DASH® SARS-CoV-2 & Flu A/B Test

K241652 · Nuclein, LLC · QOF · Dec 20, 2024 · Microbiology

Device Facts

• Record ID: K241652
• Device Name: DASH® SARS-CoV-2 & Flu A/B Test
• Applicant: Nuclein, LLC
• Product Code: QOF · Microbiology
• Decision Date: Dec 20, 2024
• Decision: SESE
• Submission Type: Dual Track
• Regulation: 21 CFR 866.3981
• Device Class: Class 2
• Attributes: Pediatric

Indications for Use

The DASH SARS-CoV-2 & Flu A/B Test is a rapid reverse transcription polymerase chain reaction (RT-PCR) assay performed on the DASH Rapid PCR Instrument and is intended for the simultaneous in vitro qualitative detection and differentiation of SARS-CoV-2, influenza A and influenza B virus ribonucleic acid (RNA) in anterior nasal swab specimens from patients with signs and symptoms of respiratory tract infection. The test is intended to aid in the differential diagnosis of SARS-CoV-2, influenza A, and influenza B in humans in conjunction with other clinical, epidemiologic and laboratory findings. Positive results of a specific target are indicative of the presence of that viral RNA and may not be the definite cause of disease. Positive results do not rule out co-infection with other pathogens. Negative results do not preclude SARS-CoV-2, influenza A or influenza B infection and should not be used as the sole basis for patient management decisions

Device Story

The DASH SARS-CoV-2 & Flu A/B Test is a rapid, point-of-care RT-PCR assay for qualitative detection of SARS-CoV-2, Flu A, and Flu B. The system consists of a single-use test cartridge and the DASH Rapid PCR Instrument. The user inserts an anterior nasal swab into the cartridge containing lysis buffer; the cartridge is then sealed and inserted into the instrument. The instrument performs automated sample preparation via sequence-specific capture using biotinylated oligomers and streptavidin-coated paramagnetic particles, followed by RT-PCR amplification (40 cycles). Fluorescence is monitored via a four-channel LED fluorometer. The device provides qualitative results to aid in differential diagnosis. It is intended for use in CLIA-waived settings by untrained operators. The output assists clinicians in managing patients with respiratory symptoms by identifying viral targets, though it does not rule out co-infections or other pathogens.

Clinical Evidence

Clinical performance was evaluated in a prospective study (N=795) at seven US CLIA-waived sites during the 2023-2024 respiratory season. Untrained operators performed testing. Compared to FDA-cleared RT-PCR comparators, the DASH test demonstrated: SARS-CoV-2 PPA 95.2% (90.9-97.6%), NPA 99.5% (98.6-99.8%); Flu A PPA 94.3% (84.6-98.1%), NPA 98.1% (96.8-98.9%); Flu B PPA 97.3% (86.2-99.9%), NPA 99.2% (98.3-99.6%). Invalid rate was 2.9%.

Technological Characteristics

The system uses sequence-specific capture with biotinylated oligomers and streptavidin-coated paramagnetic particles for RNA isolation, followed by RT-PCR amplification. Reagents are lyophilized in a single-use cartridge. The instrument features a four-channel LED fluorometer for real-time detection. It is a standalone, point-of-care system. Safety/EMC compliance: IEC 60601-1-2, IEC 61010-1, IEC 61010-2-010, IEC 61010-2-101.

Indications for Use

Indicated for patients with signs and symptoms of respiratory tract infection. Specimens are anterior nasal swabs collected by healthcare providers, adults (for children <14 years), or self-collected (aged ≥14 years).

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 SPOTFIRE Respiratory Panel Mini (K230719)

Submission Summary (Full Text)

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FDA

U.S. FOOD &amp; DRUG

ADMINISTRATION

# 510(k) SUBSTANTIAL EQUIVALENCE DETERMINATION DECISION SUMMARY

ASSAY AND INSTRUMENT

## I Background Information:

A 510(k) Number

K241652

B Applicant

Nuclein, LLC

C Proprietary and Established Names

DASH SARS-CoV-2 &amp; Flu A/B Test

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:

To obtain market clearance for the DASH SARS-CoV-2 &amp; Flu A/B Test.

B Measurand:

SARS-CoV-2 RNA

Influenza A RNA

Influenza B RNA

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 DASH SARS-CoV-2 &amp; Flu A/B Test is a 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 DASH SARS-CoV-2 &amp; Flu A/B Test is a rapid reverse transcription polymerase chain reaction (RT-PCR) assay performed on the DASH Rapid PCR Instrument and is intended for the simultaneous in vitro qualitative detection and differentiation of SARS-CoV-2, influenza A and influenza B virus ribonucleic acid (RNA) in anterior nasal swab specimens from patients with signs and symptoms of respiratory tract infection. The test is intended to aid in the differential diagnosis of SARS-CoV-2, influenza A, and influenza B in humans in conjunction with other clinical, epidemiologic and laboratory findings.

Positive results of a specific target are indicative of the presence of that viral RNA and may not be the definite cause of disease. Positive results do not rule out co-infection with other pathogens. Negative results do not preclude SARS-CoV-2, influenza A or influenza B infection and should not be used as the sole basis for patient management decisions

C Special Conditions for Use Statement(s):

Rx - For Prescription Use Only

IVD - For In Vitro Diagnostic Use Only

D Special Instrument Requirements:

DASH Rapid PCR Instrument (DASH Instrument)

IV Device/System Characteristics:

A Device Description:

The DASH SARS-CoV-2 &amp; Flu A/B Test is a reverse transcription polymerase chain reaction (RT-PCR) assay for rapid qualitative detection and differentiation of SARS-CoV-2, influenza A (Flu A) and influenza B (Flu B) from human anterior nasal swab (ANS) specimens. The test combines the technologies of sequence specific capture sample preparation and RT-PCR amplification. An anterior nasal swab with a 30-mm breakpoint is used to collect a specimen. The ANS specimen is added directly to the DASH SARS-CoV-2 &amp; Flu A/B Test cartridge sample chamber. The DASH SARS-CoV-2 &amp; Flu A/B Test cartridge contains all reagents necessary to perform the test. The cartridge is capped and inserted into the DASH Instrument to initiate the test, and all subsequent test steps are performed automatically by the DASH Instrument.

Materials included with the DASH SARS-CoV-2 &amp; Flu A/B Test consist of the following:

- Ten individually pouched DASH SARS-CoV-2 &amp; Flu A/B Cartridges per kit

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- Ten individually packaged anterior nares sample collection swabs (30 mm breakpoint, sterile elongated polyester flock) per kit
- DASH SARS-CoV-2 &amp; Flu A/B Test Quick Reference Instructions (QRI).

The DASH Instrument, DASH SARS-CoV-2 &amp; Flu A/B Positive Control Swabs Kit, and DASH Negative Control Swabs Kit are provided separately.

## B Principle of Operation:

The DASH SARS-CoV-2 &amp; Flu A/B Test combines the technologies of sequence specific capture sample preparation and RT-PCR amplification. The DASH SARS-CoV-2 &amp; Flu A/B Test cartridge contains all reagents necessary to perform the test to detect RNA from SARS-CoV-2, influenza A and influenza B.

An ANS sample is collected and added directly to the DASH SARS-CoV-2 &amp; Flu A/B Test cartridge. The Test cartridge is capped and inserted into the DASH Instrument to initiate the test. The Test cartridge sample chamber contains lysis buffer which releases the RNA target from the virus. The target RNA molecules are isolated from specimens by use of biotinylated capture oligomers (capture oligomers) which are in turn captured by streptavidin-coated paramagnetic particles. The biotinylated capture oligomers contain sequences complementary to specific regions of the RNA target molecules. During the hybridization step, the sequence specific regions of the capture oligomers bind to specific regions of the RNA target molecules. The capture oligomer:target complex is then concentrated by the addition of streptavidin-coated paramagnetic particles.

The paramagnetic particles, including the bound target molecules, are washed to remove residual specimen matrix that may contain amplification reaction inhibitors. After the target capture steps are completed, the target RNA is ready for RT-PCR amplification. Forty amplification cycles are performed, during which the fluorescence is measured with a four-color LED fluorometer. Amplification of any assay targets, Flu A, Flu B, and SARS-CoV-2, leads to fluorescent signal. An algorithm determines whether the result is positive or negative for each viral target assay. Additionally, a process control is included in the cartridge to control for adequate processing of the target virus through the steps of sample purification, nucleic acid amplification and to monitor for the presence of inhibitors in the RT-PCR reaction. Total run time, from swab insertion into the Test cartridge to display of the test results is approximately 15 minutes.

The DASH Instrument contains firmware and software that operate the instrument and analyze the assay data. Test results are displayed on the DASH Instrument screen and can be printed (optional) after the test has been completed. The procedural control will display "Valid" on the results screen for all samples, both negative and positive (see Table 1 below), to indicate that the test results may be reported.

Table 1. DASH SARS CoV-2 &amp; Flu A/B Results Interpretation

| Target | Result | Interpretation |
| --- | --- | --- |
| SARS-CoV-2 | Positive | The target nucleic acids of SARS-CoV-2 are detected. |
| | Negative | The target nucleic acids of SARS-CoV-2 are not detected. |
| | Positive | The target nucleic acids of Flu A are detected. |

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| Flu A | Negative | The target nucleic acids of Flu A are not detected. |
| --- | --- | --- |
| Flu B | Positive | The target nucleic acids of Flu B are detected. |
| | Negative | The target nucleic acids of Flu B are not detected. |
| Procedural Control | Valid | Amplification meets acceptance criteria. Test results may be reported. |
| | Invalid | Presence or absence of the target nucleic acids cannot be determined. Repeat test with a new cartridge. |
| Error | | Presence or absence of the target nucleic acids cannot be determined. Instrument detected an error and aborted the current test.
Repeat test with a new cartridge. |

## C Instrument Description Information:

1. **Instrument Name:**
DASH Rapid PCR Instrument (DASH Instrument)

2. **Specimen Identification:**
DASH Instrument is connected to a barcode reader which scans specimen and Test cartridge labels.

3. **Specimen Sampling and Handling:**
The anterior nasal swab specimen is collected and added directly to the DASH SARS-CoV-2 &amp; Flu A/B Test cartridge sample chamber. No collection media is used to store the swab. Each sample is processed individually.

4. **Calibration:**
Not applicable.

5. **Quality Control:**

- **Procedural Control**
An assay targeting the bacteriophage MS2 is built-in the Test cartridge as the Procedural Control (PRC) that undergoes identical processing to the patient specimen which provides confirmation that the test procedure was executed correctly. If the test fails for any reason, the PRC result will be reported as an INVALID and the operator is instructed in the labelling to test the sample again with another test device.

- **External Controls**
The DASH SARS-CoV-2 &amp; Flu A/B Positive Control and DASH Negative Control swabs are available separately for use as external controls. External controls are tested using the same procedure that is used to test a patient specimen.

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The DASH SARS-CoV-2 &amp; Flu A/B Positive Control Swabs Kit is comprised of five single use, individually packaged swabs with 30 mm breakpoint, containing a non-infectious solution of Flu A, Flu B, and SARS-CoV-2 viruses. The DASH Negative Control Swabs Kit is comprised of five single-use, individually packaged swabs with 30 mm breakpoint that are void of any added media.

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V Substantial Equivalence Information:

A Predicate Device Name(s):
BIOFIRE SPOTFIRE Respiratory (R) Panel Mini

B Predicate 510(k) Number(s):
K230719

C Comparison with Predicate(s):

| Device & Predicate Device(s): | K241652 | K230719 |
| --- | --- | --- |
| Device Trade Name | DASH SARS-CoV-2 & Flu A/B Test | BIOFIRE SPOTFIRE Respiratory (R) Panel Mini |
| General Device Characteristic Similarities | | |
| Regulation Number and Name | Same | 21 CFR 866.3981; Devices to detect and identify nucleic acid targets in respiratory samples from microbial agents that cause the SARS-CoV-2 respiratory infection and other microbial agents when in a multi-analyte test. |
| Product Code | Same | QOF |
| Technology/Detection | PCR with streptavidin-coated paramagnetic particles and fluorescence detection | PCR with DNA melting analysis |
| Intended Use/Indications For Use | The DASH SARS-CoV-2 & Flu A/B Test is a rapid reverse transcription polymerase chain reaction (RT-PCR) assay performed on the DASH Rapid PCR Instrument and is | The BIOFIRE SPOTFIRE Respiratory Panel Mini (SPOTFIRE R Panel Mini) is a multiplexed polymerase chain reaction (PCR) test intended for use with the BIOFIRE SPOTFIRE System for |

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| | intended for the simultaneous in vitro qualitative detection and differentiation of SARS-CoV-2, influenza A and influenza B virus ribonucleic acid (RNA) in anterior nasal swab specimens from patients with signs and symptoms of respiratory tract infection. The test is intended to aid in the differential diagnosis of SARS-CoV-2, influenza A, and influenza B in humans in conjunction with other clinical, epidemiologic and laboratory findings.

Positive results of a specific target are indicative of the presence of that viral RNA and may not be the definite cause of disease. Positive results do not rule out co-infection with other pathogens. Negative results do not preclude SARS-CoV-2, influenza A or influenza B infection and should not be used as the sole basis for patient management decisions. | the simultaneous, qualitative detection and identification of multiple respiratory viral nucleic acids in nasopharyngeal swab (NPS) specimens obtained from individuals with signs and symptoms of respiratory tract infection, including COVID-19.

The following organism types are identified and differentiated using the SPOTFIRE R Panel Mini:
• Coronavirus SARS-CoV-2
• Human rhinovirus
• Influenza A virus
• Influenza B virus
• Respiratory syncytial virus

Nucleic acids from the viral organisms identified by this test are generally detectable in NPS specimens during the acute phase of infection. The detection and identification of specific viral nucleic acids from individuals exhibiting signs and/or symptoms of respiratory infection are indicative of the presence of the identified microorganism and aids in diagnosis if used in conjunction with other |
| --- | --- | --- |

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| | | 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.

Negative results in the setting of a respiratory illness may be due to infection with pathogens that are not detected by this test, or lower respiratory tract infection that may not be detected by an NPS specimen. Positive results do not rule out coinfection with other organisms. The agent(s) detected by the SPOTFIRE R Panel Mini may not be the definite cause of disease.

Additional laboratory testing (e.g., bacterial and viral culture, immunofluorescence, and radiography) may be necessary when evaluating a patient with possible respiratory tract infection. |
| --- | --- | --- |
| Test Format | Same | Single use |
| Results Reported | Same | Qualitative |
| Time to Result | Same | ~15 minutes |
| General Device Characteristic Differences | | |
| Target viruses | SARS-CoV-2 | • Coronavirus SARS- |

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| | Influenza A
Influenza B | CoV-2
• Human rhinovirus
• Influenza A virus
• Influenza B virus
• Respiratory syncytial virus |
| --- | --- | --- |
| Specimen Type | Anterior Nasal Swab | Nasopharyngeal swab in transport media |
| Automated Test Processes | Sample extraction, amplification, and detection | Amplification and detection |
| Controls | Internal procedural Control included in the cartridge
External Controls (purchased separately) | Two controls are included in each reagent pouch to control for sample processing and both stages of PCR and melt analysis.
External Controls are provided separately (see K230868) |
| Instrument Systems | DASH Rapid PCR Instrument | SPOTFIRE System |

VI Standards/Guidance Documents Referenced:

- IEC 60601-1-2 Ed. 4.1:2020 Medical electrical equipment - Part 1-2: General requirements for basic safety and essential performance - Collateral Standard: Electromagnetic disturbances - Requirements and tests
- IEC 61010-1, 3rd edition, May 11, 2012, revision 2023-06-06: Safety requirements for electrical equipment for measurement, control, and laboratory use - Part 1: general requirements
- IEC 61010-2-010:2019 Safety requirements for electrical equipment for measurement, control, and laboratory use - Part 2-010: Particular requirements for laboratory equipment for the heating of materials
- IEC 61010-2-101:2018 Safety requirements for electrical equipment for measurement, control, and laboratory use - Part 2-101: Particular requirements for in vitro diagnostic (IVD) medical equipment
- CLSI EP12 3rd Edition, Evaluation of Qualitative, Binary Output Examination Performance
- CLSI EP12-A2, User Protocol for Evaluation of Qualitative Test Performance; Approved Guideline - Second Edition.
- CLSI MM03-3rd Edition, Molecular Diagnostic Methods for Infectious Diseases; Approved Guideline
- CLSI EP07 3rd Edition, Interference Testing in Clinical Chemistry
- CLSI EP37 1st Edition, Supplemental Tables for Interference Testing in Clinical Chemistry

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- CLSI EP25 2nd Edition, Evaluation of Stability of In Vitro Medical Laboratory Test Reagents
- CLSI EP25-A, Evaluation of Stability of In Vitro Diagnostic Reagents; Approved Guideline

## VII Performance Characteristics (if/when applicable):

## A Analytical Performance:

1. Precision/Reproducibility:

### a) Precision/Repeatability

Precision of the DASH SARS-CoV-2 &amp; Flu A/B Test was evaluated over a 12-day period, conducted by two operators at a single site, and resulted in 96 replicates per concentration of the SARS-CoV-2, Flu A and Flu B targets. Test samples were contrived in simulated clinical nasal matrix, co-spiked with SARS-CoV-2, Flu A, and Flu B to target concentrations. Three panel members were prepared: true negative (unspiked), low positive (2x LoD (near Limit of Detection)), and moderate positive (5x LoD). The test samples were prepared by applying the contrived viral dilutions onto the swabs. The samples were randomized and blind-coded. Each operator tested two replicates of each sample in each run (two runs per day), for a total of 96 replicates per panel member. Four DASH Instruments were utilized during the execution of this study. Results are summarized in the table below.

| Sample | Sample Concentration | Operator | n/N[1] | Agreement with Expected Results (%) |
| --- | --- | --- | --- | --- |
| SARS-CoV-2 | Low Positive | 1 | 48/48 | 100.0 |
| | | 2 | 48/48 | |
| | Moderate Positive | 1 | 48/48 | 100.0 |
| | | 2 | 48/48 | |
| Flu A | Low Positive | 1 | 48/48 | 100.0 |
| | | 2 | 48/48 | |
| | Moderate Positive | 1 | 48/48 | 100.0 |
| | | 2 | 48/48 | |
| Flu B | Low Positive | 1 | 48/48 | 100.0 |
| | | 2 | 48/48 | |
| | Moderate Positive | 1 | 48/48 | 100.0 |
| | | 2 | 48/48 | |
| Negative | N/A | 1 | 48/48 | 100.0 |
| | | 2 | 48/48 | |

[1]n is number of tests with expected results. N is the total number of valid tests

| Sample | Sample Conc. | n/N[1] | Mean Cq[2] | Between Operator | Between Day | Between Runs | Within Run/Repeatability | Total |
| --- | --- | --- | --- | --- | --- | --- | --- | --- |

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# b) Reproducibility

Another study was conducted to evaluate the reproducibility of the DASH SARS-CoV-2 &amp; Flu A/B Test at three external CLIA-waived sites with a total of nine untrained operators. A total of three lots of the Test cartridges were used. Operators tested a randomized blind-coded panel of contrived samples at three concentration levels (prepared in the same manner as above for the precision study), with three replicates per level tested by each operator over five days. There were 270 replicates of each concentration tested over the study period (3 replicates x 2 runs x 3 operators x 5 days x 3 sites). Each operator ran one set of external controls (i.e., positive control and negative control) on their assigned instrument prior to testing reproducibility samples daily.

The results are summarized below in Table 4 and 5. No significant differences between sites, or between operators were observed.

Table 4: Summary of Qualitative Results for Reproducibility Study Results

| Analyte | Sample Concentration | % Agreement with Expected Results[1] (n/N[2]) (95% Confidence Interval) | | | | | | |
| --- | --- | --- | --- | --- | --- | --- | --- | --- |
| | | Site 1 | Site 2 | Site 3 | Overall | | | |
| SARS-CoV-2 | Low Positive | 100.0% (90/90) (95.9% -100.0%) | 97.8% (88/90) (92.3% - 99.4%) | 100.0% (90/90) (95.9% -100.0%) | 99.3% (268/270) (97.3% -99.8%) | | | |
| | Moderate Positive | 98.9% (89/90) (94.0% - 99.9%) | 100.0% (89/89)[3] (95.9% -100.0%) | 100.0% (90/90) (95.9% -100.0%) | 99.6% (268/269) (97.9% -100.0%) | | | |
| | Negative | 100.0% (90/90) (95.9% -100.0%) | 100.0% (90/90) (95.9% -100.0%) | 100.0% (90/90) (95.9% -100.0%) | 100.0% (270/270) (98.6% -100.0%) | | | |
| Flu A | Low Positive | 100.0% (90/90) | 97.8% (88/90) | 100.0% (90/90) | 99.3% (268/270) | | | |

[1]n is number of tests with expected results. N is the total number of valid tests
[2]DASH software identifies the cycle at which an amplification curve meets a predefined mathematical criterion as Cq.
$\mathrm{Cq} =$ quantification cycle, $\mathrm{SD} =$ standard deviation, $\% \mathrm{CV} =$ percent coefficient of variation

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Table 5: Summary of Reproducibility Results (Cq Variability Analysis)

| Analyte | Sample Conc. | n/N[1] | Mean Cq[2] | Between Site | Between Lot | Between Day | Between Operator | Between Run | Within-Run | Total |
| --- | --- | --- | --- | --- | --- | --- | --- | --- | --- | --- |
| SD | %CV | SD | %CV | SD | %CV | SD | %CV | SD | %CV | SD | %CV | SD | %CV |
| SARS-CoV-2 | Low Positive | 268/270 | 29.6 | 0.2 | 0.7 | 0.6 | 2.1 | 0 | 0 | 0.0 | 0.0 | 0.0 | 0.0 | 2.7 | 9.2 | 2.8 | 9.4 |
| Moderate Positive | 268/269 | 28.4 | 0.0 | 0.0 | 0.5 | 1.7 | 0 | 0 | 0.4 | 1.5 | 0.0 | 0.0 | 1.9 | 6.5 | 2.0 | 6.9 |
| Flu A | Low Positive | 268/270 | 24.2 | 0.0 | 0.2 | 0.0 | 0.0 | 0 | 0 | 0.0 | 0.0 | 0.4 | 1.6 | 2.3 | 9.5 | 2.3 | 9.6 |
| Moderate Positive | 269/269 | 23.1 | 0.1 | 0.3 | 0.1 | 0.6 | 0 | 0 | 0.3 | 1.3 | 0.0 | 0.0 | 0.9 | 3.7 | 0.9 | 4.0 |
| Flu B | Low Positive | 268/270 | 26.1 | 0.0 | 0.0 | 0.3 | 1.1 | 0 | 0 | 0.3 | 1.2 | 0.0 | 0.0 | 2.3 | 8.9 | 2.4 | 9.1 |
| Moderate Positive | 269/269 | 25.3 | 0.2 | 0.8 | 0.2 | 0.7 | 0 | 0 | 0.2 | 0.7 | 0.2 | 0.8 | 0.7 | 2.9 | 0.8 | 3.3 |

[1]A total of 15 samples generated invalid results on the first attempt; 14 samples generated valid results upon repeat and were included in the data analysis.
[2]n is number of tests with expected results. N is the total number of valid tests
[3]One moderate positive sample produced an error/no result and was excluded from the analysis, yielding a total of 89 results for this panel member at Site 2.

[1]n is number of tests with expected results. N is the total number of valid tests
[2]DASH software identifies the cycle at which an amplification curve meets a predefined mathematical criterion as Cq
$\mathrm{Cq} =$ quantification cycle, $\mathrm{SD} =$ standard deviation, $\% \mathrm{CV} =$ percent coefficient of variation

# 2. Linearity:

Not applicable; this is a qualitative assay.

# 3. Analytical Specificity/Interference:

# a) Cross-Reactivity Study

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# Wet Testing

Cross-Reactivity was evaluated for the DASH SARS-CoV-2 &amp; Flu A/B Test on the DASH Instrument by wet testing of 50 different viruses, bacteria, and fungi that could potentially interfere with the assay targets. All samples were prepared in simulated clinical nasal matrix and tested individually, without the presence of the test viral targets. The testing concentrations for potentially interfering microorganisms were $\geq 10^{5}$ units/mL for viruses and $\geq 10^{6}$ units/mL for other microorganisms, unless otherwise noted in the table below.

None of the evaluated organisms demonstrated cross-reactivity with the assay at the tested concentrations as shown in the table below.

Table 6: Cross-Reactivity Testing Results

| Microorganism | Microorganism Concentration (per Swab) | Microorganism Concentration (per mL) |
| --- | --- | --- |
| Adenovirus type C1 | 1.44E+04 TCID50/swab | 3.6E+05 TCID50/mL |
| Cytomegalovirus (also known as HHV-5) | 2.82E+03 TCID50/swab | 7.05E+04 TCID50/mL |
| Epstein-Barr virus (also known as HHV-4) | 1.51E+04 TCID50/swab | 3.78E+05 TCID50/mL |
| Human coronavirus 229E | 4.00E+03 TCID50/swab | 1.00E+05 TCID50/mL |
| Human coronavirus OC43 | 4.75E+03 TCID50/swab | 2.19E+05 TCID50/mL |
| Human coronavirus HKU1 | 1.10E+04 Copies/swab | 2.75E+05 Copies/mL |
| Human coronavirus NL63 | 4.00E+03 TCID50/swab | 1.00E+05 TCID50/mL |
| Human Metapneumovirus | 1.19E+04 TCID50/swab | 2.98E+05 TCID50/mL |
| Measles virus | 1.54E+04 TCID50/swab | 3.85E+05 TCID50/mL |
| Middle East Respiratory Syndrome coronavirus (MERS-CoV) | N/A[1] | |
| Genomic RNA Middle East Respiratory Syndrome coronavirus (MERS-CoV) EMC/2012 | 1.35 ng RNA/swab | 33.8 ng RNA/mL |
| Mumps virus | 1.19E+04 TCID50/swab | 2.98E+05 TCID50/mL |
| Parainfluenza virus 1 | 4.75E+03 TCID50/swab | 1.19E+05 TCID50/mL |
| Parainfluenza virus 2 | 4.00E+03 TCID50/swab | 1.00E+05 TCID50/mL |
| Parainfluenza virus 3 | 1.44E+04 TCID50/swab | 3.6E+05 TCID50/mL |
| Parainfluenza virus 4 | 2.82E+03 TCID50/swab | 7.05E+04 TCID50/mL |
| Respiratory syncytial virus | 2.82E+03 TCID50/swab | 7.05E+04 TCID50/mL |
| Rhinovirus type 1A | 2.82E+03 TCID50/swab | 7.05E+04 TCID50/mL |

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| Enterovirus type 68 | 3.40E+03 TCID_{50}/swab | 8.5E+04 TCID_{50}/mL |
| --- | --- | --- |
| Severe Acute Respiratory Syndrome coronavirus (2003) RNA | 9.80E+03 Copies/swab | 2.45E+05 Copies/mL |
| Bordetella pertussis | 5.00E+04 CFU/swab | 1.25E+06 CFU/mL |
| Bordetella parapertussis | 5.00E+04 CFU/swab | 1.25E+06 CFU/mL |
| Chlamydia pneumoniae | 5.00E+04 CFU/swab | 1.25E+06 CFU/mL |
| Corynebacterium diphtheriae | 5.00E+04 CFU/swab | 1.25E+06 CFU/mL |
| Escherichia coli | 5.00E+04 CFU/swab | 1.25E+06 CFU/mL |
| Haemophilus influenzae | 5.00E+04 CFU/swab | 1.25E+06 CFU/mL |
| Fusobacterium necrophorum | 5.00E+04 CFU/swab | 1.25E+06 CFU/mL |
| Lactobacillus acidophilus | 5.00E+04 CFU/swab | 1.25E+06 CFU/mL |
| Legionella pneumophila | 5.00E+04 CFU/swab | 1.25E+06 CFU/mL |
| Moraxella catarrhalis | 5.00E+04 CFU/swab | 1.25E+06 CFU/mL |
| Mycobacterium tuberculosis (heat-inactivated) | 5.00E+04 CFU/swab | 1.25E+06 CFU/mL |
| Mycoplasma pneumoniae | 5.00E+04 CFU/swab | 1.25E+06 CFU/mL |
| Mycoplasma genitalium | 5.00E+04 CFU/swab | 1.25E+06 CFU/mL |
| Neisseria meningitidis | 5.00E+04 CFU/swab | 1.25E+06 CFU/mL |
| Neisseria elongata subsp. glycolytica) | 5.00E+04 CFU/swab | 1.25E+06 CFU/mL |
| Neisseria mucosa | 5.00E+04 CFU/swab | 1.25E+06 CFU/mL |
| Nocardia asteroides | 1.74E+05 CFU/swab | 4.35E+06 CFU/swab |
| Pseudomonas aeruginosa | 5.00E+04 CFU/swab | 1.25E+06 CFU/mL |
| Staphylococcus aureus | 5.00E+04 CFU/swab | 1.25E+06 CFU/mL |
| Staphylococcus epidermidis | 5.00E+04 CFU/swab | 1.25E+06 CFU/mL |
| Streptococcus pneumoniae | 5.00E+04 CFU/swab | 1.25E+06 CFU/mL |
| Streptococcus pyogenes | 5.00E+04 CFU/swab | 1.25E+06 CFU/mL |
| Streptococcus salivarius | 5.00E+04 CFU/swab | 1.25E+06 CFU/mL |
| Streptococcus mitis | 5.00E+04 CFU/swab | 1.25E+06 CFU/mL |
| Streptococcus mutans | 5.00E+04 CFU/swab | 1.25E+06 CFU/mL |
| Aspergillus niger | 5.00E+04 CFU/swab | 1.25E+06 CFU/mL |
| Candida albicans | 5.00E+04 CFU/swab | 1.25E+06 CFU/mL |

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In silico

In silico cross-reactivity of the DASH SARS-CoV-2 &amp; Flu A/B Test was explored by evaluating genomes representing the viral, bacterial, and fungal organisms in the wet testing cross-reactivity study (see table above). 10% of complete genomes of these microorganisms were randomly selected and downloaded from GenBank to be included in this analysis. Any microorganism showing &gt;80% homology to a single primer, probe, or capture oligonucleotide was further evaluated for homology for the other target primers and probes for amplification. DASH SARS-CoV-2 &amp; Flu A/B Test's forward and reverse primers for the SARS-CoV-2 target exhibited &gt;80% homology to SARS-coronavirus Tor2 isolate and therefore may be amplified but will not be detected due to inability of the test target probes to bind. Performance of DASH SARS-CoV-2 &amp; Flu A/B Test with SARS-coronavirus Tor2 has not been confirmed by wet testing analysis.

### b) Microbial Interference Study

Microbial Interference was evaluated for the DASH SARS-CoV-2 &amp; Flu A/B Test on the DASH Instrument by wet testing over 50 different viruses, bacteria, and fungi that are common causes of respiratory infections. All samples were prepared in simulated clinical nasal matrix and tested with individual microorganisms or with pooled groups of up to four microorganisms in the presence of the test viral targets, SARS-CoV-2, influenza A and influenza B, co-spiked at 3x LoD. Microbial non-interference was determined if the target had a 100% positive call rate for all three replicates tested (3/3).

None of the evaluated microorganisms demonstrated interference with the assay at the tested concentrations as shown in the table below.

Table 7: Microbial Interference Testing Results

| Microorganism | Microorganism Concentration (per Swab) | Microorganism Concentration (per mL) |
| --- | --- | --- |
| Adenovirus C1 | 1.44E+04 TCID_{50}/swab | 3.6E+05 TCID_{50}/mL |
| Cytomegalovirus (also known as HHV-5) | 2.48E+03 TCID_{50}/swab | 6.2E+04 TCID_{50}/mL |
| Epstein-Barr virus (also known as HHV-4) | 1.51E+04 TCID_{50}/swab | 3.78E+05 TCID_{50}/mL |
| Human coronavirus 229E | 4.00E+03 TCID_{50}/swab | 1.0E+05 TCID_{50}/mL |
| Human coronavirus OC43 | 4.75E+03 TCID_{50}/swab | 2.19E+05 TCID_{50}/mL |
| Human coronavirus HKU1 | 9.68E+03 Copies/swab | 2.41E+05 Copies/mL |
| Human coronavirus NL63[1] | 4.0E+03 TCID_{50}/swab | 1.0E+05 TCID_{50}/mL |
| Measles virus[1] | 1.54E+04 TCID_{50}/swab | 3.85E+05 TCID_{50}/mL |

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| Genomic RNA Middle East Respiratory Syndrome coronavirus (MERS-CoV) | 1.35 ng RNA/swab | 33.8 ng RNA/mL |
| --- | --- | --- |
| Middle East Respiratory Syndrome coronavirus (MERS-CoV) | N/A^{[2]} | |
| Mumps virus | 1.19E+04 TCID_{50}/swab | 2.98E+05 TCID_{50}/mL |
| Human Metapneumovirus | 1.19E+04 TCID_{50}/swab | 2.98E+05 TCID_{50}/mL |
| Parainfluenza virus 1 | 4.75E+03 TCID_{50}/swab | 1.19E+05 TCID_{50}/mL |
| Parainfluenza virus 2 | 4.0E+03 TCID_{50}/swab | 1.0E+05 TCID_{50}/mL |
| Parainfluenza virus 3 | 1.44E+04 TCID_{50}/swab | 3.6E+05 TCID_{50}/mL |
| Parainfluenza virus 4 | 2.48E+03 TCID_{50}/swab | 6.20E+04 TCID_{50}/mL |
| Respiratory syncytial virus | 2.48E+03 TCID_{50}/swab | 6.2E+04 TCID_{50}/mL |
| Rhinovirus type 1A | 2.48E+03 TCID_{50}/swab | 6.2E+04 TCID_{50}/mL |
| Enterovirus type 68 | 2.99E+03 TCID_{50}/swab | 7.48E+04 TCID_{50}/mL |
| Severe Acute Respiratory Syndrome coronavirus (2003) RNA | 8.62E+03 Copies/swab | 2.16E+05 Copies/mL |
| Bordetella pertussis | 5.0E+04 CFU/swab | 1.25E+06 CFU/mL |
| Bordetella parapertussis | 5.0E+04 CFU/swab | 1.25E+06 CFU/mL |
| Candida albicans | 5.0E+04 CFU/swab | 1.25E+06 CFU/mL |
| Chlamydia pneumoniae | 5.0E+04 CFU/swab | 1.25E+06 CFU/mL |
| Corynebacterium diphtheriae | 5.0E+04 CFU/swab | 1.25E+06 CFU/mL |
| Escherichia coli | 5.0E+04 CFU/swab | 1.25E+06 CFU/mL |
| Haemophilus influenzae | 5.0E+04 CFU/swab | 1.25E+06 CFU/mL |
| Fusobacterium necrophorum | 5.0E+04 CFU/swab | 1.25E+06 CFU/mL |
| Lactobacillus acidophilus | 5.0E+04 CFU/swab | 1.25E+06 CFU/mL |
| Legionella pneumophila | 5.0E+04 CFU/swab | 1.25E+06 CFU/mL |
| Moraxella catarrhalis | 5.0E+04 CFU/swab | 1.25E+06 CFU/mL |
| Mycobacterium tuberculosis (heat- inactivated) | 5.0E+04 CFU/swab | 1.25E+06 CFU/mL |
| Mycoplasma pneumoniae | 5.0E+04 CFU/swab | 1.25E+06 CFU/mL |
| Mycoplasma genitalium | 5.0E+04 CFU/swab | 1.25E+06 CFU/mL |
| Neisseria meningitidis | 5.0E+04 CFU/swab | 1.25E+06 CFU/mL |
| Neisseria elongata subsp. glycolytica | 5.0E+04 CFU/swab | 1.25E+06 CFU/mL |
| Pseudomonas aeruginosa | 5.0E+04 CFU/swab | 1.25E+06 CFU/mL |
| Staphylococcus aureus | 5.0E+04 CFU/swab | 1.25E+06 CFU/mL |
| Staphylococcus epidermidis | 5.0E+04 CFU/swab | 1.25E+06 CFU/mL |
| Streptococcus pneumoniae | 5.0E+04 CFU/swab | 1.25E+06 CFU/mL |

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| Streptococcus pyogenes | 5.0E+04 CFU/swab | 1.25E+06CFU/mL |
| --- | --- | --- |
| Streptococcus salivarius | 5.0E+04 CFU/swab | 1.25E+06 CFU/mL |
| Aspergillus niger | 5.0E+04 CFU/swab | 1.25E+06 CFU/mL |
| Cryptococcus neoformans | 5.0E+04 CFU/swab | 1.25E+06 CFU/mL |
| Pneumocystis jirovecii | 5.0E+04 CFU/swab | 1.25E+06 CFU/mL |
| Eikenella corrodens | 5.0E+04 CFU/swab | 1.25E+06 CFU/mL |
| Neisseria mucosa | 5.0E+04 CFU/swab | 1.25E+06 CFU/mL |
| Streptococcus mitis | 5.0E+04 CFU/swab | 1.25E+06 CFU/mL |
| Streptococcus mutans | 5.0E+04 CFU/swab | 1.25E+06 CFU/mL |
| Nocardia asteroides | 1.74E+05 CFU/swab | 4.35E+06 CFU/mL |

Note: Thicker outer border indicates groups of pooled microorganisms that were tested.
[1] Tested individually as interference was observed when tested as a group.
[2] Concentration not available from vendor. Sample is qualitative non-infectious purified intact viral particles.

## c) Competitive Interference Study

The impact of competitive interference, caused by co-infections with on-target analytes, was evaluated for the DASH SARS-CoV-2 &amp; Flu A/B Test on the DASH Instrument by testing contrived samples consisting of individual SARS-CoV-2, Flu A or Flu B strains at high concentrations in the presence of the other two target strains at 3x LoD. For this study, competitive interference was assessed using one strain each of SARS-CoV-2, influenza A and influenza B. Testing for each target strain (at 3x LoD concentration) and each potential competitive strain (at high concentration) was performed in triplicate. No competitive interference was observed if all replicates for the low concentration (3x LoD) target yielded positive results.

The study showed that SARS-CoV-2 at concentrations &gt;1.41E+06 copies/mL inhibited detection of Flu A at 3x LoD, and at SARS-CoV-2 concentrations of &gt;5.65E+06 copies/mL inhibited detection of Flu B at 3x LoD. Flu A at concentration of 1.38E+06 TCID₅₀/mL inhibited detection of Flu B at 3x LoD. In addition, Flu B at concentrations above 1.01E+03 TCID₅₀/mL inhibited detection of both SARS-CoV-2 and Flu A at 3x LoD. The highest coinfection concentration of the competing virus, where the remaining two targets are detected for three replicates at 3x LoD, are reported in the table below.

Table 8: Competitive Interference Study Results

| Competing Virus | Concentration at which All 3 Targets are Positive | | |
| --- | --- | --- | --- |
| | SARS-CoV-2 | Flu A | Flu B |
| SARS-CoV-2 | 1.41E+06 copies/mL ~47x LoD | 3x LoD | 3x LoD |
| Flu A | 3x LoD | 1.38E+06 TCID₅₀/mL ~3405x LoD | 3x LoD |

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# d) Exogenous/Endogenous Interfering Substances

The potential for interference for the DASH SARS-CoV-2 &amp; Flu A/B Test was evaluated with substances that may be present in respiratory specimens. A total of 22 potentially interfering endogenous and exogenous substances were tested at or above clinically relevant levels in simulated clinical nasal matrix co-spiked at ~3x LoD with SARS-CoV-2, Flu A and Flu B inactivated viral particles. Each sample was tested in triplicate. The FluMist nasal vaccine was not tested.

Two of the substances tested in the study, Biotin (vitamin B-7) at 4.58 µg/mL and Flonase (active ingredient Fluticasone Propionate) at 5% (v/v) gave false negative results with SARS-CoV-2 target. Additional testing at lower test substance concentrations were performed to determine the concentration where interference is no longer observed. No interference was observed for any of the substances tested at the concentrations noted in the table below.

Table 9: Exogenous/Endogenous Interfering Substances Study Results

| Interfering Substance | Concentration |
| --- | --- |
| Afrin Original Nasal Spray | 15% v/v |
| Chloroseptic Sore Throat spray | 20% v/v |
| Flonase | 2.5% v/v[1] |
| Relenza | 0.3 mg/ml |
| Tobramycin | 4 µg/mL |
| Mupirocin | 10 mg/mL |
| Mucin | 0.1% w/v |
| Blood | 2% v/v |
| Biotin | 2.29 µg/mL[1] |
| Viral Transport Medium | 50% v/v |
| Neo-Synephrine Nasal Spray | 15% v/v |
| Walgreens Saline Nasal Spray | 15% v/v |
| Beclomethasone Dipropionate, USP (Micronized) | 0.068 mg/mL |
| Flunisolide Nasal Solution, USP 0.025% | 0.04 mg/mL |
| Nasacort Nasal Spray | 0.04 mg/mL |
| Walgreens 24 Hour Budesonide Nasal Spray | 0.051 mg/mL |
| Nasonex | 0.04 mg/mL |
| Zicam Allergy Relief Nasal Spray | 20% v/v |
| Snuff | 0.1% w/v |
| Latex glove powder | 1% w/v |

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| Mucinex Insta Sooth Kickstart Sore Throat | 1.7 mg/mL |
| --- | --- |
| Zinc gluconate hydrate | 0.1 μg/mL |

[1]Interference was observed at concentrations higher than these concentrations.

4. Assay Reportable Range:

Not applicable.

5. Traceability, Stability, Expected Values (Controls, Calibrators, or Methods):

a) External Controls

The DASH SARS-CoV-2 &amp; Flu A/B Positive Control and DASH Negative Control swabs are available separately for use as external controls. Please refer to Instrument Description Information (Section C.5) above for assay controls. A study was conducted to evaluate these external controls with DASH SARS-CoV-2 &amp; Flu A/B Test. A total of 45 DASH SARS-CoV-2 &amp; Flu A/B Positive Control and 45 DASH Negative Control swabs were tested daily by nine untrained operators at three CLIA-Waived sites across five testing days. Three lots of external controls were distributed evenly across all the three sites. All 45 external positive controls and all 45 external negative controls yielded accurate results.

b) Sample Stability

A study was conducted to determine the specimen stability on the swab after collection and prior to testing with the DASH SARS-CoV-2 &amp; Flu A/B Test on the DASH Instrument. Specimens included SARS-CoV-2, Flu A and Flu B, co-spiked in pooled negative nasal matrix at a concentration of 2x LoD. Twenty replicates of the positive samples were tested fresh (T0) and after storage at multiple time points (5, 10, 25, 30 and 40 minutes) at room temperature. Negative samples were also included and were tested in three replicates for each storage condition. Results indicated that positive samples remained stable, and all targets were detected up to twenty-five minutes after collection and storage at room temperature. Negative samples tested correctly for each timepoint.

Per test labeling, patient samples should be tested immediately after collection for optimal test performance.

c) Matrix Equivalency

The objective of this study was to establish equivalent performance of the DASH SARS-CoV-2 &amp; Flu A/B Test on the DASH Instrument between the two matrices used in the analytical studies: clinical pooled nasal matrix (PNM) and simulated clinical nasal matrix (SCNM). SCNM was formulated in phosphate-buffered saline (PBS) with glycerol and mucin, added to mimic the nasal cavity fluid. For this study, SARS-CoV-2 (Isolate hCoV-19/USA/GAEHC-2811C/2021), Flu A (A/Victoria/2570/2019) and Flu B (B/Utah/9/2014) were co-spiked in both matrices at 2x and 5x LoD. Aliquots of negative matrices were also included in the evaluation. For both matrices, ten replicates of the negative samples, 40 replicates of the samples prepared at 2x LoD and ten replicates of the samples prepared at 5x LoD were tested. The acceptance criteria to demonstrate equivalency was ≥95% detection for

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samples at 2x LoD, $100\%$ detection for the samples at 5x LoD for each target, and $0\%$ detection of the negative samples. The results obtained in this study are summarized below. The data demonstrated equivalent performance of the test with both the matrices.

Table 10: Matrix Equivalency Study Results

| Viral Strain | Concentration | Matrix | n/N[1] | Detection Rate (% Positive) |
| --- | --- | --- | --- | --- |
| SARS CoV-2 | 2x LoD | PNM | 39/40 | 98 |
| | | SCNM | 38/40 | 95 |
| | 5x LoD | PNM | 10/10 | 100 |
| | | SCNM | 10/10 | 100 |
| Flu A | 2x LoD | PNM | 40/40 | 100 |
| | | SCNM | 40/40 | 100 |
| | 5x LoD | PNM | 10/10 | 100 |
| | | SCNM | 10/10 | 100 |
| Flu B | 2x LoD | PNM | 39/40 | 98 |
| | | SCNM | 40/40 | 100 |
| | 5x LoD | PNM | 10/10 | 100 |
| | | SCNM | 10/10 | 100 |
| Negative Matrix | N/A | PNM | 10/10 | 0 |
| | | SCNM | 10/10 | 0 |

$^{[1]}$ n is number of tests with expected results. N is the total number of valid tests

# 6. Detection Limit:

Limit of detection (LoD) of the DASH SARS-CoV-2 &amp; Flu A/B Test on the DASH Rapid PCR Instrument was determined utilizing five viral strains representing the test targets. All samples were prepared in pooled negative nasal matrix. Initially, three-fold serial dilutions of each strain in pooled negative nasal matrix were prepared and tested in replicates of four to find the lowest level detectable by the DASH SARS-CoV-2 &amp; Flu A/B Test. The final LoD, a concentration producing $95\%$ positivity, was confirmed by testing twenty replicates of each of the strains at the selected preliminary LoD. The final LoD results are summarized in the tables below.

Table 11: Limit of Detection Study Results

| Viral Target | Strain | Source/ Product Type | LoD (copies or TCID50/swab) | LoD (copies or TCID50/mL) |
| --- | --- | --- | --- | --- |
| SARS-CoV-2 | hCoV- 19/USA/GA-EHC- 2811C/2021 (Lineage B.1.1.529; Omicron Variant) | BEI Resources/ Gamma-Irradiated | 1200 copies/swab | 3.0E+04 copies/mL |
| Flu A | H1N1 Victoria/2570/19 | Zeptometrics/ Culture Fluid | 1.35 TCID50/swab | 33.75 TCID50/mL |
| | H3N2 Darwin/9/21 | Zeptometrics/ Culture Fluid | 0.225 TCID50/swab | 5.63 TCID50/mL |
| Flu B | Washington/02/19 - Victoria Lineage | Zeptometrics/ Culture Fluid | 0.1 TCID50/swab | 2.50 TCID50/mL |

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7. Inclusivity (Analytical Reactivity):

Wet Testing

The inclusivity of the DASH SARS-CoV-2 &amp; Flu A/B Test on the DASH Instrument was evaluated by testing seven strains of SARS-CoV-2, 21 strains of Flu A, and 10 strains of Flu B. Positive samples were contrived by spiking each virus strain, individually, into negative pooled nasal matrix at concentrations of ~3x LoD and testing in triplicate in a blinded fashion. Inclusivity was determined if the target had a 100% detection (3/3) for all three replicates tested. If less than 100% detection was observed, the strain was prepared at a higher concentration and tested until 100% detection was achieved. The strains evaluated and the lowest concentration that met the inclusivity criteria outlined above are shown in the table below.

Table 12: Inclusivity Wet Testing Results

| Test Target Variant/Subtype | Virus Concentration (per Swab) | Virus Concentration (per mL) |
| --- | --- | --- |
| SARS-CoV-2
B.1.1.7 (England/204820464/2020) | 0.0828 TCID_{50}/swab^{[1]} | 2.07 TCID_{50}/mL^{[1]} |
| SARS-CoV-2
B.1.351 (South Africa/KRISP-K005325/2020) | 0.0414 TCID_{50}/swab | 1.04 TCID_{50}/mL |
| SARS-CoV-2
B.1.617.2; Delta Variant (USA/PHC658/2021) | 0.1656 TCID_{50}/swab^{[2]} | 4.14 TCID_{50}/mL^{[2]} |
| SARS-CoV-2
P1; Gamma Variant (Japan/TY7-503/2021) | 0.0414 TCID_{50}/swab | 1.04 TCID_{50}/mL |
| SARS-CoV-2
B.1.617.1; Kappa Variant (USA/CA-Stanford-15_S02/2021) | 0.0828 TCID_{50}/swab^{[1]} | 2.07 TCID_{50}/mL^{[1]} |
| SARS-CoV-2
BA 2.3; Omicron Variant | 0.0414 TCID_{50}/swab | 1.04 TCID_{50}/mL |
| Gamma-irradiated SARS-CoV-2
isolate USA-WA1/2020 | 7200 copies/swab | 1.8E+05 copies/mL |
| Influenza A H1N1 (New Cal/20/99) | 4.05 TCID_{50}/swab | 101.25 TCID_{50}/mL |
| Influenza A H1N1 (PR/8/34) | 4.05 TCID_{50}/swab | 101.25 TCID_{50}/mL |
| Influenza A H1N1 (Brisbane/59/07) | 4.05 TCID_{50}/swab | 101.25 TCID_{50}/mL |
| Influenza A H1N1 (Taiwan/42/06) | 4.05 TCID_{50}/swab | 101.25 TCID_{50}/mL |
| Influenza A H1N1 (Singapore/63/04) | 4.05 TCID_{50}/swab^{[3]} | 101.25 TCID_{50}/mL^{[3]} |
| Influenza A H1N1pdm (California/07/09) | 4.05 TCID_{50}/swab | 101.25 TCID_{50}/mL |
| Influenza A H1N1pdm (New York/18/09) | 4.05 TCID_{50}/swab | 101.25 TCID_{50}/mL |
| Influenza A H1N1pdm Virus (Guangdong-Maonan/SWL 1536/19) | 4.05 TCID_{50}/swab | 101.25 TCID_{50}/mL |
| Influenza A H1N1pdm (Mexico/4108/09) | 4.05 TCID_{50}/swab | 101.25 TCID_{50}/mL |
| Influenza A H1N1pdm (Victoria/2570/19) | 4.05 TCID_{50}/swab | 101.25 TCID_{50}/mL |

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| Influenza A H3N2 (Wisconsin/67/05) | 4.05 TCID50/swab | 101.25 TCID50/mL |
| --- | --- | --- |
| Influenza A H3N2 Virus (Hong Kong/2671/19) | 8.1 TCID50/swab[1] | 202.50 TCID50/mL[1] |
| Influenza A H3N2 (Stockholm/6/14) | 4.05 TCID50/swab | 101.25 TCID50/mL |
| Influenza A H3N2 (Hong Kong/8/68) | 4.05 TCID50/swab | 101.25 TCID50/mL |
| Influenza A H3N2 (Texas/50/12) | 4.05 TCID50/swab | 101.25 TCID50/mL |
| Influenza A H3N2 (Kansas/14/17) | 8.1 TCID50/swab[1] | 202.50 TCID50/mL[1] |
| Influenza A H3N2 (California/7/04) | 4.05 TCID50/swab | 101.25 TCID50/mL |
| Influenza A H3N2 (Norway/466/14) | 4.05 TCID50/swab | 101.25 TCID50/mL |
| Influenza A H3N2 (Kumamoto/102/02) | 4.05 TCID50/swab | 101.25 TCID50/mL |
| Influenza A H3N2 (Brisbane/10/07) | 8.1 TCID50/swab[1] | 202.5 TCID50/mL[1] |
| Influenza A H5N3 (Genomic RNA from Kilbourne F3:A/duck/Singapore/645/1997 mutant) | 1.76E+03 ng RNA/swab | 4.4E+04 ng RNA/mL |
| Influenza B (Lee/40) | 4.05 TCID50/swab[1] | 101.25 TCID50/mL[1] |
| Influenza B (Massachusetts/2/12) | 2.025 TCID50/swab | 50.63 TCID50/mL |
| Influenza B Victoria Lineage (Austria/1359417/21) | 2.025 TCID50/swab | 50.63 TCID50/mL |
| Influenza B Victoria Lineage (Victoria/2/87) | 2.025 TCID50/swab | 50.63 TCID50/mL |
| Influenza B Victoria Lineage (Alabama/2/17) | 2.025 TCID50/swab | 50.63 TCID50/mL |
| Influenza B Yamagata Lineage (Panama/45/90) | 2.025 TCID50/swab | 50.63 TCID50/mL |
| Influenza B Yamagata Lineage (Florida/04/06) | 2.025 TCID50/swab | 50.63 TCID50/mL |
| Influenza B Yamagata Lineage (Brisbane/9/14) | 2.025 TCID50/swab | 50.63 TCID50/mL |

[1]Testing at a lower concentration (i.e., 3x LoD) failed to yield $100\%$ detection. The lowest concentration that yielded $100\%$ detection was 6x LoD.
[2]Testing at a lower concentration (i.e., 3x LoD) failed to yield $100\%$ detection. The lowest concentration that yielded $100\%$ detection was 12x LoD.
[3]Influenza A H1N1 (Singapore/63/04) was detected in 2/3 replicates at 3x LoD. The strain was reprepared at 3x LoD and tested in 7 replicates for $100\%$ detection.

# In silico

The inclusivity of the DASH SARS-CoV-2 &amp; Flu A/B Test was also evaluated using in silico analysis of the sample preparation capture oligonucleotides and the PCR primers and probes for SARS-CoV-2 and Flu A and B in relation to sequences available in the National Center for Biotechnology Information (NCBI) SARS-CoV-2 and influenza virus database. Genomic sequences for SARS-CoV-2 were retrieved from the NCBI on May 8, 2024, and included all lineages and variants of concern (VOC) or variants of interest (VOI). The influenza genomic sequences were retrieved from the NCBI on May 13, 2024. Flu A sequences were limited to subtypes H1N1 (excluding Swine flu) and H3N2, based upon completeness. Flu B sequences were required to be full-length, with no subtype limitations applied. All genomic sequences containing ambiguous nucleotides were excluded from the analysis.

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The query oligonucleotides for the specific virus target were aligned to the filtered sets of viral genome sequences using the Burrows-Wheeler Aligner (BWA) program, where any difference would be a mismatch or an insertion/deletion. The genomic sequences were ranked according to the number of capture oligonucleotides and PCR primers and probes containing zero, one, and two or more mismatches. Melting temperature (Tm) analysis was performed for any oligonucleotide:genome pairing that contained a mismatch, and the pairings that passed the Tm analysis criteria were combined with pairings containing no mismatches, then grouped into amplifiable sets. The resulting oligonucleotide:genome pairings were counted to determine the number of sequences predicted to be captured and, if captured, predicted to be detected by the DASH SARS-CoV-2 &amp; Flu A/B Test. The results of this predicted-to-detect analysis are summarized in the table below:

Table 13: Sequences Predicted by In Silico Analysis to Be Detected by DASH SARS-CoV-2 &amp; Flu A/B Test

| Target | Sequences Predicted to be Detected |
| --- | --- |
| Flu A
(Subtypes H1N1 and H3N2) | ≥99.97% (18112 of 18117 sequences analyzed) |
| Flu B | ≥98.27% (8136 of 8279 sequences analyzed) |
| SARS-CoV-2 | ≥99.99% (994778 of 994846 sequences analyzed) |

Based on the analysis, 99.99% (994778/994846) of SARS-CoV-2 sequences, 99.97% (18112/18117) of influenza A subtype H1N1 and H3N2 sequences and 98.3% (8136/8279) of influenza B sequences are expected to be detected by the DASH SARS-CoV-2 &amp; Flu A/B Test.

Inclusivity Information for Non-Seasonal Influenza A Subtypes (in silico Analysis)
DASH SARS-CoV-2 &amp; Flu A/B Test in silico inclusivity analysis was performed by aligning with sequences corresponding to the gene target in Flu A subtype H5N3 and H7N7 sequences downloaded from GISAID (Global Initiative on Sharing All Influenza Data). Reactivity was predicted based on the number and location of the mismatches in the targeted region. The results of the BLAST analysis demonstrated that the subtype H5N3 and H7N7 of Flu A are expected to be detected by DASH SARS-CoV-2 &amp; Flu A/B Test. Wet testing for H5N3 strain of Flu A was also performed, confirming inclusivity of DASH SARS-CoV-2 &amp; Flu A/B Test with the H5N3 strain of Flu A (see Table 12 above). The Flu A H7N7 strain was not available for wet testing confirmation.

Additional Flu A in silico inclusivity analysis was performed for Flu A subtypes available from the GISAID database from November 5, 2023 to November 5, 2024 that have infected humans. In this analysis, all possible combinations of hemagglutinin and neuraminidase were considered, and all available sequences were required to contain the Flu A target region, found in humans, and to be full-length as reported by the submitters. All sequences containing ambiguous nucleotides were excluded from the analysis. Based on these criteria, a total of 37 sequences including subtype H5N1 (26), H5N6 (3), H9N2 (6), H10N3 (1), and H10N5 (1) were analyzed in the study. The resulting oligonucleotide:genome pairings were counted to determine the number of sequences predicted to be captured and, if captured, predicted to be detected by the DASH SARS-CoV-2 &amp; Flu A/B Test. Based on in silico

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inclusivity analysis of the Flu A capture oligonucleotides and PCR primers and probes, it is predicted that the DASH SARS-CoV-2 &amp; Flu A/B Test will detect 97.3% (36/37) of all human host Flu A sequences collected between November 5, 2023 and November 5 2024. The performance characteristics of DASH SARS-CoV-2 &amp; Flu A/B Test with these Flu A subtypes have not been confirmed by wet testing analysis.

## 8. Assay Cut-Off:

The DASH Rapid PCR Instrument has four fluorescence detectors also known as channels and an algorithm that is utilized for detection of the targets. The DASH SARS-CoV-2 &amp; Flu A/B Test does not have a simple Ct or Cq threshold below which the target is 'Detected', and above which 'Not Detected'. The Cq threshold for the test is 40, same as the total number of PCR cycles. If Cq is &gt;40, then the target is always 'Not Detected'.

## 9. Carry-Over:

An analytical study was performed to assess potential carryover or cross-contamination in the DASH SARS-CoV-2 &amp; Flu A/B Test by testing high positive and negative samples in an alternating fashion on the DASH Instrument. The high positive samples consisted of a single viral target (either SARS-CoV-2, Flu A or Flu B), prepared by spiking the viral target at ≥1×10⁵ copies/mL or ≥1×10⁵ TCID₅₀/mL in simulated clinical nasal matrix. The study utilized eight viral strains representing the test targets (two strains of SARS-CoV-2, and three strains each of Flu A and Flu B).

Samples were tested across five DASH Instruments, where one positive sample was followed by one negative sample. The process was repeated eight times to cover two strains of SARS-CoV-2, and three strains each of Flu A and Flu B. In total, 40 high positive and 40 negative samples were evaluated. All high positive samples yielded positive results for the viral targets in the sample, while all negative samples were negative. These results confirm that there is no evidence of carryover from samples tested with the DASH SARS-CoV-2 &amp; Flu A/B Test on the DASH Rapid PCR Instrument.

## B Comparison Studies:

### 1. Method Comparison with Predicate Device:

Refer to the 'Clinical Studies' section of this document.

## C Clinical Studies:

### 1. Clinical Performance:

The clinical performance of the DASH SARS-CoV-2 &amp; Flu A/B Test on DASH Instrument was established in a multi-center study conducted with anterior nasal swab (ANS) specimens collected from individuals with signs and symptoms of respiratory infection. Specimens were prospectively collected (i.e., all comers between two time points that met the clinical study inclusion criteria) from January and March 2024. Seven geographically diverse CLIA-waived testing sites within the United States participated in the study. Informed consent was obtained for all patients prior to testing. AN swabs for comparator testing were collected by

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the healthcare provider from both sides of the nose and placed into transport media for comparator testing. For testing with the DASH SARS-CoV-2 &amp; Flu A/B Test, AN swabs from both sides of the nose were either healthcare provider-collected or adult-collected from individuals below 14 years, or self-collected from individuals aged 14 years or older. The order of swab collection was alternated between the comparator swab and the DASH SARS-CoV-2 &amp; Flu A/B Test. Performance of the DASH SARS-CoV-2 &amp; Flu A/B Test was assessed by a comparison of results with an FDA cleared RT-PCR test for SARS-CoV-2, and a second FDA cleared test for Flu A and Flu B (expressed as positive percent agreement (PPA) and negative percent agreement (NPA)). All discordant results between the DASH SARS-CoV-2 &amp; Flu A/B Test and the comparator were investigated using a third highly sensitive FDA cleared test and are footnoted under the Performance table below.

A total of 819 subjects were enrolled in the study from the seven clinical sites. Of these, two subjects were later found ineligible for enrollment. Of the remaining 817 enrolled subjects, 22 were excluded due to protocol deviations (e.g., specimen shipping issues, comparator sample being unable to be tested within the stability window of the test IFU, etc.), leaving 795 evaluable specimens to be included in the calculations of estimates of assay performance. Of those, additional three specimens were deemed unevaluable for Flu A and B due to unavailability of comparator test results. In total, 795 specimens were included in the calculations of performance for SARS-CoV-2, and 792 specimens for calculations of performance for Flu A and Flu B when tested on the DASH SARS-CoV-2 &amp; Flu A/B Test.

Of the 833 samples tested on DASH SARS-CoV-2 &amp; Flu A/B Test, including 16 repeat samples, there were 24 instances of invalid results (2.9%; 95% CI: 1.9% - 4.3%). The clinical performance of the DASH SARS-CoV-2 &amp; Flu A/B Test is summarized in the table below.

Table 14: Clinical Performance of DASH SARS-CoV-2 &amp; Flu A/B Test

| Test Targets | Positive Percent Agreement | | Negative Percent Agreement | |
| --- | --- | --- | --- | --- |
| | TP/(TP+FN) | PPA (95% CI) | TN/(TN+FP) | NPA (95% CI) |
| SARS-CoV-2 | 160/168[1] | 95.2% (90.9% - 97.6%) | 624/627 | 99.5% (98.6% - 99.8%) |
| Flu A | 50/53[2] | 94.3% (84.6% - 98.1%) | 725/739[3] | 98.1% (96.8% - 98.9%) |
| Flu B | 36/37[4] | 97.3% (86.2% - 99.9%) | 749/755[5] | 99.2% (98.3% - 99.6%) |

TP-true positive; FN-false negative; TN-true negative; FP-false positive
[1]SARS-CoV-2 (PPA): One (1) sample was negative by another FDA cleared test, agreeing with the DASH.
[2]Flu A PPA: Two (2) samples were negative by another FDA cleared test, agreeing with the DASH.
[3]Flu A NPA: 13 samples were positive by another FDA cleared test, agreeing with the DASH.
[4]Flu B PPA: One (1) sample was negative by another FDA cleared test, agreeing with the DASH.
[5]Flu B NPA: Four (4) samples were positive by another FDA cleared test, agreeing with the DASH.

2. Other Clinical Supportive Data (When 1. and 2. Are Not Applicable):

Not applicable.

D Clinical Cut-Off:

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Not applicable.

## E Expected Values:

The positivity for SARS-CoV-2, Flu A and Flu B, as determined by DASH SARS-CoV-2 &amp; Flu A/B Test, are shown below, stratified by the study site and by patient age group.

Table 15: Overall Positivity Rates Observed During the Clinical Study

| Analyte | Overall | | | Site 1 (N=89) | | Site 2 (N=178) | | Site 3 (N=152) | | Site 4 (N=59) | | Site 5 (N=124) | | Site 6 (N=93) | | Site 7 (N=97) | |
| --- | --- | --- | --- | --- | --- | --- | --- | --- | --- | --- | --- | --- | --- | --- | --- | --- | --- |
| | N | # | % | # | % | # | % | # | % | # | % | # | % | # | % | # | % |
| SARS-CoV-2 | 795 | 160 | 20.1% | 10 | 11.2% | 19 | 10.7% | 25 | 16.4% | 6 | 10.2% | 29 | 23.4% | 25 | 26.9% | 46 | 47.4% |
| Flu A | 792 | 50 | 6.3% | 5 | 5.6% | 13 | 7.3% | 5 | 3.3% | 2 | 3.4% | 12 | 9.7% | 4 | 4.3% | 9 | 9.3% |
| Flu B | 792 | 36 | 4.5% | 9 | 10.1% | 6 | 3.4% | 12 | 7.9% | 2 | 3.4% | 2 | 1.6% | 4 | 4.3% | 1 | 1.0% |

N = Total evaluable specimens; # = Number of Positives; % = % positivity rate

Table 16: Positivity Rates by Age Group

| Analyte | Overall | Subject Age | | | | | | | | | |
| --- | --- | --- | --- | --- | --- | --- | --- | --- | --- | --- | --- |
| | | | | 2-<14 years | | 14-24 years | | >24-64 years | | ≥65 years | |
| | N | # | % | # | % | # | % | # | % | # | % |
| SARS-CoV-2 | 795 | 160 | 20.1% | 19 | 11.8% | 28 | 17.5% | 97 | 60.6% | 16 | 10.0% |
| Flu A | 792 | 50 | 6.3% | 8 | 16.0% | 10 | 20.0% | 29 | 58.0% | 3 | 6.0% |
| Flu B | 792 | 36 | 4.5% | 15 | 41.70% | 5 | 13.9% | 16 | 44.4% | 0 | 0.0% |

N = Total evaluable specimens; # = Number of Positives; % = % positivity rate

## VIII Proposed Labeling:

The labeling supports the finding of substantial equivalence for this device.

## IX Conclusion:

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

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