV-2 (3x LoD) | 3 / 3 (100%) | RSV (1.0 × 10^{5} TCID_{50}/mL) | 3 / 3 (100%) |
| Influenza A (low), SARS-CoV-2 (high) | Influenza A (3x LoD) | 3 / 3 (100%) | SARS-CoV-2 (1.0 × 10^{5} TCID_{50}/mL) | 3 / 3 (100%) |
| Influenza A (low), Influenza B (high) | Influenza A (3x LoD) | 3 / 3 (100%) | Influenza B (1.0 × 10^{5} TCID_{50}/mL) | 3 / 3 (100%) |
| Influenza A (low), RSV (high) | Influenza A (3x LoD) | 3 / 3 (100%) | RSV (1.0 × 10^{5} TCID_{50}/mL) | 3 / 3 (100%) |
| Influenza B (low), SARS-CoV-2 (high) | Influenza B (3x LoD) | 3 / 3 (100%) | SARS-CoV-2 (1.0 × 10^{5} TCID_{50}/mL) | 3 / 3 (100%) |
| Influenza B (low), Influenza A (high) | Influenza B (3x LoD) | 3 / 3 (100%) | Influenza A (1.0 × 10^{5} TCID_{50}/mL) | 3 / 3 (100%) |
| Influenza B (low), RSV (high) | Influenza B (3x LoD) | 3 / 3 (100%) | RSV (1.0 × 10^{5} TCID_{50}/mL) | 3 / 3 (100%) |
| RSV (low), SARS-CoV-2 (high) | RSV (3x LoD) | 3 / 3 (100%) | SARS-CoV-2 (1.0 × 10^{5} TCID_{50}/mL) | 3 / 3 (100%) |
| RSV (low), Influenza A (high) | RSV (3x LoD) | 3 / 3 (100%) | Influenza A (1.0 × 10^{5} TCID_{50}/mL) | 3 / 3 (100%) |
| RSV (low), Influenza B (high) | RSV (3x LoD) | 3 / 3 (100%) | Influenza B (1.0 × 10^{5} TCID_{50}/mL) | 3 / 3 (100%) |
| SARS-CoV-2 (low) | SARS-CoV-2 (3x LoD) | 3 / 3 (100%) | N/A | N/A |
| SARS-CoV-2 (high) | N/A | N/A | SARS-CoV-2 (1.0 × 10^{5} TCID_{50}/mL) | 3 / 3 (100%) |
| Influenza A (low) | Influenza A (3x LoD) | 3 / 3 (100%) | N/A | N/A |
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| Influenza A (high) | N/A | N/A | Influenza A (1.0 × 10^{5} TCID_{50}/mL) | 3 / 3 (100%) |
| --- | --- | --- | --- | --- |
| Influenza B (low) | Influenza B (3x LoD) | 3 / 3 (100%) | N/A | N/A |
| Influenza B (high) | N/A | N/A | Influenza B (1.0 × 10^{5} TCID_{50}/mL) | 3 / 3 (100%) |
| RSV (low) | RSV (3x LoD) | 3 / 3 (100%) | N/A | N/A |
| RSV (high) | N/A | N/A | RSV (1.0 × 10^{5} TCID_{50}/mL) | 3 / 3 (100%) |
| Negative (NP alone) | N/A | N/A | N/A | N/A |
### c. Cross-Reactivity and Microbial Interference Wet-Testing
Cross-reactivity and microbial interference was assessed for the TaqPath COVID-19, Flu A, Flu B, RSV Select Panel. Testing included microbes that are commonly found in upper respiratory samples, including commensal flora and pathogenic microbes. In the cross-reactivity portion of the study, contrived samples were prepared by spiking microbes into negative pooled nasopharyngeal (NP) swab samples in UTM/VTM to determine whether false positive results may be produced for any of the targets of the test. In addition to contrived samples, results from samples tested during the Clinical Validation study that had been called positive for coronavirus HKU1 (CoV-HKU1) by the comparator test were assessed for evidence of cross-reactivity with SARS-CoV-2 (USA-WA1/2020), influenza A (Michigan/45/15), influenza B (Florida/04/06) or RSV (CH93(18)-18). In the microbial interference portion of the study, the same microorganisms were tested in the presence of one representative viral strain for each of SARS-CoV-2, influenza A, influenza B, and RSV at 3x LoD to determine whether false negative results may be produced for the spiked targets. Each sample was tested in triplicate through the full workflow with one lot of reagents on the QS5Dx instrument. Bacteria and fungi were tested at 10$^{6}$ CFU/mL and viruses were tested at 10$^{5}$ TCID$_{50}$/mL, or the highest concentration possible. As shown in **Table 10**, no cross-reactivity or microbial interference was observed at the concentrations tested.
**Table 10.** Summary of Cross-Reactivity and Microbial Interference Study Results
| Organism type | Species | Test Concentration | Negative Samples | Number positive / total (% Positive) | | | |
| --- | --- | --- | --- | --- | --- | --- | --- |
| | | | Number Negative / Total (% Negative) | SARS-CoV-2 | Influenza A | Influenza B | RSV |
| | *Bordetella parapertussis* | 1 x 10^{6} CFU/mL | 3/3 (100%) | 3/3 (100%) | 3/3 (100%) | 3/3 (100%) | 3/3 (100%) |
| | *Bordetella pertussis* | 1 x 10^{6} CFU/mL | 3/3 (100%) | 3/3 (100%) | 3/3 (100%) | 3/3 (100%) | 3/3 (100%) |
| | *Chlamydophila pneumoniae* | 1 x 10^{6} CFU/mL | 3/3 (100%) | 3/3 (100%) | 3/3 (100%) | 3/3 (100%) | 3/3 (100%) |
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| Bacteria | Corynebacterium diphtheriae | 1 x 10^{6} CFU/mL | 3/3 (100%) | 3/3 (100%) | 3/3 (100%) | 3/3 (100%) | 3/3 (100%) |
| --- | --- | --- | --- | --- | --- | --- | --- |
| | Escherichia coli | 1 x 10^{6} CFU/mL | 3/3 (100%) | 3/3 (100%) | 3/3 (100%) | 3/3 (100%) | 3/3 (100%) |
| | Fusobacterium necrophorum | 1 x 10^{6} CFU/mL | 3/3 (100%) | 3/3 (100%) | 3/3 (100%) | 3/3 (100%) | 3/3 (100%) |
| | Haemophilus influenzae type b | 1 x 10^{6} CFU/mL | 3/3 (100%) | 3/3 (100%) | 3/3 (100%) | 3/3 (100%) | 3/3 (100%) |
| | Klebsiella pneumoniae | 1 x 10^{6} CFU/mL | 3/3 (100%) | 3/3 (100%) | 3/3 (100%) | 3/3 (100%) | 3/3 (100%) |
| | Lactobacillus acidophilus | 1 x 10^{6} CFU/mL | 3/3 (100%) | 3/3 (100%) | 3/3 (100%) | 3/3 (100%) | 3/3 (100%) |
| | Legionella pneumophila | 1 x 10^{6} CFU/mL | 3/3 (100%) | 3/3 (100%) | 3/3 (100%) | 3/3 (100%) | 3/3 (100%) |
| | Moraxella catarrhalis | 1 x 10^{6} CFU/mL | 3/3 (100%) | 3/3 (100%) | 3/3 (100%) | 3/3 (100%) | 3/3 (100%) |
| | Mycobacterium tuberculosis | 5.4 x 10^{7} copies/mL | 3/3 (100%) | 3/3 (100%) | 3/3 (100%) | 3/3 (100%) | 3/3 (100%) |
| | Mycoplasma genitalium | 4 x 10^{5} cells/mL | 3/3 (100%) | 3/3 (100%) | 3/3 (100%) | 3/3 (100%) | 3/3 (100%) |
| | Mycoplasma pneumoniae | 1 x 10^{6} CFU/mL | 3/3 (100%) | 3/3 (100%) | 3/3 (100%) | 3/3 (100%) | 3/3 (100%) |
| | Neisseria elongata | 1 x 10^{6} CFU/mL | 3/3 (100%) | 3/3 (100%) | 3/3 (100%) | 3/3 (100%) | 3/3 (100%) |
| | Neisseria meningitidis serogroup A | 1 x 10^{6} CFU/mL | 3/3 (100%) | 3/3 (100%) | 3/3 (100%) | 3/3 (100%) | 3/3 (100%) |
| | Pseudomonas aeruginosa | 1 x 10^{6} CFU/mL | 3/3 (100%) | 3/3 (100%) | 3/3 (100%) | 3/3 (100%) | 3/3 (100%) |
| | Staphylococcus aureus MRSA | 1 x 10^{6} CFU/mL | 3/3 (100%) | 3/3 (100%) | 3/3 (100%) | 3/3 (100%) | 3/3 (100%) |
| | Staphylococcus epidermidis MRSE | 1 x 10^{6} CFU/mL | 3/3 (100%) | 3/3 (100%) | 3/3 (100%) | 3/3 (100%) | 3/3 (100%) |
| | Streptococcus pneumoniae | 1 x 10^{6} CFU/mL | 3/3 (100%) | 3/3 (100%) | 3/3 (100%) | 3/3 (100%) | 3/3 (100%) |
| | Streptococcus pyogenes | 1 x 10^{6} CFU/mL | 3/3 (100%) | 3/3 (100%) | 3/3 (100%) | 3/3 (100%) | 3/3 (100%) |
| | Streptococcus salivarius | 1 x 10^{6} CFU/mL | 3/3 (100%) | 3/3 (100%) | 3/3 (100%) | 3/3 (100%) | 3/3 (100%) |
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| Fungi | *Aspergillus flavus* | 1 x 10^{6} CFU/mL | 3/3 (100%) | 3/3 (100%) | 3/3 (100%) | 3/3 (100%) | 3/3 (100%) |
| --- | --- | --- | --- | --- | --- | --- | --- |
| | *Candida albicans* | 1 x 10^{6} CFU/mL | 3/3 (100%) | 3/3 (100%) | 3/3 (100%) | 3/3 (100%) | 3/3 (100%) |
| | *Pneumocystis carinii* | 1 x 10^{6} Cells/mL | 3/3 (100%) | 3/3 (100%) | 3/3 (100%) | 3/3 (100%) | 3/3 (100%) |
| Virus | Adenovirus type 1, species C | 1 x 10^{5} TCID_{50}/mL | 3/3 (100%) | 3/3 (100%) | 3/3 (100%) | 3/3 (100%) | 3/3 (100%) |
| | Mastadenovirus B type 7, Gomen | 1 x 10^{5} TCID_{50}/mL | 3/3 (100%) | 3/3 (100%) | 3/3 (100%) | 3/3 (100%) | 3/3 (100%) |
| | Cytomegalovirus (HHV-5) | 3.2 x 10^{4} TCID_{50}/mL | 3/3 (100%) | 3/3 (100%) | 3/3 (100%) | 3/3 (100%) | 3/3 (100%) |
| | Enterovirus type 68 | 1 x 10^{5} TCID_{50}/mL | 3/3 (100%) | 3/3 (100%) | 3/3 (100%) | 3/3 (100%) | 3/3 (100%) |
| | Epstein-Barr virus (HHV-4) | 1.57 x 10^{7} copies/mL | 3/3 (100%) | 3/3 (100%) | 3/3 (100%) | 3/3 (100%) | 3/3 (100%) |
| | Hepatitis B virus | 4.2 x 10^{7} IU/mL | 3/3 (100%) | 3/3 (100%) | 3/3 (100%) | 3/3 (100%) | 3/3 (100%) |
| | Hepatitis C virus | 3.3 x 10^{7} IU/mL | 3/3 (100%) | 3/3 (100%) | 3/3 (100%) | 3/3 (100%) | 3/3 (100%) |
| | Herpes simplex virus 1 (HSV-1) | 1 x 10^{5} TCID_{50}/mL | 3/3 (100%) | 3/3 (100%) | 3/3 (100%) | 3/3 (100%) | 3/3 (100%) |
| | Human coronavirus 229E | 2.82 x 10^{4} TCID_{50}/mL | 3/3 (100%) | 3/3 (100%) | 3/3 (100%) | 3/3 (100%) | 3/3 (100%) |
| | Human coronavirus HKU1^{[1]} | 2 x 10^{6} copies/mL | 3/3 (100%) | 3/3 (100%) | 3/3 (100%) | 3/3 (100%) | 3/3 (100%) |
| | Human coronavirus HKU1 | 39 positive clinical specimens | 38/39 (97.4%)^{[2]} | Not tested | Not tested | Not tested | Not tested |
| | Human coronavirus NL63 | 2.34 x 10^{4} TCID_{50}/mL | 3/3 (100%) | 3/3 (100%) | 3/3 (100%) | 3/3 (100%) | 3/3 (100%) |
| | Human coronavirus OC43 | 1 x 10^{5} TCID_{50}/mL | 3/3 (100%) | 3/3 (100%) | 3/3 (100%) | 3/3 (100%) | 3/3 (100%) |
| | Human immunodeficiency virus type IIIB | 1 x 10^{4} IU/mL | 3/3 (100%) | 3/3 (100%) | 3/3 (100%) | 3/3 (100%) | 3/3 (100%) |
| | Human metapneumovirus | 1 x 10^{5} TCID_{50}/mL | 3/3 (100%) | 3/3 (100%) | 3/3 (100%) | 3/3 (100%) | 3/3 (100%) |
| Measles virus | 1 x 10^{5} TCID_{50}/mL | 3/3 (100%) | 3/3 (100%) | 3/3 (100%) | 3/3 (100%) | 3/3 (100%) | |
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| | MERS coronavirus strain Florida/USA-2 Saudi Arabia 2014 | Not provided ^{[3]} | 3/3 (100%) | 3/3 (100%) | 3/3 (100%) | 3/3 (100%) | 3/3 (100%) |
| --- | --- | --- | --- | --- | --- | --- | --- |
| | MERS coronavirus strain EMC/2012 | 1 x 10^{5} TCID_{50}/mL | 3/3 (100%) | 3/3 (100%) | 3/3 (100%) | 3/3 (100%) | 3/3 (100%) |
| | Mumps virus | 1 x 10^{5} TCID_{50}/mL | 3/3 (100%) | 3/3 (100%) | 3/3 (100%) | 3/3 (100%) | 3/3 (100%) |
| | Parainfluenza virus type 1 | 1 x 10^{5} TCID_{50}/mL | 3/3 (100%) | 3/3 (100%) | 3/3 (100%) | 3/3 (100%) | 3/3 (100%) |
| | Parainfluenza virus type 2 | 1 x 10^{5} TCID_{50}/mL | 3/3 (100%) | 3/3 (100%) | 3/3 (100%) | 3/3 (100%) | 3/3 (100%) |
| | Parainfluenza virus type 3 | 1 x 10^{5} TCID_{50}/mL | 3/3 (100%) | 3/3 (100%) | 3/3 (100%) | 3/3 (100%) | 3/3 (100%) |
| | Parainfluenza virus type 4A | 1 x 10^{5} TCID_{50}/mL | 3/3 (100%) | 3/3 (100%) | 3/3 (100%) | 3/3 (100%) | 3/3 (100%) |
| | Rhinovirus type 1A | 2.82 x 10^{4} TCID_{50}/mL | 3/3 (100%) | 3/3 (100%) | 3/3 (100%) | 3/3 (100%) | 3/3 (100%) |
| | SARS coronavirus strain 2003-00592 | Not provided ^{[3]} | 3/3 (100%) | 3/3 (100%) | 3/3 (100%) | 3/3 (100%) | 3/3 (100%) |
| | SARS coronavirus strain Urbani | 1 x 10^{5} PFU/mL | 3/3 (100%) | 3/3 (100%) | 3/3 (100%) | 3/3 (100%) | 3/3 (100%) |
| | Varicella-zoster virus (HHV-3) | 1.94 x 10^{8} copies/mL | 3/3 (100%) | 3/3 (100%) | 3/3 (100%) | 3/3 (100%) | 3/3 (100%) |
| Biofluid | Pooled human nasal wash | Neat | 3/3 (100%) | 3/3 (100%) | 3/3 (100%) | 3/3 (100%) | 3/3 (100%) |
$^{[1]}$ Subgenomic synthetic control (ATCC VR-3262SD)
$^{[2]}$ One sample produced a positive for SARS-CoV-2.
$^{[3]}$ Concentration not provided by the vendor.
#### 4. Assay Reportable Range:
Not applicable; this is a qualitative assay.
#### 5. Traceability, Stability, Expected Values (Controls, Calibrators, or Methods):
##### a. Quality Controls
Please refer to Instrument Description Information (Section C.4) above for assay controls.
##### b. Specimen Stability
Specimen stability testing was performed on pooled, negative NP and AN swab specimens in VTM/UTM, contrived with influenza A (H1N1pdm, California/07/09), influenza B (Yamagata, Florida/04/06), SARS-CoV-2 (USA-WA1/2020) and RSV (RSVA, 2013 Isolate) individually spiked in each respective matrix at 3x LoD. Samples were stored at room temperature (30°C), refrigerated (8°C) and frozen (-30°C to -10°C
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and -70°C) temperatures for specified time periods prior to testing with the TaqPath SARS-CoV-2, Flu A, Flu B, RSV Select Panel. The test condition for samples stored at 30°C included an additional storage of those samples at 8°C for 80 hours after completion of the specified time periods at 30°C. Five (5) replicates of samples for each storage condition in each specimen matrix were tested. Two (2) replicates of pooled negative NP and AN swab specimens in UTM/VTM were also tested at all storage conditions. Samples were extracted using KingFisher Apex Dx Purification System, and real-time PCR was performed on the QS5Dx with one lot of assay reagents.
All targets for both NP and AN specimens in VTM/UTM produced 100% positivity at all the time points tested for the 8°C, 30°C and the ≤-70°C temperatures. The AN specimens also demonstrated 100 % agreement with the expected results for all the targets at all time points for the -30°C to -10°C storage condition. The specimen stability results are summarized in **Tables 11-13** below.
These data support the claimed specimen stability in the Instruction for Use of 24 hours at room temperature (15–25°C), 72 hours refrigerated (2–8°C) and 30 days at ≤-70°C.
**Table 11.** Results for Sample Stability at the 8°C Storage Condition
| Matrix | Storage Condition | Time point | Negative | Influenza A | Influenza B | SARS-CoV-2 | RSV |
| --- | --- | --- | --- | --- | --- | --- | --- |
| | | | % Positive (nPositive/Total) | % Positive (nPositive/Total) | % Positive (nPositive/Total) | % Positive (nPositive/Total) | % Positive (nPositive/Total) |
| NP | 8°C | 0 Hour | 0% (0/2) | 100% (5/5) | 100% (5/5) | 100% (5/5) | 100% (5/5) |
| | | 24 Hours | 0% (0/2) | 100% (5/5) | 100% (5/5) | 100% (5/5) | 100% (5/5) |
| | | 48 Hours | 0% (0/2) | 100% (5/5) | 100% (5/5) | 100% (5/5) | 100% (5/5) |
| | | 72 Hours | 0% (0/2) | 100% (5/5) | 100% (5/5) | 100% (5/5) | 100% (5/5) |
| | | 80 Hours | 0% (0/2) | 100% (5/5) | 100% (14/14)* | 100% (5/5) | 100% (5/5) |
| AN | 8°C | 0 Hour | 0% (0/2) | 100% (5/5) | 100% (5/5) | 100% (5/5) | 100% (5/5) |
| | | 24 Hours | 0% (0/2) | 100% (5/5) | 100% (5/5) | 100% (5/5) | 100% (5/5) |
| | | 48 Hours | 0% (0/2) | 100% (5/5) | 100% (5/5) | 100% (5/5) | 100% (5/5) |
| | | 72 Hours | 0% (0/2) | 100% (5/5) | 100% (5/5) | 100% (5/5) | 100% (5/5) |
| | | 80 Hours | 0% (0/2) | 100% (5/5) | 100% (5/5) | 100% (5/5) | 100% (5/5) |
\*Includes a total of 4 replicates from the original run (1 replicate was excluded) and 10 replicates from Repeat 2.
**Table 12.** Results for Sample Stability at the 30°C Storage Condition
| Matrix | Storage Condition | Time point | Negative | Influenza A | Influenza B | SARS-CoV-2 | RSV |
| --- | --- | --- | --- | --- | --- | --- | --- |
| | | | % Positive (nPositive/Total) | % Positive (nPositive/Total) | % Positive (nPositive/Total) | % Positive (nPositive/Total) | % Positive (nPositive/Total) |
| NP | 30°C | 0 Hour | 0% (0/2) | 100% (5/5) | 100% (5/5) | 100% (5/5) | 100% (5/5) |
| | | 24 Hours | 0% (0/2) | 100% (5/5) | 100% (5/5) | 100% (5/5) | 100% (5/5) |
| | | 27 Hours | 0% (0/2) | 100% (5/5) | 100% (5/5) | 100% (5/5) | 100% (5/5) |
| AN | 30°C | 0 Hour | 0% (0/2) | 100% (5/5) | 100% (5/5) | 100% (5/5) | 100% (5/5) |
| | | 24 Hours | 0% (0/2) | 100% (5/5) | 100% (5/5) | 100% (5/5) | 100% (5/5) |
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| | 27 Hours | 0% (0/2) | 100% (5/5) | 100% (5/5) | 100% (5/5) | 100% (5/5) |
| --- | --- | --- | --- | --- | --- | --- |
**Table 13.** Positivity values for the ≤-70°C storage condition
| Matrix | Storage Condition | Time point | Negative | Influenza A | Influenza B | SARS-CoV-2 | RSV |
| --- | --- | --- | --- | --- | --- | --- | --- |
| | | | % Positive (nPositive/Total) | % Positive (nPositive/Total) | % Positive (nPositive/Total) | % Positive (nPositive/Total) | % Positive (nPositive/Total) |
| NP | ≤-70°C | 0 day | 0% (0/2) | 100% (5/5) | 100% (5/5) | 100% (5/5) | 100% (5/5) |
| | | 15 days | 0% (0/2) | 100% (5/5) | 100% (5/5) | 100% (5/5) | 100% (5/5) |
| | | 30 days | 0% (0/2) | 100% (5/5) | 100% (5/5) | 100% (5/5) | 100% (5/5) |
| | | 33 days | 0% (0/2) | 100% (5/5) | 100% (5/5) | 100% (5/5) | 100% (5/5) |
| AN | ≤-70°C | 0 day | 0% (0/2) | 100% (5/5) | 100% (5/5) | 100% (5/5) | 100% (5/5) |
| | | 15 days | 0% (0/2) | 100% (5/5) | 100% (5/5) | 100% (5/5) | 100% (5/5) |
| | | 30 days | 0% (0/2) | 100% (5/5) | 100% (5/5) | 100% (5/5) | 100% (5/5) |
| | | 33 days | 0% (0/2) | 100% (5/5) | 100% (5/5) | 100% (5/5) | 100% (5/5) |
### c. Extraction Eluate Freeze-thaw Stability
The eluate stability study evaluated the performance of stored and frozen-and-thawed nucleic acid eluates with the TaqPath COVID-19, Flu A, Flu B, RSV Select Panel. NP and AN specimens in VTM/UTM were contrived with one strain individually of SARS-CoV-2 (USA-WA1/2020), influenza A (California/07/09), influenza B (Florida/04/06) and RSV (RSV A, 2013 isolate) to 3x LoD prior to being extracted with the MagMAX Dx Viral/Pathogen Nucleic Acid Isolation Kit on the KingFisher Apex to collect the eluates. Fifteen replicates each of NP and AN specimen eluates were either tested fresh or tested after storage at 2°C to 8°C for up to 27 hours followed by storage at ≤ -70°C for more than 6 days and undergoing two (2) freeze/thaw cycles. Stability was determined from three (3) time points. Testing was performed with one lot of the TaqPath COVID-19, Flu A, Flu B, RSV Select Panel on the QS5 Dx instrument.
100% of contrived positive samples produced the expected results at all three (3) time points evaluated. The results demonstrate that extracted sample eluates from contrived NP and AN swab samples containing SARS-CoV-2, influenza A, influenza B or RSV were stable when held for 27 hours at 8°C followed by more than 6 days at ≤ -70°C while undergoing two (2) freeze/thaw cycles. Therefore, the results support eluates storage claims at 2°C to 8°C for up to 24 hours, storage at ≤ -70°C for up to 5 days and/or up to one (1) freeze/thaw cycle.
### d. Fresh vs Frozen Specimen Study
Fresh vs. frozen equivalency was assessed by testing contrived positive samples consisting of one representative strain of influenza A (H1N1pdm, Michigan/45/15), influenza B (Yamagata, Florida/04/06), SARS-CoV-2 (USA-WA1/2020) and RSV (RSVB, CH93(18)-18) at 2x and 5x their respective LoDs. Contrived samples were prepared in pooled, negative NP and AN specimens in VTM/UTM, respectively. Each sample for each condition was tested with twenty replicates (N=20) for the 2x LoD concentration and with ten replicates (N=10) for the 5x LoD concentration through the
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full TaqPath COVID-19, Flu A, Flu B, RSV Select Panel workflow. Testing was performed on freshly contrived samples and samples that had undergone two (2), three (3) and six (6) freeze/thaw cycles with one lot of reagents on the QS5Dx instrument. Additionally, negative samples were also tested for all the conditions with five replicates (N=5). The results of this study support the use of frozen specimens in the clinical performance study and the claim in the TaqPath COVID-19, Flu A, Flu B, RSV Select Panel Instructions for Use that specimens collected in Copan UTM Universal Transport Medium or BD Universal Viral Transport (UVT) Medium may undergo up to 2 freeze/thaw cycles. Do not exceed 1 freeze/thaw cycle for specimens collected in other transport media types.
# 6. Detection Limit:
The Limit of Detection (LoD) of the TaqPath COVID-19, Flu A, Flu B, RSV Select Panel was determined for two (2) strains of influenza A H1N1, two (2) strains of influenza A H3N2, one (1) strain of influenza B of Victoria lineage, two (2) strains of influenza B of Yamagata lineage, one (1) strain of SARS-CoV- 2, one (1) strain of RSV A and one (1) strain of RSV B. Each virus was spiked into pooled, negative nasopharyngeal (NP) swab specimens in VTM/UTM, extracted on the KingFisher Apex Dx Purification System, then PCR was performed on the QS5Dx instrument across three (3) days, with two (2) reagent lots. LoDs for all viruses except the influenza B Victoria lineage strain were also determined in anterior nasal (AN) swab specimens in VTM/UTM.
The preliminary LoD was determined by probit analysis separately for each virus in each specimen type with each reagent lot, and the highest (i.e., worst-case) LoD value across reagent lots was selected as the preliminary LoD for that virus in the respective specimen type.
The preliminary LoDs determined by probit analysis were confirmed by testing at least 24 replicates at, above, and below the preliminary LoDs. Contrived specimens were prepared with negative clinical matrix (NP and AN swab specimens in VTM/UTM) and tested on the QS5Dx instrument. The LoD for the TaqPath COVID-19, Flu A, Flu B, RSV Select Panel is shown in **Table 14** below.
**Table 14.** Limit of Detection for the TaqPath COVID-19, Flu A, Flu B, RSV Select Panel
| Virus | Subtype or Lineage | Strain | LoD in GCE/mL | LoD in TCID_{50}/mL |
| --- | --- | --- | --- | --- |
| **SARS-CoV-2** | **N/A** | **USA-WA1/2020** | 107 GCE/mL | 0.158 TCID_{50}/mL |
| **Influenza A** | **H1N1** | **Michigan/45/15** | 340 GCE/mL | 0.045 TCID_{50}/mL |
| | | **California/07/09** | 282 GCE/mL | 0.026 TCID_{50}/mL |
| | **H3N2** | **South Australia/55/14** | 281 GCE/mL | 0.034 TCID_{50}/mL |
| | | **Texas/50/12** | 268 GCE/mL | 0.079 TCID_{50}/mL |
| **Influenza B** | **Yamagata** | **Victoria/504/00** | 1405 GCE/mL | 0.120 TCID_{50}/mL |
| | | **Florida/04/06** | 464 GCE/mL | 0.048 TCID_{50}/mL |
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| | Victoria | Malaysia/2506/2004 | 3500 GCE/mL | 0.075 TCID_{50}/mL |
| --- | --- | --- | --- | --- |
| RSV | A | 2013 Isolate | 1614 GCE/mL | 0.013 TCID_{50}/mL |
| | B | CH93(18)-18 | 1478 GCE/mL | 0.046 TCID_{50}/mL |
### a. Limit of Detection with WHO Standard
The study was performed using two-phase approach to determine the TaqPath COVID-19, Flu A, Flu B, RSV Select Panel LoD using the WHO Standard on the QS5Dx instrument. Contrived specimens were formulated with the First WHO International Standard for SARS-CoV-2 RNA spiked into pooled, negative anterior nasal (AN) swab specimens in VTM/UTM at various concentrations.
Once formulated, contrived specimens were extracted and detected using the TaqPath COVID-19, Flu A, Flu B, RSV Select Panel in Phase I (range finding). The results of Phase I were used to establish the preliminary LoD (which was the lowest concentration to generate 100% positivity when tested in triplicate). In Phase II, the preliminary LoD was confirmed as the lowest concentration that results in at least 95% positive detection (≥95% positive) when tested with 20 replicates. Phase II testing was performed in a blinded format.
The results of Phase I testing produced a preliminary LoD of 5.012 × 10¹ IU/mL, which was the lowest (most dilute) concentration that yielded 100% positivity. In Phase II, the LoD was confirmed at 150.36 IU/mL as the lowest concentration that resulted in ≥ 95% positive detection as summarized in Table 15 below.
Table 15. Limit of Detection with SARS-CoV-2 WHO Standard
| Concentration | Positive / Total Replicates (% Positive) |
| --- | --- |
| 451.08 IU/mL (9x Preliminary LoD) | 20 / 20 (100%) |
| 150.36 IU/mL (3x Preliminary LoD) | 20 / 20 (100%) |
| 50.12 IU/mL (Preliminary LoD) | 18 / 20 (90%) |
| 16.71 IU/mL (0.33x Preliminary LoD) | 12 / 20 (60%) |
| Negative | 0 / 3 (0%) |
### 7. Analytical Reactivity (Inclusivity)
#### a. In-silico Inclusivity
Table 16 summarizes the genome sequences for SARS-CoV-2, influenza A virus, influenza B virus and respiratory syncytial virus that were downloaded from GISAID and GenBank on the indicated dates and analyzed for identity and melting temperature (Tm). Genomes with 100% identity and/or Tm greater than annealing temperature (Ta) for at least one assay were designated as reactive. For influenza A, wet testing with a predominant mismatch demonstrated no significant difference in detection between the mismatched sequence and reference sequence, therefore genomes with that mismatch were also predicted to be reactive.
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**Table 16.** *In silico* Inclusivity (Reactivity) for the TaqPath COVID-19, Flu A, Flu B, RSV Select Panel
| Virus | Specimen collection date range | Sequences with 100% identity to ≥ 1 assay | Sequences with Tm > Ta for ≥ 1 assay | Predicted inclusivity |
| --- | --- | --- | --- | --- |
| SARS-CoV-2 | All through 25 March 2024 (GISAID and GenBank) | >99.99% (11,485,741/11,486,294) | >99.99% (11,486,283/11,486,294) | >99.9% |
| Influenza A | 1 January 2009 through 25 March 2024 (GenBank) and 23 April 2024 (GISAID) | 83.76% (110,802/132,283) | 89.6% (118,481/132,283) | 95.27%^{[1]} |
| Influenza B | 1 January 2009 through 25 March 2024 (GenBank) and 23 April 2024 (GISAID) | 98.56% (31,480/31,939) | >99.9% (31,935/31,939) | >99.9% |
| RSV | All through 25 March 2024 (GISAID and GenBank) | 87.07% (8,460/9,716) | 99.49% (9,666/9,716) | 99.49% |
$^{[1]}$ Mismatch substitution was wet tested and showed no significant reduction in sensitivity compared with reference. Including strains with this mismatch in the counts produces a predicted inclusivity of 95.27%.
#### b. Inclusivity Wet-Testing
Reactivity (inclusivity) was determined for eleven (11) strains of influenza A (H1N1), twelve (12) strains of influenza A (H3N2), five (5) strains of influenza B (two (2) of Victoria lineage and three (3) of Yamagata lineage), six (6) strains of RSV (three (3) each of RSV A and B), five (5) strains of SARS-CoV-2 and five (5) avian influenza strains (one (1) each of H5N1, H5N2, H7N2, H7N7 and H7N9). Each virus was introduced at near-LoD levels into negative, pooled NP swab specimens in VTM/UTM, then extracted and detected with the TaqPath COVID-19, Flu A, Flu B, RSV Select Panel. Sample extraction was performed using the KingFisher Apex, and real-time RT-PCR was performed with the QS5Dx using one (1) lot of TaqPath COVID-19, Flu A, Flu B, RSV Select Panel reagents. Each sample was formulated with live virus, inactivated virus or genomic RNA, depending upon availability and tested in triplicate (N=3). Virus strains that did not produce 100% detection at approximately 3x LoD were further tested at progressively higher concentrations until 100% detection was achieved. The results are summarized in **Table 17**.
**Table 17.** Reactivity (Inclusivity) Study Results
| Strain | Positive Replicates / Total (% Positivity) | Concentration | |
| --- | --- | --- | --- |
| | | GCE/mL | TCID_{50}/mL |
| Influenza A H1N1 (Georgia/M5081/2012) | 3 / 3 (100%) | 1020 | 42.83 |
| Influenza A H1N1 (New Caledonia/20/99) | 3 / 3 (100%) | 1020 | 0.44 |
| Influenza A H1N1 (NY/03/09)^{[1],[2]} | 3 / 3 (100%) | 4080 | 6.88 |
| Influenza A H1N1 (Singapore/63/04) | 3 / 3 (100%) | 1020 | 0.05 |
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| Influenza A H1N1 (Brisbane/02/2018)^{[2]} | 3 / 3 (100%) | 1020 | 0.11 |
| --- | --- | --- | --- |
| Influenza A H1N1 (Solomon Islands/3/2006) | 3 / 3 (100%) | 1020 | 1.67 |
| Influenza A H1N1 (Puerto Rico/08/1934) | 3 / 3 (100%) | 1020 | 0.11 |
| Influenza A H1N1 (Mexico/4108/09)^{[2]} | 3 / 3 (100%) | 1020 | 7.75 |
| Influenza A H1N1 (Taiwan/42/06) | 3 / 3 (100%) | 1020 | 0.99 |
| Influenza A H1N1 (Brisbane/59/07) | 3 / 3 (100%) | 1020 | 0.07 |
| Influenza A H1N1 (Victoria/2570/19)^{[2]} | 3 / 3 (100%) | 1020 | 0.03 |
| Influenza A H3N2 (Wisconsin/67/2005) | 3 / 3 (100%) | 1278 | 0.20 |
| Influenza A H3N2 (Switzerland/9715293/13) | 3 / 3 (100%) | 1278 | 0.007 |
| Influenza A H3N2 (Kansas/14/2017) | 3 / 3 (100%) | 1278 | 0.69 |
| Influenza A H3N2 (Singapore/INFIMH-16-0019/16) | 3 / 3 (100%) | 1278 | 0.57 |
| Influenza A H3N2 (Perth/16/09) | 3 / 3 (100%) | 1278 | 0.07 |
| Influenza A H3N2 (Victoria/361/2011) | 3 / 3 (100%) | 1278 | 0.02 |
| Influenza A H3N2 (Hong Kong/8/68) | 3 / 3 (100%) | 1278 | 0.12 |
| Influenza A H3N2 (Brisbane/10/07) | 3 / 3 (100%) | 1278 | 0.20 |
| Influenza A H3N2 (California/7/04) | 3 / 3 (100%) | 1278 | 0.72 |
| Influenza A H3N2 (Hong Kong/4801/14) | 3 / 3 (100%) | 1278 | 2.32 |
| Influenza A H3N2 (Hong Kong/2671/19) | 3 / 3 (100%) | 1280 | 1.07 |
| Influenza A H3N2 (Macha/O1237/2021) | 3 / 3 (100%) | 1280 | 2.62 |
| Influenza B Victoria (Colorado/06/2017) | 3 / 3 (100%) | 4215^{[4]} | 0.08 |
| Influenza B Victoria (Victoria/2/87) | 3 / 3 (100%) | 4215^{[4]} | 1.74 |
| Influenza B Yamagata (Massachusetts/02/2012) | 3 / 3 (100%) | 1392 | 0.02 |
| Influenza B Yamagata (Phuket/3073/13) | 3 / 3 (100%) | 1392 | 0.004 |
| Influenza B Yamagata (Wisconsin/01/2010) | 3 / 3 (100%) | 1392 | 0.06 |
| RSV A (Long) | 3 / 3 (100%) | 4842 | 4.56 |
| RSV A (2014 Isolate 342) | 3 / 3 (100%) | 4842 | 0.26 |
| RSV A (2006 isolate) | 3 / 3 (100%) | 4842 | 0.05 |
| RSV B (9320)^{[5]} | 3 / 3 (100%) | 4434 | Unknown^{[3]} |
| RSV B (3/2015 Isolate #2) | 3 / 3 (100%) | 4434 | 2.07 |
| RSV B (12/2014 Isolate #1) | 3 / 3 (100%) | 4434 | 0.56 |
| SARS-CoV-2 (Alpha Variant (B.1.1.7) England/204820464/2020)^{[5]} | 3 / 3 (100%) | 321 | 0.26 |
| SARS-CoV-2 (Beta (B.1.351) South Africa/KRISP- K005325/2020)^{[5]} | 3 / 3 (100%) | 321 | 0.09 |
| SARS-CoV-2 (Gamma (P1) Japan/TY7- | 3 / 3 (100%) | 321 | 0.03 |
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| 503/2021)^{[5]} | | | |
| --- | --- | --- | --- |
| SARS-CoV-2 (Delta (B.1.617.2) USA/PHC658/2021)^{[5]} | 3 / 3 (100%) | 321 | 0.02 |
| SARS-CoV-2 (Omicron (BA.2.3) USA/MD-HP24556/2022)^{[5]} | 3 / 3 (100%) | 321 | 0.02 |
| Avian influenza H5N1 (India/NIV/2006)^{[5]} | 3 / 3 (100%) | 1278 | Unknown^{[3]} |
| Avian influenza H5N2 (pheasant/New Jersey/1355/1998)^{[5]} | 3 / 3 (100%) | 1278 | Unknown^{[3]} |
| Avian influenza H7N2 (turkey/Virginia/4529/2002)^{[5]} | 3 / 3 (100%) | 1278 | Unknown^{[3]} |
| Avian influenza H7N7 (mallard/Netherlands/12/2000)^{[5]} | 3 / 3 (100%) | 1278 | Unknown^{[3]} |
| Avian influenza H7N9 (Anhui/1/2013)^{[5]} | 3 / 3 (100%) | 1278 | Unknown^{[3]} |
[1] Reactive with the TaqPath COVID-19, Flu A, Flu B, RSV Select Panel at 12x LoD.
[2] H1N1pdm09 strain.
[3] TCID$_{50}$/mL concentrations unknown (not reported by supplier).
[4] 1.2x LoD relative to LoD of 3500 GCE/mL determined for influenza B Victoria lineage strain Malaysia/2506/2004.
[5] Viral genomic RNA or inactivated virus.
## 8. RNase P Internal Control Cutoff Confirmation
In the RNase P Internal Control Cutoff Confirmation study, viral target-negative “used” (i.e., collected from individual donors and containing RNase P) nasopharyngeal (NP) swab specimens and “unused” (i.e., unopened and not containing RNase P) NP swabs were tested according to the TaqPath COVID-19, Flu A, Flu B, RSV Select Panel Instructions for Use. The IC cutoff was confirmed at Cq = 33.0 while testing individually collected NP swabs. The results demonstrated an overall positive percent agreement (PPA) of 98.07% (95% CI of 97.31-98.63%) and negative percent agreement (NPA) of 100% (95% CI of 97.49-100%) for RNase P detection in NP swabs. For AN swabs, PPA for RNase P detection was 92.30% (95% Score CI of 90.83-93.54%) and NPA was 100% (95% Score CI of 97.50-100%).
Due to the lower than expected PPA observed for AN specimens in the RNase P Cutoff confirmation study, an additional study was conducted utilizing an improved AN swab collection technique. All of the evaluated samples produced valid calls. Two (2) out of the one hundred thirty-eight (138) AN samples were positive for COVID-19, while the rest were negative for all viral targets. RNase P was detected in 100% of both negative and positive samples. The results are summarized in **Table 18**.
**Table 18.** Invalid Rate in AN Specimen Swab Collection Study Results
| Sample Call | RNase P Call | Number of Samples | Percent Validity | RNase P (Cq) | | |
| --- | --- | --- | --- | --- | --- | --- |
| | | | | Mean | Median | SD |
| Negative | Positive | 136 | 100% | 27.1 | 26.7 | 2.1 |
| SARS-CoV-2 Positive | Positive | 2 | 100% | 26.9 | 26.9 | 1.9 |
| All | Positive | 138 | 100% | 27.1 | 26.7 | 2.1 |
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This study has demonstrated that the improved AN swab collection technique reduces the invalid rate for AN swabs when tested with the TaqPath COVID-19, Flu A, Flu B, RSV Select Panel. The TaqPath COVID-19, Flu A, Flu B, RSV Select Panel Instructions for Use include the improved AN swab collection instructions.
# 9. Carry-Over:
The study was conducted using negative and contrived specimens of pooled nasopharyngeal (NP) swabs collected in VTM/UTM in a modified checkerboard pattern so that 12 positive samples per extraction event were included amongst 82 negative samples. Extraction was completed using contrived specimens with high-titer inactivated SARS-CoV-2 viral material and downstream real-time PCR with the TaqPath COVID-19, Flu A, Flu B, RSV Select Panel assay and workflow. The entire workflow was evaluated from sample extraction to real-time PCR results.
Six plates containing 12 high-positive samples (SARS-CoV-2 at 1 x 10⁵ PFU/mL in pooled NP specimens in VTM/UTM) surrounded by negative NP samples were extracted by two operators on two different KingFisher Apex Dx Purification System instruments and tested sequentially with the TaqPath COVID-19, Flu A, Flu B, RSV Select Panel.
None of the 492 negative sample replicates tested produced a positive result for SARS-CoV-2, demonstrating a carryover/cross-contamination rate of 0%.
# **B Comparison Studies:**
# 1. Method Comparison with Predicate Device:
Refer to Clinical Studies Section of this document.
# 2. Matrix Comparison:
This study was conducted to establish matrix equivalency for the TaqPath COVID-19, Flu A, Flu B, RSV Select Panel among nasopharyngeal (NP) and anterior nasal (AN) swab specimens collected in VTM/UTM.
Contrived samples prepared with one strain each of SARS-CoV-2, influenza A, influenza B and RSV. Forty (40) replicates at 5x LoD, 20 replicates at 2x LoD and 20 negative replicates were tested in NP and AN sample matrices with one lot of the TaqPath COVID-19, Flu A, Flu B, RSV Select Panel reagents on the QS5Dx instrument. Matrix equivalency was demonstrated with a percent positivity of 100% at 5x LoD and 2x LoD for both matrices. All negative replicates gave negative results, producing a 0% rate of positivity (**Table 19**).
**Table 19.** Matrix Equivalency Study Result Summary
| Virus | Virus Strain | Condition | Concentration (GCE/mL) | Matrix | # Positive/# Tested (% Positivity) |
| --- | --- | --- | --- | --- | --- |
| SARS-CoV-2 | USA-WA1/2020 | 5x LoD | 535 | NP | 20/20 (100%) |
| | | | | AN | 20/20 (100%) |
| | | 2x LoD | 214 | NP | 40/40 (100%) |
| | | | | AN | 40/40 (100%) |
| Influenza A H1N1 | California/07/09 | 5x LoD | 1410 | NP | 20/20 (100%) |
| | | | | AN | 20/20 (100%) |
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| | | 2x LoD | 564 | NP | 40/40 (100%) |
| --- | --- | --- | --- | --- | --- |
| | | | | AN | 40/40 (100%) |
| Influenza B | Florida/04/06 | 5x LoD | 2320 | NP | 20/20 (100%) |
| | | | | AN | 20/20 (100%) |
| | | 2x LoD | 928 | NP | 40/40 (100%) |
| | | | | AN | 40/40 (100%) |
| RSVA | 2013 Isolate Culture Fluid | 5x LoD | 8070 | NP | 20/20 (100%) |
| | | | | AN | 20/20 (100%) |
| | | 2x LoD | 3228 | NP | 40/40 (100%) |
| | | | | AN | 40/40 (100%) |
| Negative | N/A | 0x LoD | 0 | NP | 0/20 (0%) |
| | | | | AN | 0/20 (0%) |
## C Clinical Studies:
### 1. Prospective Clinical Study
One NP swab and one AN swab was prospectively collected from each subject enrolled in the study from 14 diverse sites in the US. NP and AN swabs were collected in UTM, and the collection order (first specimen collected) was alternated between NP and AN swabs. Specimens were collected by trained healthcare professionals between February 20, 2023 and February 29, 2024. AN swabs were collected by swabbing both nares with the same swab, and NP swabs were collected from either of the two nostrils.
One thousand eight hundred and forty (1,840) subjects who were experiencing signs and symptoms of respiratory viral infection were enrolled into prospective collection of this study. Data collected from one duplicate enrollment was excluded from the study and two additional subjects withdrew from the study before samples could be collected. AN and NP samples were collected from each of the remaining 1,837 subjects. In addition, 54 AN swabs were excluded due to collection site protocol deviations, four AN swabs were excluded due to testing site protocol deviations, 132 AN swabs were excluded due to a contamination event at one of the testing sites, one AN swab was excluded due to invalid comparator test result, 105 AN swabs were excluded due to invalid TaqPath COVID-19, Flu A, Flu B, RSV Select Panel results. These exclusions resulted in 1,541 AN swabs that were included in final performance analysis, of which 1,192 specimens were tested fresh (Category I specimens) with the TaqPath COVID-19, Flu A, Flu B, RSV Select Panel and 349 were tested after freezing (Category II specimens). Two (2) of the 1,541 AN specimens observed inconclusive comparator results for influenza A, leaving 1,539 evaluable AN specimens for influenza A target. Of the 1,840 subjects enrolled in the prospective study, 1,647 had their AN swab specimens evaluated with the candidate device. One hundred seventy nine (179) of these specimens were invalid by the candidate device during testing, for an initial invalid rate of 10.9% (179/1,647). Upon retesting, the invalid rate for AN specimens decreased to 6.4% (105/1,647).
Of the 1,837 NP swabs that were collected, 55 were excluded due to collection site protocol deviations, three due to testing site protocol deviations, 132 due to contamination event at one of the testing sites and 27 due to invalid TaqPath COVID-19, Flu A, Flu B, RSV Select Panel results. A total of 1,620 NP swabs were included in final performance analysis, of which 1,246 specimens were tested fresh (Category I specimens) with the TaqPath COVID-19, Flu A, Flu B, RSV Select Panel and 374 were tested after freezing (Category II
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specimens). Four (4) of the 1,620 NP specimens observed inconclusive comparator results for Flu A, leaving 1,616 evaluable NP specimens for Flu A. Of the 1,840 subjects enrolled in the prospective study, 1,647 had their NP swab specimens evaluated with the candidate device. Forty five (45) of these specimens were invalid by the candidate device during testing, for an initial invalid rate of 2.7% (45/1,647). Upon retesting, the invalid rate for NP specimens decreased to 1.6% (27/1,647).
Demographics of subjects included in the prospective study that were included in the final performance analysis are summarized in **Table 20**.
**Table 20.** Demographics of Subjects from Prospective Cohort Study Included in the Final Performance Analysis
| Category | AN | AN% | NP | NP% |
| --- | --- | --- | --- | --- |
| **Sex** | | | | |
| Female | 882 | 57.2% | 924 | 57.0% |
| Male | 655 | 42.5% | 692 | 42.7% |
| Prefer Not to Answer | 4 | 0.3% | 4 | 0.3% |
| **Ethnicity** | | | | |
| Hispanic or Latino | 363 | 23.6% | 370 | 22.8% |
| Not Hispanic or Latino | 1133 | 73.5% | 1204 | 74.3% |
| Prefer not to answer | 45 | 2.9% | 46 | 2.8% |
| **Race** | | | | |
| American Indian or Alaska Native | 16 | 1.0% | 16 | 1.0% |
| Asian | 38 | 2.4% | 40 | 2.4% |
| Black or African American | 434 | 28.2% | 470 | 29.0% |
| Native Hawaiian or Other Pacific Islander | 10 | 0.7% | 12 | 0.7% |
| Other | 23 | 1.5% | 24 | 1.5% |
| Prefer not to answer | 59 | 3.8% | 62 | 3.8% |
| White | 961 | 62.4% | 996 | 61.5% |
| **Age (yrs.)** | | | | |
| Mean | 39.6 | N/A | 40.0 | N/A |
| Std Dev | 19.9 | N/A | 19.9 | N/A |
| Min | 0 | N/A | 0 | N/A |
| Max |…
