Predicate Devices

Not Found

Reference Devices

AI/ML (Artificial Intelligence / Machine Learning)

No
The device description and performance studies focus on a lateral flow immunochromatographic assay, which is a traditional biochemical test method. There is no mention of AI, ML, image processing, or any computational analysis of the results. The interpretation is based on visual lines appearing on the test strip.

Therapeutic

No
Explanation: This device is for diagnostic purposes, specifically for detecting antigens related to COVID-19 and influenza A/B, not for treating or alleviating a disease.

Diagnostic

Yes

The device is intended for the "qualitative detection and differentiation of influenza A, and influenza B nucleoprotein antigens and SARS-CoV-2 nucleocapsid antigen," which directly identifies infectious agents in individuals with symptoms. This is a diagnostic function.

SaMD (Software as a Medical Device)

No

The device description clearly states it is an immunochromatographic assay composed of a plastic housing (cassette) containing a test strip with various physical components and reagents. This is a hardware-based diagnostic test, not a software-only device.

IVD (In Vitro Diagnostic)

Based on the provided information, the Healgen Rapid Check COVID-19/Flu A&B Antigen Test is indeed an IVD (In Vitro Diagnostic).

Here's why:

Intended Use: The intended use explicitly states it's for the "qualitative detection and differentiation of influenza A, and influenza B nucleoprotein antigens and SARS-CoV-2 nucleocapsid antigen directly in anterior nasal swab samples". This is a classic description of an in vitro diagnostic test, which analyzes samples taken from the human body to provide information about a person's health.
Device Description: The description details an "immunochromatographic assay" that uses antibodies to detect specific antigens in a biological sample (anterior nasal swab). This process is performed in vitro (outside the body).
Clinical Performance Assessment: The study conducted to assess performance involved testing samples collected from individuals, which is a standard practice for evaluating IVDs.
Key Metrics: The performance metrics reported (Positive Percent Agreement, Negative Percent Agreement) are typical measures used to evaluate the accuracy of diagnostic tests.

Therefore, the Healgen Rapid Check COVID-19/Flu A&B Antigen Test fits the definition of an In Vitro Diagnostic device.

PCCP (Predetermined Change Control Plan) Authorized

N/A

Overview

Intended Use / Indications for Use

The Healgen Rapid Check COVID-19/Flu A&B Antigen Test is a lateral flow immunochromatographic assay intended for the qualitative detection and differentiation of influenza A, and influenza B nucleoprotein antigens and SARS-CoV-2 nucleocapsid antigen directly in anterior nasal swab samples from individuals with signs and symptoms of respiratory tract infection. Symptoms of respiratory infections due to SARS-CoV-2 and influenza can be similar. This test is for non-prescription home use by individuals aged 14 years or older testing themselves, or adults testing individuals aged 2 years or older.

All negative results are presumptive and should be confirmed with an FDA-cleared molecular assay when determined to be appropriate by a healthcare provider. Negative results do not rule out infection with influenza, SARS-CoV-2 or other pathogens. Individuals who test negative and experience continued or worsening respiratory symptoms, such as fever, cough and/or shortness of breath, should therefore seek follow-up care from their healthcare provider.

Positive results do not rule out co-infection with other respiratory pathogens and therefore do not substitute for a visit to a healthcare provider or appropriate follow-up.

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

SCA

Device Description

The Healgen Rapid Check COVID-19/Flu A&B Antigen Test is an immunochromatographic assay that uses highly sensitive monoclonal antibodies to detect nucleoprotein antigens from SARS-CoV-2, influenza virus types A and B in anterior nasal swab (ANS) samples from symptomatic individuals. The test device is composed of a plastic housing, known as a cassette that contains a test strip with the following parts: sample pad, reaction membrane, and absorbing pad. The reagent pad contains colloidal gold conjugated with monoclonal antibodies (mAb) specific to SARS-CoV-2, Influenza A, and Influenza B target proteins. The reaction membrane contains different analyte specific antibodies to capture the target proteingold-mAb complexes at the respective test lines. Excess liquid and reagents are absorbed by the absorbing pad. The Healgen Rapid Check COVID-19/Flu A&B Antigen Test does not use biotin-Streptavidin/avidin chemistry in any of the steps for coupling reagents.

Mentions image processing

Not Found

Mentions AI, DNN, or ML

Not Found

Input Imaging Modality

Not Found

Anatomical Site

anterior nasal swab samples

Indicated Patient Age Range

individuals aged 14 years or older testing themselves, or adults testing individuals aged 2 years or older.

Intended User / Care Setting

non-prescription home use by individuals aged 14 years or older testing themselves, or adults testing individuals aged 2 years or older.

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

Not Found

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

Not Found

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

A multi-center, prospective clinical study was conducted with lay users to assess the performance of the Healgen Rapid Check COVID-19/Flu A&B Antigen Test in detecting nucleoprotein antigens extracted from COVID-19, influenza virus types A and B in selfcollected and self-tested anterior nasal swab samples. The study only enrolled subjects with two or more symptoms of respiratory infection consistent with COVID-19 or influenza. Ten clinical sites across the U.S. conducted the study from February to April 2024. Both the comparator and the candidate test used anterior nasal swab samples and the collection order was alternated by study subject. Comparator test samples were collected by health care professionals at the clinical study sites and inserted into Universal Transport Media per the IFU of the comparator test. Samples were then sent to a central site for testing with highly sensitive RT-PCR tests separately detecting SARS-CoV-2 and Flu A/B. Samples for the candidate antigen test were collected per the test's quick reference instructions and were either self-collected by a lay user aged >14 years or collected by an adult (parent/guardian) from individuals aged 2 to

Regulation Number and Section

N/A

Summary

0

Image /page/0/Picture/0 description: The image contains the logo of the U.S. Food and Drug Administration (FDA). The logo consists of two parts: the Department of Health & Human Services logo on the left and the FDA logo on the right. The FDA logo features the letters "FDA" in blue, followed by the words "U.S. FOOD & DRUG ADMINISTRATION" in a smaller font size.

EVALUATION OF AUTOMATIC CLASS III DESIGNATION FOR Healgen Rapid Check COVID-19/Flu A&B Antigen Test DECISION SUMMARY

I. Background Information:

A De Novo Number

DEN240029

B Applicant

Healgen

C Proprietary and Established Names

Healgen Rapid Check COVID-19/Flu A&B Antigen Test

D Regulatory Information

| Product
Code(s) | Classification | Regulation
Section | Panel |
|--------------------|----------------|---------------------------------------------------------------------------------|--------------|
| SCA | Class II | 21 CFR 866.3987 – Multi-
analyte respiratory virus
antigen detection test | Microbiology |

II. Submission/Device Overview:

A Purpose of Submission

De Novo request for evaluation of automatic class II designation for the Healgen Rapid Check COVID-19/Flu A&B Antigen Test

B Measurand

Influenza type A and type B nucleoprotein and SARS-CoV-2 nucleocapsid antigens

C Type of Test

Qualitative Lateral flow Immunoassay

III. Indications for Use:

A Intended Use(s):

See Indications for Use below

Food and Drug Administration 10903 New Hampshire Avenue Silver Spring, MD 20993-0002 www.fda.gov

1

B Indication(s) for Use:

The Healgen Rapid Check COVID-19/Flu A&B Antigen Test is a lateral flow immunochromatographic assay intended for the qualitative detection and differentiation of influenza A, and influenza B nucleoprotein antigens and SARS-CoV-2 nucleocapsid antigen directly in anterior nasal swab samples from individuals with signs and symptoms of respiratory tract infection. Symptoms of respiratory infections due to SARS-CoV-2 and influenza can be similar. This test is for non-prescription home use by individuals aged 14 years or older testing themselves, or adults testing individuals aged 2 years or older.

All negative results are presumptive and should be confirmed with an FDA-cleared molecular assay when determined to be appropriate by a healthcare provider. Negative results do not rule out infection with influenza, SARS-CoV-2 or other pathogens. Individuals who test negative and experience continued or worsening respiratory symptoms, such as fever, cough and/or shortness of breath, should therefore seek follow-up care from their healthcare provider.

Positive results do not rule out co-infection with other respiratory pathogens and therefore do not substitute for a visit to a healthcare provider or appropriate follow-up.

C Special conditions for Use Statement(s):

OTC - Over The Counter

D Special Instrument Requirements:

Not Applicable

IV. Device/System Characteristics:

A Device Description:

The Healgen Rapid Check COVID-19/Flu A&B Antigen Test is an immunochromatographic assay that uses highly sensitive monoclonal antibodies to detect nucleoprotein antigens from SARS-CoV-2, influenza virus types A and B in anterior nasal swab (ANS) samples from symptomatic individuals. The test device is composed of a plastic housing, known as a cassette that contains a test strip with the following parts: sample pad, reaction membrane, and absorbing pad. The reagent pad contains colloidal gold conjugated with monoclonal antibodies (mAb) specific to SARS-CoV-2, Influenza A, and Influenza B target proteins. The reaction membrane contains different analyte specific antibodies to capture the target proteingold-mAb complexes at the respective test lines. Excess liquid and reagents are absorbed by the absorbing pad. The Healgen Rapid Check COVID-19/Flu A&B Antigen Test does not use biotin-Streptavidin/avidin chemistry in any of the steps for coupling reagents.

B Principle of Operation:

When the test sample is added into the sample well (S) of the cassette, mAb conjugates dried in the reagent pad are dissolved and migrate along with the sample, across the reaction lines on the membrane. The reaction lines contain different antibodies that are also analyte specific and bind

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to the target protein-gold-mAb complexes and immobilize them on the membrane, resulting in a visible red test line.

Results completely develop after 15 minutes. Reactions for each virus occur independently at their respective locations on the test reaction membrane. If the sample contains influenza type A or B antigens, a pink-to-red test line (A or B) will develop; if SARS-CoV-2 antigens are present, a pink-to-red test line (T) will develop. The procedural control lines (C) must always appear. Healgen Rapid Check COVID-19/Flu A&B Antigen Test is validated for testing direct samples without transport media.

The technical principle for influenza A virus and influenza B virus is identical to that of the SARS-CoV-2 antigen test strip as shown in figure 1 below.

Image /page/2/Figure/3 description: The image shows a diagram of a COVID-19 test strip. The diagram illustrates how the test works with and without the presence of the COVID-19 antigen. The test strip includes a sample pad, conjugate pad, test line, control line, and absorbent pad. The diagram also shows the different components of the test, such as gold particles, COVID-19 antigens, gold-labeled COVID-19 specific antibodies, COVID-19 specific capture antibodies, and anti-mouse IgG.

Figure 1: Schematic of the Healgen Rapid Check COVID-19/Flu A&B Antigen Test Strip

Both the test strips enclosed in the test device independently feature an internal control, denoted directly on the test device as "C" for user interface. Test strip specific control lines are needed to indicate that each respective test strip is working adequately in each lay user performed test. The control line contains goat anti-mouse IgG antibodies that capture the excess gold-labeled mouse antibody preloaded in the reagent pad. The controls must be positive for a sample to provide a valid result to demonstrate that the test reagents are functional and correctly performed. If the control line is not detected, the sample result is invalid. Test results are displayed as Positive, Negative, or Invalid.

C Interpretation of Results:

The qualitative results of the Healgen Rapid Check COVID-19/Flu A&B Antigen Test are visually interpreted by the user. Examples of the positive, and invalid results interpretations are provided within the "Interpreting the Result" section of the QRI. Individuals can scan a QR code within the QRI. This code directs the test user to a complete list of test result interpretations prepared and provided online at: https://www.healgen.com/covid19-influenza-a-b. Results interpretation is described in the below figure.

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Visual for Positions of Result Lines
Image: test cassette	C = Control Line
Flu A, Flu B, COVID = Test Line indicating presence/absence of
Flu B = Influenza B
Flu A = Influenza A
COVID = SARS-CoV-2
Invalid (No Result)	Invalid
A valid result must display a pinkish-red Control Line in the control region 'C' of both the result windows. If no or only one window of the test cassette shows a pinkish-red Control Line in the control region 'C', the assay is invalid and cannot be interpreted no matter the presence of any positive Test lines in the cassette windows.

Invalid tests should be repeated with a new test. | Missing 'C' line on ONE or BOTH strips
Image: three invalid test cassettes
Note: The 3 images displayed are examples only; additional invalid outcomes are possible and can include test strips with or without test lines for Flu A, Flu B and/or COVID. |
| Negative Result | Valid Negative Result |
| A negative sample will produce a single pinkish-red Control Line in the control region 'C' of the window, indicating a negative result. This Control Line means that the detection part of the test was done correctly, sample was added, but no COVID-19 antigen was detected. | BOTH "C" lines must be PRESENT
Image: valid negative test cassette |
| Positive Result | Valid Positive Result |
| A positive specimen will produce a single pinkish-red Test Line and a single pinkish-red Control Line. This means that COVID-19 antigen was detected and that the detection part of the test performed correctly. Specimens with low levels of antigen may give a faint Test Line | BOTH "C" lines must be PRESENT
Image: text |

4

Image /page/4/Figure/0 description: The image shows a diagram of COVID-19 and Flu test results. There are six different test results shown, including COVID-19 Positive, Flu A Positive, Flu B Positive, COVID-19 & Flu A Positive, COVID-19 & Flu B Positive, COVID-19 & Flu A & Flu B Positive, and Flu A & Flu B Positive. Each test result shows the control line (C) and the test lines for Flu A, Flu B, and COVID-19. The positive results are indicated by the presence of a red line in the test line area.

Figure 2: Results Interpretation

Standards/Guidance Documents Referenced: V.

| Document Number | Title | Publishing
Organization |
|-------------------|--------------------------------------------------------------------------------------------------------------------------------------------------------------------------------|----------------------------|
| EP25-A | Evaluation of Stability of In Vitro Diagnostics Reagents | CLSI |
| EP05-A3 | Evaluation of Precision of Quantitative Measurement Procedures | CLSI |
| EP12-A2 | User Protocol for Evaluation of Qualitative Test Performance | CLSI |
| ISO 15223-1 | Medical Devices - Symbols to be used with information to be
supplied by the manufacturer | ANSI AAMI ISO |
| ISO 10993-5:2009 | Biological evaluation of medical devices - Part 5: Tests for in vitro
cytotoxicity | ANSI AAMI ISO |
| ISO 10993-10:2010 | Biological evaluation of medical devices - Part 10: Tests for irritation
and skin sensitization | ANSI AAMI ISO |
| ISO 11135:2014 | Sterilization of health-care products – Ethylene oxide – Requirements
for the development, validation and routine control of a sterilization
process for medical devices | ANSI AAMI ISO |
| ISO 10993-7 | Biological evaluation of medical devices -Part 7: Ethylene oxide
sterilization residuals. | ANSI AAMI ISO |

5

VI. Performance Characteristics:

A Analytical Performance:

1. Precision/Reproducibility:

The Precision study for the Healgen Rapid Check COVID-19/Flu A&B Antigen Test was evaluated in two different in-house studies using the same 3 lots of test kits and the same operators. The strains used for testing were PROtrol inactivated SARS-CoV-2 lineage BA.5; omicron variant, PROtrol inactivated influenza A/Guangdong-Maonan/SWL1536/19, and PROtrol inactivated influenza B/Washington/02/19.

Study 1 was conducted by 2 trained operators. Three sample levels (2X LoD co-spiked, 5X LoD co-spiked and Negative Pooled Nasal Wash) were tested on each day, one replicate per run, per operator, and per lot of devices. Two (2) runs (morning and afternoon) were conducted each day per operator, per lot, per day. This exact testing scheme was carried out over 10 days (same 3 sample levels tested, on the same 3 lots, by the same 2 operators, in 2 runs per day). This resulted in 120 total tests per sample level. All samples were randomized and blinded for each day. For all three lots and operators, the results for this study shown in the Table 1 below were identical and concordant with the expected results.

Study 2 was specifically conducted to further evaluate potential differences between lots. The study used negative samples (without virus analytes) and very low positive samples at 0.75x LoD, commonly referred to as high negative sample. Samples were prepared near the C95 concentration for all three analytes and were randomized and blinded. This supplemental precision testing was carried out over 3 days only, but otherwise followed the same study design as above. This resulted in 72 total tests per analyte and sample level (24 replicates for each analyte with each lot). Data from this testing are integrated into Table 1 below.

The random errors of the testing procedure across different days and runs, paired with an operator's ability to read the intensity for samples with very low analyte concentration (commonly referred to as 'high negative samples') is expected to confound lot-specific variability and to have a significant impact on the precision estimates for high negative samples such as the 0.75 x LoD sample tested in this second part of the precision assessment. This is supported by the stratified data for study 2 that demonstrated imprecision also between runs and days (data not shown).

Taken together, the results of both precision assessments demonstrate a test precision and a lotto-lot precision that are consistent with the expectations for the analyte concentration in the samples, the test's technology, and the test's LoD. The between-lot variability does not impact low concentrated samples equal to or above 2 x LoD of the test.

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		Lot 1		Lot 2		Lot 3		Total
Sample	Analyte	Count*	%
Agreement	Count*	%
Agreement	Count*	%
Agreement	Percent
Lot-to-Lot
Agreement	95% CI
Negative	SARS-
CoV-2	0/64	100%	0/64	100%	0/64	100%	100%	98.0-100%
	Flu A	0/64	100%	0/64	100%	0/64	100%	100%	98.0-100%
	Flu B	0/64	100%	0/64	100%	0/64	100%	100%	98.0-100%
0.75 x
LoD	SARS-
CoV-2	20/24	83.3%	22/24	91.7%	17/24	70.8%	81.9%	71.5-89.1%
	Flu A	15/24	62.5%	15/24	62.5%	15/24	62.5%	62.5%	50.9-72.8%
	Flu B	18/24	75.0%	17/24	70.8%	14/24	58.3%	68.0%	56.6-76.7%
2 x LoD	SARS-
CoV-2	40/40	100%	40/40	100%	40/40	100%	100%	93.9-100%
	Flu A	40/40	100%	40/40	100%	40/40	100%	100%	93.9-100%
	Flu B	40/40	100%	40/40	100%	40/40	100%	100%	93.9-100%
5 x LoD	SARS-
CoV-2	40/40	100%	40/40	100%	40/40	100%	100%	93.9-100%
	Flu A	40/40	100%	40/40	100%	40/40	100%	100%	93.9-100%
	Flu B	40/40	100%	40/40	100%	40/40	100%	100%	93.9-100%

Table 1: Summary Results for Lot-Lot Precision study (Operators 1 and 2 Combined)

The total number of replicates included in this table is different sample concentrations due to the study being performed in two parts. Please refer to the study description above for more information.

2. Linearity:

This is a qualitative test without numerical data output and linearity is not applicable.

3. Analytical Specificity/Interference:

Cross-reactivity and Microbial Interference:

Cross Reactivity and Microbial Interference studies were conducted to determine if other respiratory pathogens/flora that could be present in a direct nasal swab samples could cause a false-positive test result or interfere with a true positive result. A panel of viruses, bacteria, fungi, and pooled nasal wash was used for these studies. Final target organism concentrations were 1.0 x 105 PFU/mL/1 x 105 TCID50/mL for viruses, and 1.0 x 106 cfu/mL for bacteria and fungi. When the target concentration was not achievable due to the stock culture, the highest concentration possible was tested without dilution. Dilutions for cross-reactivity testing were made in pooled negative swab matrix in saline (NCM Saline). Each organism was tested in replicates of three (3) without SARS-CoV-2/ FluA/FluB present in the sample. All testing was randomized and blinded.

Organisms that did not cause a false-positive result were further evaluated for microbial interference by testing PNW spiked with low-level UV inactivated SARS-CoV-2, live Flu A virus, and live Flu B virus isolate (3X single analyte LoD) in the presence of potentially interfering organism at a high titer in triplicate. If interference was observed at the level tested, an additional titration study was performed to determine the highest microorganism concentration that does not produce interference with the Healgen Rapid Check COVID-19/Flu A&B Antigen test device.

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Neither cross-reactivity nor interference was observed for any of the organisms at the concentrations tested with the Healgen Rapid Check COVID-19/Flu A&B Antigen test device.

The summary of cross-reactivity and microbial interference results are shown in the table below.

	Concentrations		Cross-	Microbial
Organism	Tested	Units	Reactivity	Interference
SARS-CoV-1	1.25E+05	PFU/ml	ND*	ND
MERS-coronavirus	1.47E+05	TCID50/mL	ND	ND
Human coronavirus OC43	7.00E+05	TCID50/mL	ND	ND
Human coronavirus 229E	1.58E+05	TCID50/mL	ND	ND
Human coronavirus NL63	8.00E+04	TCID50/mL	ND	ND
Human coronavirus HKU1 a	1:20 dilution	NA	NA	ND
Adenovirus, Type 1 (Adenoid 71)	2.23E+05	TCID50/mL	ND	ND
Adenovirus Type 7, Type 7A
(Species B)	1.58E+05	TCID50/mL	ND	ND
Cytomegalovirus, Strain AD-169	7.05E+04	TCID50/mL	ND	ND
Epstein Barr Virus, Strain B95-8	1.83E+06	CP/mL	ND	ND
Human Metapneumovirus (hMPV),
Strain TN/91-316	3.50E+05	TCID50/mL	ND	ND
Parainfluenza virus 1,
Strain FRA/29221106/2009	2.00E+05	TCID50/mL	ND	ND
Parainfluenza virus 2, Strain Greer	1.75E+05	TCID50/mL	ND	ND
Parainfluenza virus 3, Strain C243	7.00E+05	TCID50/mL	ND	ND
Parainfluenza virus 4, Strain N/A	2.39E+05	TCID50/mL	ND	ND
Enterovirus Type (e.g. 68),
Species D Type 68	2.23E+05	TCID50/mL	ND	ND
Respiratory syncytial virus A,
Strain A-2	3.50E+05	TCID50/mL	ND	ND
Respiratory syncytial virus B,
Strain CH93(18)-18	2.29E+05	TCID50/mL	ND	ND
Rhinovirus 1A, Strain N/A	7.05E+04	TCID50/mL	ND	ND
Bordetella pertussis, Strain A639	2.50E+08	CFU/mL	ND	ND
Candida albicans, Strain Z006	6.03E+06	CFU/mL	ND	ND
Chlamydia pneumoniae, Strain Z500	4.33E+06	IFU/mL	ND	ND
Corynebacterium xerosis	2.30E+07	CFU/mL	ND	ND
Escherichia coli, Strain mcr-1	1.79E+08	CFU/mL	ND	ND
Hemophilus influenzae, type b;
Eagan	9.68E+06	CFU/mL	ND	ND
Lactobacillus sp., Lactobacillus
Acidophilus, Strain Z048	1.21E+07	CFU/mL	ND	ND
Legionella spp pneumophila,
Strain Philadelphia-1	6.50E+06	CFU/mL	ND	ND
Moraxella catarrhalis, Strain 59632	2.50E+08	CFU/mL	ND	ND
Mycoplasma pneumoniae,
Strain PI 1428	2.50E+07	CFU/mL	ND	ND
Mycobacterium tuberculosis
avirulent, Strain H37Ra-1	4.15E+06	CFU/mL	ND	ND
Neisseria meningitidis, serogroup A	3.43E+06	CFU/mL	ND	ND
Neisseria sp. Elongata Z071	2.68E+08	CFU/mL	ND	ND
Pneumocystis jirovecii,	1.30E+07	CFU/mL	ND	ND
Organism	Concentrations
Tested	Units	Cross-
Reactivity	Microbial
Interference
Strain W303-Pji
Pseudomonas aeruginosa, Strain N/A	3.45E+08	CFU/mL	ND	ND
Staphylococcus aureus Protein A
producer, e.g., Cowan strain, NCTC
8530 [S11]; Cowan's serotype 1	2.60E+08	CFU/mL	ND	ND
Staphylococcus epidermidis (PCI

                                                           | 9.00E+07                 | CFU/mL    | ND                   | ND                        |

| Streptococcus salivarius,
Strain C699 [S30D] | 1.01E+06 | CFU/mL | ND | ND |
| Streptococcus pneumoniae,
Strain Z022 | 1.81E+07 | CFU/mL | ND | ND |
| Streptococcus pyogenes,
Strain MGAS 8232 | 7.50E+07 | CFU/mL | ND | ND |
| Measles, Strain Edmonston | 8.48E+05 | TCID50/mL | ND | ND |
| Mumps (Isolate 1) | 8.48E+05 | TCID50/mL | ND | ND |

Table 2:Summary of Cross-reactivity and Microbial Interference Results

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*ND - Not Detected

a1:10 dilution of cultured stock HKU1 sample from Emory

Competitive Interference:

Competitive interference of the test's analytes was tested with different combinations of low (3x LoD) and high concentrations of Flu A, Flu B and SARS-CoV-2 spiked together onto a swab and then tested with one lot of Healgen Rapid Check COVID-19/Flu A&B Antigen test device. The study used inactivated SARS-CoV-2 but live influenza A and B virus strains.

The table below summarizes the results of the competitive interference study. For each condition tested all three replicates tested at the low target analyte condition tested positive in the presence of a second target analyte at high concentrations. No false positive results were observed for analytes not present in the sample.

	Analyte Concentration Added to Sample*
	(No. positive replicates / No. of total replicates)
	Flu A	Flu B	SARS-CoV-2
Analyte Concentration Added
Results	High
3/3	Low
3/3	-
0/3
Analyte Concentration Added
Results	High
3/3	-
0/3	Low
3/3
Analyte Concentration Added
Results	Low
3/3	High
3/3	-
0/3
Analyte Concentration Added
Results	-
0/3	High
3/3	Low
3/3
Analyte Concentration Added
Results	Low
3/3	-
0/3	High
3/3
Analyte Concentration Added
Results	-
0/3	Low
3/3	High
3/3

Table 3: Competitive Interference Results Summary

SARS-CoV-2 strain - 1X LoD - 3.95E+02 TCID50/mL

Flu A - H3N2:A/Darwin/6/2021 - 1X LoD - 2.09E+02 TCIDso/mL

Flu B - Yamagata: B/Florida/4/2006 - 1X LoD-1.46E+01 TCID50/mL

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Exogenous and Endogenous Interference Study

The Healgen Rapid Check COVID-19/Flu A&B Antigen test was evaluated for performance in the presence and absence of potentially interfering substances that might be present in a respiratory specimen. Interfering substances testing was performed using a panel of endogenous and exogenous substances tested at concentrations listed in the below table.

Negative specimens were evaluated in triplicates to confirm that the potentially interfering substances would not cause false positive results with the test.

Negative clinical matrix (pooled nasal wash) was co-spiked with SARS-CoV-2 USA WA1/2020, Flu A H1N1pdm09/A Victoria/4897/2022, and Flu B Yamagata/B/Florida/4/2006, and then mixed 1:1 with interfering substance. Final concentration for each analyte was 3x LoD (based on the established single analyte LoD). Negative nasal wash has been demonstrated to be equivalent to the anterior nasal swab matrix in a matrix equivalency study. Testing was performed in triplicate to confirm that SARS-CoV-2, Flu A and Flu B could still be detected if the test substances were present in the sample. All testing was randomized and blinded. Test results are summarized in the table below.

With the exception of Flu Mist Quadrivalent live influenza vaccine, none of the substances caused a false-positive test result in unspiked samples. While the presence of Flu Mist Quadrivalent live influenza vaccine at 15% v/v concentration did not interfere with the detection of true positive results of the 3x LoD co-spiked samples, the vaccine also resulted in positive results for Flu A and Flu B (as expected based on the composition of the vaccine). Hand sanitizer cream lotion and hand sanitizer 80% ethanol fast drying at 15% v/v showed false negative results for Flu B, but detected all analytes at 7.5% v/v.

| Interfering Substance | Concentration | Cross-Reactivity
(no analyte)
(# pos/ # total) | | | Interference
(3x co-spiked analyte LoD)
(# pos/ # total) | | |
|--------------------------------------------------|-------------------------|------------------------------------------------------|-------|-------|----------------------------------------------------------------|-------|-------|
| | | SARS-
CoV-2 | Flu A | Flu B | SARS-
CoV-2 | Flu A | Flu B |
| Human Whole Blood
(EDTA tube) | 4% v/v | 0/3 | 0/3 | 0/3 | 3/3 | 3/3 | 3/3 |
| Leukocytes | 1.67 x 10^6
cells/mL | 0/3 | 0/3 | 0/3 | 3/3 | 3/3 | 3/3 |
| Throat Lozenges
(Menthol/Benzocaine) | 3 mg/mL | 0/3 | 0/3 | 0/3 | 3/3 | 3/3 | 3/3 |
| Mucin, bovine
submaxillary gland | 2.5 mg/mL | 0/3 | 0/3 | 0/3 | 3/3 | 3/3 | 3/3 |
| Zinc (Therazinc throat
Spray) | 15% v/v | 0/3 | 0/3 | 0/3 | 3/3 | 3/3 | 3/3 |
| Naso GEL (NeilMed) | 15% v/v | 0/3 | 0/3 | 0/3 | 3/3 | 3/3 | 3/3 |
| Nasal Drops
(Phenylephrine) | 15% v/v | 0/3 | 0/3 | 0/3 | 3/3 | 3/3 | 3/3 |
| Nasal Spray
(Oxymetazoline) | 15% v/v | 0/3 | 0/3 | 0/3 | 3/3 | 3/3 | 3/3 |
| Nasal Spray (Cromolyn) | 15% v/v | 0/3 | 0/3 | 0/3 | 3/3 | 3/3 | 3/3 |
| Nasal Corticosteroid
(Dexamethasone) | 1 mg/mL | 0/3 | 0/3 | 0/3 | 3/3 | 3/3 | 3/3 |
| Nasal Corticosteroid
(Fluticasone Propionate) | 15% v/v | 0/3 | 0/3 | 0/3 | 3/3 | 3/3 | 3/3 |

Table 4: Interfering Substances Study Results

10

| Interfering Substance | Concentration | Cross-Reactivity
(no analyte)
(# pos/ # total) | | | Interference
(3x co-spiked analyte LoD)
(# pos/ # total) | | |
|------------------------------------------------------------------------------------------|---------------|------------------------------------------------------|-------|-------|----------------------------------------------------------------|-------|-------|
| | | SARS-
CoV-2 | Flu A | Flu B | SARS-
CoV-2 | Flu A | Flu B |
| Nasal gel (Galphimia
glauca, Histanium
hydrocloricum, Luffa
operculate, Sulfur) | 1.25% | 0/3 | 0/3 | 0/3 | 3/3 | 3/3 | 3/3 |
| Homeopathic allergy
relief (Histaminum
hydrochloricum) | 15% v/v | 0/3 | 0/3 | 0/3 | 3/3 | 3/3 | 3/3 |
| Zicam nasal spray
(Galphimia glauca, Luffa
operculata) | 15% v/v | 0/3 | 0/3 | 0/3 | 3/3 | 3/3 | 3/3 |
| Nasal spray (Alkalol) | 15% v/v | 0/3 | 0/3 | 0/3 | 3/3 | 3/3 | 3/3 |
| Sore Throat Phenol Spray | 15% v/v | 0/3 | 0/3 | 0/3 | 3/3 | 3/3 | 3/3 |
| Tobramycin | 4 $μ$ g/mL | 0/3 | 0/3 | 0/3 | 3/3 | 3/3 | 3/3 |
| Mupirocin | 10 mg/mL | 0/3 | 0/3 | 0/3 | 3/3 | 3/3 | 3/3 |
| Anti-viral drug
(Remdesvir) | 10 mg/mL | 0/3 | 0/3 | 0/3 | 3/3 | 3/3 | 3/3 |
| Tamiflu (Oseltamivir) | 5 mg/mL | 0/3 | 0/3 | 0/3 | 3/3 | 3/3 | 3/3 |
| FluMist | 15% v/v | 0/3 | 3/3 | 3/3 | 3/3 | 3/3 | 3/3 |
| (Quadrivalent/Live) | 0.15% v/v | 0/3 | 0/3 | 0/3 | NA | NA | NA |
| Zanamivir | 282 ng/mL | 0/3 | 0/3 | 0/3 | 3/3 | 3/3 | 3/3 |
| Biotin | 3500 ng/mL | 0/3 | 0/3 | 0/3 | 3/3 | 3/3 | 3/3 |
| Body & Hand Lotion | 0.5% w/v | 0/3 | 0/3 | 0/3 | 3/3 | 3/3 | 3/3 |
| Body Lotion, with 1.2%
dimethicone | 0.5% w/v | 0/3 | 0/3 | 0/3 | 3/3 | 3/3 | 3/3 |
| Hand Lotion | 5% w/v | 0/3 | 0/3 | 0/3 | 3/3 | 3/3 | 3/3 |
| Hand Sanitizer with Aloe,
62% ethyl alcohol | 5% w/v | 0/3 | 0/3 | 0/3 | 3/3 | 3/3 | 3/3 |
| Hand Sanitizer cream | 15% v/v | 0/3 | 0/3 | 0/3 | 3/3 | 3/3 | 0/3 |
| lotion | 7.5% v/v | 0/3 | 0/3 | 0/3 | 3/3 | 3/3 | 3/3 |
| Hand Sanitizer, 80% | 15% v/v | 0/3 | 0/3 | 0/3 | 3/3 | 3/3 | 0/3 |
| ethanol | 7.5% v/v | 0/3 | 0/3 | 0/3 | 3/3 | 3/3 | 3/3 |
| Hand soap
liquid gel | 10% w/v | 0/3 | 0/3 | 0/3 | 3/3 | 3/3 | 3/3 |

4. Assay Reportable Range:

This section is not applicable as this device is a qualitative assay.

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

Internal Controls:

Both the test strips enclosed in the test device independently feature an internal control, denoted directly on the test device as "C" for user interface. Test strip specific control lines are needed to indicate that each respective test strip is working adequately in each lay user performed test. The control line contains IgG antibodies that capture the excess gold-labeled mouse antibody preloaded in the reagent pad. These controls have to be positive for all valid test results to

11

demonstrate that the test reagents are functional, and the tests correctly performed. If the control lines are not detected, the sample result is invalid.

External Controls:

External Control testing is not performed by lay users and is therefore not applicable to OTC tests. External controls are therefore not included in the test kit.

Stability

Real Time Stability:

Three lots of the Healgen Rapid Check COVID-19/Flu A&B Antigen Test kits were subjected to a summer profile (40°C for 8 hours and then 30°C for 4 hours) and then a winter profile (- 10°C for 8 hours and then 18°C for 4 hours) to simulate the anticipated shipping/handling times and temperatures expected for unopened kits, after which they were stored at 2-8℃ or 30+3℃, respectively. The test panel comprised of negative clinical matrix, 2x LoD and 5x LoD of inactivated SARS-CoV-2, and live Flu A and Flu B viruses. Testing was performed at time 0 (baseline) and month 1, 3, 6, 9, 12, 15, and 18. Testing will continue for months 21, 24 and 27. All study data are 100% concordant with expected results and support a shelf-life of up to 15 months. The shelf life will be updated as additional passing time points will become available.

Open Kit Stability Study:

In this study, the amount of time a test device can be left outside of its packaging was assessed using a test panel comprised of five (5) negative samples (clinical matrix: PNW) and five (5) cospiked low positive samples (2X single analyte LoD of SARS-CoV-2, Flu A, and Flu B cospiked together into PNW). PNW was demonstrated to be equivalent to negative nasal swab matrix in a matrix equivalency study. Device packaging was opened and testing was performed at zero (0) hours to establish baseline. Thereafter, devices were stored for one-hour and twohour, respectively at 30±1℃ (the worst-case condition for a room temperature claim). All study data before and after storage of the open kits were 100% concordant with the expected results.

Transport Stability:

Simulated winter and summer transport temperature conditions were used to evaluate the worstcase shipping and handling of unopened components of the Healgen COVID-19/Flu A&B Ag Combo Rapid test over an extended period. The functional performance of Healgen's test device is assessed by comparing the pre- (T0) and post-distribution (Td) results of a test panel comprised of pooled negative nasal wash (PNW) samples and co-spiked low positive samples (3X single analyte LoD with SARS-CoV-2, Flu A, and Flu B, together contrived in PNW). PNW was demonstrated to be an equivalent negative clinical matrix to negative nasal swab matrix in a matrix equivalency study. All results were as expected for all time points.

6. Detection Limit:

Single Analyte LoD:

The LoD of the device was performed to determine the lowest detectable concentration of SARS-CoV-2, influenza A and influenza B at which at least 95% of all true positive replicates are consistently detected as positive. The LoD was assessed for each analyte in two parts, a preliminary range finding study, followed by a confirmatory LoD study. A preliminary LoD was

12

determined by first testing serial ten-fold dilutions of live influenza A and B, and inactivated SARS-CoV-2 virus stocks diluted into either pooled negative swab matrix (PNSM) or pooled nasal wash (PNW) in 3 replicates per dilution. Single analyte virus dilutions (50 µL/swab) were each spiked onto dry sterile swabs and tested per the IFU.

The preliminary LoD results for each individual virus strain is shown in below tables.

Table 5: Preliminary LoD - SARS-CoV-2

| Isolate/Lineage | (TCID50/mL) | SARS-CoV-2
(TCID50/Swab) | #Positive/#
Total | # of device
lots tested |
|---------------------------------------------------|--------- | | 3.16E+05 | | 3.16E+04 | | 3.16E+03 | | 1.58E+03 | USA-WA1-2020 (UV inactivated) | 7.90E+02 | | 3.95E+02 | | 3.16E+02 | | 1.58E+02 | | 3.09E+06 | | 3.09E+05 | | 3.09E+04 | USA-WA1-2020 (Heat inactivated) | 3.09E+03 | | 1.5E+03 | | 3.09E+02 | | 2.19E+03 | USA/COR-22-063113/2022 (BA.5,
Omicron variant) | 1.09E+03 | 5.45E+01 | | 5.47E+02 | | 2.19E+02 SARS-CoV-2
------------------|-----------------------------|----------------------|----------------------------|
| 1.58E+04 | 3/3 | |
| 1.58E+03 | 3/3 | |
| 1.58E+02 | 3/3 | |
| 7.90E+01 | 3/3 | |
| 3.95E+01 | 3/3 | 1 |
| 1.98E+01 | 3/3 | |
| 1.58E+01 | 2/3 | |
| 7.90E+00 | 0/3 | |
| 1.5E+05 | 3/3 | |
| 1.5E+04 | 3/3 | |
| 1.5E+03 | 3/3 | 1 |
| 1.5E+02 | 3/3 | |
| 7.5E+01 | 1/3 | |
| 1.5E+01 | 0/3 | |
| 1.09E+02 | 9/9 | |
| 9/9 | 3 |
| 2.73E+01 | 4/9 | |
| 1.09E+01 | 2/3 | |

Table 6: Preliminary LoD - Influenza A

Total | # of device

lots tested |
|-----------------|--------------------------------|---------------------------|-----------------------------|-----------------------|----------------------------|
| | | 4.17E+04 | 2.09E+03 | 3/3 | |
| | | 4.17E+03 | 2.09E+02 | 3/3 | |
| | | 4.17E+02 | 2.09E+01 | 3/3 | |
| H3N2 | A/Darwin/6/2021 | 2.09E+02 | 1.05E+01 | 3/3 | 1 |
| | | 1.04E+02 | 5.20E+00 | 3/3 | |
| | | 5.21E+01 | 2.61E+00 | 3/3 | |
| | | 4.17E+01 | 2.09E+00 | 2/3 | |
| | | 4.17E+04 | 2.09E+03 | 0/3 | |
| | | 2.02E+04 | 1.01E+03 | 3/3 | |
| | | 2.02E+03 | 1.01E+02 | 3/3 | |
| | | 2.02E+02 | 1.01E+01 | 3/3 | |
| H1N1 | pdm09:A/Victoria/48
97/2022 | 1.01E+02 | 5.05E+00 | 1/3 | 1 |
| | | 5.05E+01 | 2.53E+00 | 0/3 | |
| | | 2.53E+01 | 1.27E+00 | 0/3 | |
| | | 2.02E+01 | 1.01E+00 | 0/3 | |

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Total | # of device

lots tested |
|-----------------|-------------------------------------|---------------------------|-----------------------------|-----------------------|----------------------------|
| | A/California/07/2009
pdm09 | 2.1E+05 | 5.85E+02 | 3/3 | 1 |
| | | 2.1E+04 | 5.85E+01 | 3/3 | |
| | | 2.1E+03 | 5.85E | 3/3 | |
| | | 1.05E+03 | 5.25E+01 | 3/3 | |
| | | 5.25E+02 | 2.6E+01 | 1/3 | |
| | | 2.1E+02 | 1.05E+01 | 0/3 | |
| | Guangdong-
Maonan/SWL
1536/19 | 5.62E+01 | 2.81 | 9/9 | 3 |
| | | 5.62 | 2.81E-01 | 0/3 | |

Table 7: Preliminary LoD - Influenza B

| Isolate/Lineage | Strain | SARS-CoV-2
(TCID50/
mL) | SARS-CoV-2
(TCID50/Swab) | #Positive/#
Total | # of device lots
tested |
|-----------------|-----------------------------------------------|-------------------------------|-----------------------------|----------------------|----------------------------|
| Yamagata | B/Florida/4/2006 | 1.17E+04 | 5.85E+02 | 3/3 | |
| | | 1.17E+03 | 5.85E+01 | 3/3 | |
| | | 1.17E+02 | 5.85E+00 | 3/3 | |
| | | 5.85E+01 | 2.93E+00 | 3/3 | 1 |
| | | 2.93E+01 | 1.47E+00 | 3/3 | |
| | | 1.46E+01 | 7.30E-01 | 3/3 | |
| | | 1.17E+01 | 5.85E-01 | 1/3 | |
| | B/Washington/02/2019 | 3.16E+05 | 1.58E+04 | 3/3 | |
| | | 3.16E+04 | 1.58E+03 | 3/3 | |
| | | 3.16E+03 | 1.58E+02 | 3/3 | |
| | | 1.58E+03 | 7.90E+01 | 1/3 | 1 |
| | | 7.90E+02 | 3.95E+01 | 0/3 | |
| | | 3.95E+02 | 1.98E+01 | 0/3 | |
| | | 3.16E+02 | 1.58E+01 | 0/3 | |
| Victoria | | 1.75E+04 | 8.75E+02 | 9/9 | |
| | B/Washington/02/2019
(PROtrol inactivated) | 8.75E+03 | 4.37E+02 | 0/3 | 3 |
| | | 1.75E+03 | 8.75E+01 | 0/3 | |
| | | | | | |
| | B/Florida/78/2015 | 1.7E+06 | 8.5E+04 | 3/3 | |
| | | 1.7E+05 | 8.5E+03 | 3/3 | |
| | | 1.7E+04 | 8.5E+02 | 3/3 | 1 |
| | | 8.5E+03 | 4.25E+02 | 1/3 | |
| | | 1.7E+03 | 8.5E+01 | 0/3 | |

LoD confirmatory testing was then performed individually for each virus by testing 20 replicates at the virus' preliminary (1X) LoD concentration, as determined above. For the LoD to be confirmed, at least 95% of the replicates (≥19/20) needed to test positive. Results of the LoD confirmation testing for each virus are summarized in the table below.

14

| Analyte | Isolate/
Lineage | Strain | LoD
Concentration
(TCID50/mL) | LoD
Concentration
(TCID50/swab) | #Positive
/# Total | #
device
lots
tested |
|----------------|-------------------------------------------------------|--------------------------------------------------------------|-------------------------------------|---------------------------------------|-----------------------|-------------------------------|
| | USA-WA1/2020
(UV inactivated) | NA | 3.95E+02 | 1.98E+01 | 20/20 | 1 |
| SARS-
CoV-2 | USA-WA1/2020
(Heat inactivated) | NA | 3.09E+03 | 1.5E+02 | 60/60 | 3 |
| | USA/COR-22-
063113/2022 (BA.5,
Omicron variant) | NA | 1.09E+03 | 5.45E+01 | 58/60 | 3 |
| Flu A | H3N2 | Darwin/6/21 | 2.09E+02 | 1.05E+01 | 20/20 | 1 |
| | H1N1 | Victoria/4897/22 | 2.02E+02 | 1.01E+01 | 20/20 | 1 |
| | | A/California/07/200
9 pdm09 | 1.05E+03 | 5.25 | 60/60 | 3 |
| | | Guangdong-
Maonan/SWL
1536/19 (PROtrol
inactivated) | 5.62E+01 | 2.81 | 60/60 | 3 |
| | Yamagata | Florida/04/06 | 1.46E+01 | 7.30E-01 | 20/20 | 1 |
| Flu B | Victoria | Washington/02/19 | 1.58E+03 | 7.90E+01 | 20/20 | 1 |
| | Victoria | Washington/02/19
(PROtrol
inactivated) | 1.75E+04 | 8.75E+02 | 58/60 | 3 |
| | Victoria | B/Florida/78/2015 | 1.7E+04 | 8.5E+02 | 60/60 | 3 |

Table 8: Confirmatory LoD

Co-spiked LoD:

After the single analyte LoDs were established for the candidate device, co-spiked LoD equivalency testing with all three test analytes present in the sample, was conducted to characterize performance with samples that contain more than one analyte at low concentrations. All analytes that are successfully detected by the candidate device when co-spiked at their single analyte LoD, may be co-spiked into positive sample/s used in the analytical studies.

Based on the individual analyte specific 1x LoDs, co-spiked samples were prepared by mixing all three viruses (one strain each of SARS-CoV-2, Flu A and Flu B). The 1x co-spiked LoD concentration was tested with the candidate device in twenty (20) replicates and was considered confirmed (i.e., equivalent to the established single analyte LoD) if ≥19/20 replicates were positive for concentrations within 2x LoD of the established single analyte LoD.

The Healgen Rapid check COVID-19/Flu A&B Antigen Test demonstrated co-spike equivalency for all analytes, SARS-CoV-2, Flu A and Flu B, to their respective established single analyte 1X LoD. The summary of the co-spike LoD is shown in the below table.

15

| Virus | Fold LoD | LoD Concentration
(TCID50/mL) | LoD Concentration
per Swab
(TCID50/swab) | # Positive
Replicates |
|--------------------------------------------|----------|----------------------------------|------------------------------------------------|--------------------------|
| SARS-CoV-2
(USA-WA1/2020) | 1X | 3.95 x 102 | 2.0 x 101 | 20/20 |
| Flu A H1N1
(pdm09:A/Victoria/4897/2022) | 1X | 2.02 x 102 | 1.0 x 101 | 20/20 |
| Flu B Yamagata
(B/Florida/4/2006) | 1X | 1.46 x 101 | 7.3 | 20/20 |

Table 9: Summary of Co-Spike Equivalency LoD Results

NIBSC 21/368 -WHO International Standard:

The sponsor tested the sensitivity of the test with the 1st WHO International Standard for SARS-CoV-2 antigen (NIBSC code: 21/368) spiked into pooled negative swab matrix (PNSM). The unitage of this material has an assigned value of 5,000 International Units of SARS-CoV-2 antigen per ampoule when reconstituted per instructions. A 2-fold dilution series was made to determine the preliminary LoD, which was measured using one device lot and triplicate measurements (n=3). The measurements were done by adding 50μl of each dilution directly to the test swab and processing the sample per the test's ORI. The preliminary LoD was determined to be 250 IU/ml (or 12.5 IU/swab).

The LoD confirmatory study was performed using 20 replicates (n=20) per dilution. The lowest concentration at which a minimum of 95% of results were positive was confirmed to be 250 IU/ml or 12.5 IU/Swab as shown below.

Table 10: LOD with the 1st WHO International Standard for SARS-CoV-2 Antigen (NIBSC
code:21/368)

Dilution (IU/ml)	Preliminary LoD		Confirmatory LoD
	Dilution
(IU/swab)	Results	Dilution
(IU/ml)	Dilution
(IU/swab)	Results
$4x10^3$	200	3/3
$2x10^3$	100	3/3
$1x10^3$	50	3/3
$5x10^2$	25	3/3	$5x10^2$	25	20/20
$2.5x10^2$	12.5	3/3	$2.5x10^2$	12.5	19/20
$1.25x10^2$	6.25	2/3	$1.25x10^2$	6.25	10/20
$6.25x10^1$	3.125	0/3

High-dose Hook Effect Study:

The hook effect study was conducted to evaluate if high levels of antigen present in the sample could result in a false negative test result. In this study, 50uL of the highest concentration possible for UV inactivated SARS-CoV-2 virus stock and for live influenza A and influenza B virus stocks were spiked onto sterile swabs for triplicate measurements, and swabs were tested on the device per IFU of the candidate device.

Testing showed no hook effect for SARS-CoV-2, Flu A, Flu B at the concentrations listed in the table below.

16

Tested |

|-------------|----------------------|----------------------|----------------------------------------|-----------------------------------------|-------------------------|
| SARS-CoV-2 | USA-WA1/2020 | N/A | 3.16E+06 | 1.58E+05 | 3/3 |
| Influenza A | A/Victoria/4897/2022 | H1N1 | 2.02E+05 | 1.01E+04 | 3/3 |
| Influenza A | A/Darwin/6/2021 | H3N2 | 4.17E+05 | 2.09E+04 | 3/3 |
| Influenza B | B/Washington/02/2019 | Victoria | 3.16E+06 | 1.58E+05 | 3/3 |
| Influenza B | B/Florida/4/2006 | Yamagata | 1.17E+05 | 5.85E+03 | 3/3 |

Table 11: Summary of High Dose Hook Effect Results

*Concentration in the sample solution applied to dry swab

Inclusivity Study:

Inclusivity testing was conducted to determine if the candidate device can detect different strains of SARS-CoV-2, Flu A and Flu B.

The Healgen Rapid Check COVID-19/Flu A&B Antigen Test employs the identical test strip as included in the previously cleared COVID-19-only Healgen Rapid COVID-19 Antigen Test (K232377). Inclusivity data for SARS-CoV-2 as obtained for K232377 therefore apply equally to the Healgen Rapid Check COVID-19/Flu A&B Antigen Test. Data derived from testing of commercially obtained Alpha (B.1.1.7), delta (B.1.617.2), omicron (B.1.1.529), beta (B.1.351), gamma (P1) and kappa (B.1.617.1) SARS-CoV-2 virus strains demonstrated the test strip's inclusivity of all tested variants at low concentrations (see FDA's Decision Summary for K232377 (fda.gov)).

A selection of temporal, geographic and genetically diverse Influenza A and B strains were tested on the Healgen Rapid Check COVID-19/Flu A&B Antigen Test for inclusivity. A series of ten-fold dilutions of each virus strain was made in pooled negative nasal matrix (PNSM). For each replicate tested in the study, 50uL of the dilution was pipetted on a fresh sterile swab. Once the ten-fold breakpoint was established for each of the strains, an additional series of three twofold dilutions was made from the lowest positive ten-fold dilution of each virus and triplicates were tested to demonstrate inclusivity. Contemporary strains (within the past 5 years) were prioritized over older strains.

The lowest concentrations that tested positive for relevant influenza virus strains by the candidate device are shown in the table below.

Virus	Virus Strains	Concentration	Units
Flu A - H1N1	A/ California/04/2009	2.80E+03	TCID50/mL
	A/ Brisbane/02/2018	1.51E+02	TCID50/mL
	A/ Michigan/45/2015	9.30E+00	TCID50/mL
	A/ Guangdong-Maonan/SWL
1536/2019	1.04E+03	TCID50/mL
	A/ NY/03/2009	2.29E+04	TCID50/mL
	A/ Indiana/02/2020	9.70E+06	CEID50/mL
	A/Wisconsin/588/2019	1.4E+04	FFU/mL
	A/ Sydney/5/2021	4.80E+03	TCID50/mL
	A/ Hawaii/66/2019	3.70E+07	CEID50/mL
	A/ Wisconsin/67/2022	1.05E+03	TCID50/mL
	A/New York/21/2020	2.6E+05	FFU/mL

Table 12: Inclusivity Results - Minimal Detectable Concentrations of Flu Variants

17

Virus	Virus Strains	Concentration	Units
Flu A – H3N2	A/Tasmania/503/2020	6.5E+04	FFU/mL
	A/Hong Kong/2671/2019	3.1E+06	CEID50/mL
	A/Hong Kong/45/2019	1.5E+04	FFU/mL
	A Alaska/01/2021	1.50E+04	FFU/mL
Flu A– H1N1	A/Indiana/08/2011	8.10E+02	TCID50/mL
Flu A– H1N2	A/Ohio/09/2015	7.0E+05	CEID50/mL
Flu A- H5N1	A/Minnesota/19/2011	8.00E+06	CEID50/mL
	A/mallard /Wisconsin/2576/2009	2.10E+05	GE/mL
	A/mallard /Wisconsin/2576/2009	800,000	CEID50/mL
	A/Bovine/Ohio/B24OSU-
439/2024	1,550	TCID50/mL
Flu A- H5N6	A/duck/Guangxi/S11002/2024	3.38E+05	EID50/mL
Flu A- H5N8	A/duck/Guangxi/S10888/2024	7.90E+05	EID50/mL
Flu A- H7N3	A/goose/Liaoning/S1266/2021	1.69E+05	EID50/mL
	A/northern
pintail/Illinois/10OS3959/2010	7.0E+05	CEID50/mL
Flu B -Victoria Lineage	B/ Brisbane/60/2008	6.45E-01	TCID50/mL
	B/Colorado/6/2017	5.85E+00	TCID50/mL
	B/Texas/02/2013	6.13E+00	TCID50/mL
	B/ Michigan/01/2021	2.85E+03	TCID50/mL
Flu B - Yamagata Lineage	B/Texas/06/2011	8.00E+05	CEID50/mL
	B/Utah/09/2014	1.26E+02	TCID50/mL
	B/Wisconsin/1/10	1.78E+01	TCID50/mL
Flu B - non-Victoria,
non-Yamagata	B/Maryland/1/1959	1.69E+03	CEID50/mL

Assay Cut-Off:

Not applicable as this is a qualitative visually read assay without numeric raw data.

B Comparison Studies:

Method Comparison:

Please refer to section VI.C (Clinical Studies) below for the clinical validation, regarding the method comparison studies.

Matrix Comparison:

The candidate device is only intended for qualitative detection of nucleocapsid protein antigen from SARS-CoV-2, and nucleoprotein from Flu A and Flu B in direct anterior nasal swab specimens. As no other sample types are claimed herein, a matrix comparison study is not applicable.

18

However, the sponsor performed the matrix equivalency study between pooled negative nasal swab matrix (PNSM) and the surrogate pooled negative nasal wash (PNW) that was used in multiple analytical studies. The data demonstrated equivalent performance of the test with both matrices.

C Clinical Study:

1. Clinical Performance Assessment:

A multi-center, prospective clinical study was conducted with lay users to assess the performance of the Healgen Rapid Check COVID-19/Flu A&B Antigen Test in detecting nucleoprotein antigens extracted from COVID-19, influenza virus types A and B in selfcollected and self-tested anterior nasal swab samples. The study only enrolled subjects with two or more symptoms of respiratory infection consistent with COVID-19 or influenza. Ten clinical sites across the U.S. conducted the study from February to April 2024. Both the comparator and the candidate test used anterior nasal swab samples and the collection order was alternated by study subject. Comparator test samples were collected by health care professionals at the clinical study sites and inserted into Universal Transport Media per the IFU of the comparator test. Samples were then sent to a central site for testing with highly sensitive RT-PCR tests separately detecting SARS-CoV-2 and Flu A/B. Samples for the candidate antigen test were collected per the test's quick reference instructions and were either self-collected by a lay user aged >14 years or collected by an adult (parent/guardian) from individuals aged 2 to 65 years of age | 0 (0.0%) | 2 (7.7%) | 2 (3.9%) |

Table 18: Demographics of the Usability Study Population

The human factors assessment portion of the study was completed per the protocol. Fifty-one (51) subjects (26 self-collecting and testing, and 25 lay users collecting and testing from another) were enrolled in the human factors assessment. Evaluation of the human user experience indicated high usability of the investigational test. All subjects who participated found the instructions to be clear and easy to follow and found the sample collection easy to perform, as well as having no difficulty reading the test results. Additionally, 97.5% (796 out of 816 tests) of the mock tests were interpreted correctly. Overall, 96.8% of all critical tasks associated with sample collection and the running of the Healgen COVID-19/Flu A&B Ag Combo Rapid Test Cassette (Swab) were performed correctly. Additionally, 87.6% of all non-critical tasks were

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performed correctly. The human factors assessment met the predetermined targets for the percentage of critical and noncritical tasks performed correctly as shown in the table below.

| Steps | Tasks performed
correctly | Total number
of tasks | Percentage of tasks
performed correctly |
|---------------|------------------------------|--------------------------|--------------------------------------------|
| Critical | 642 | 663 | 96.8% |
| Non- Critical | 134 | 153 | 87.6% |
| Total | 776 | 816 | 95.1% |

Table 19:Usability Study Results

3. Lay User Readability Assessment:

All 51 subjects who participated in the human factors assessment (Usability study) also interpreted a panel of mock investigational tests with various results that reflected the test concentration levels at 1.9x and 5x the limits of detection (LOD) in a blinded and random fashion. Each panel of mock tests included 16 investigational tests with various negative and positive results for each analyte. The percentage of total human factor subjects with vision impairment is 39.2% (20/51). Vision impairments encountered in study subjects are listed in the table below with their respective frequency of occurrence. The study did not include individuals with any of the following: macular degeneration, color blindness, diabetic retinopathy, glaucoma, cataracts, or amblyopia/strabismus. The overall accuracy of the results interpreted by the lay users is 97.5% (796/816): 95% CI (96.2% - 98.4%).

| Vision Impairment | # of
Subjects | Percentage of total human factors
subjects with vision
impairment (N=51) |
|---------------------------------------------------|------------------|--------------------------------------------------------------------------------|
| Near sightedness only
(with lens prescription) | 6 | 11.8% |
| Far sightedness only (with lens
prescription) | 6 | 11.8% |
| Astigmatism | 1 | 2.0% |
| More than one visual impairment
condition | 7 | 13.7% |
| Total subjects/testers with vision
impairment | 20 | 39.2% |

Table 20: Vision Impairment of Readability Study Subjects

The comparison of result interpretation data between lay users with and without visual impairment is tabulated below.

	Accuracy of Mock Test Interpretations [%]
Mock Results Type	Subjects without
vision impairment
(N=31)	Subjects with
vision impairment
(N=20)
1.9x LoD - Flu A+ & Flu B+	100.0%	100.0%
1.9x LoD - COV-19+/Flu A+	93.5%	100.0%

Table 21: Lay User Readability Study Results

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Mock Results Type	Accuracy of Mock Test Interpretations [%]
	Subjects without
vision impairment
(N=31)	Subjects with
vision impairment
(N=20)
1.9x LoD - COV-19+/Flu A+ & Flu B+	93.5%	100.0%
1.9x LoD - COV-19+/Flu B+	90.3%	100.0%
1.9x LoD - COV+	93.5%	100.0%
1.9x LoD - Flu A+	100.0%	100.0%
1.9x LoD - Flu B+	96.8%	100.0%
5x LoD - Flu A+ & Flu B+	100.0%	100.0%
5x LoD - COV-19+/Flu A+	96.8%	100.0%
5x LoD - COV-19+/Flu A+ & Flu B+	100.0%	100.0%
5x LoD - COV-19+/Flu B+	96.8%	95.0%
5x LoD - COV+	100.0%	100.0%
5x LoD - Flu A+	100.0%	95.0%
5x LoD - Flu B+	100.0%	100.0%
Invalid	93.5%	95.0%
Negative	93.5%	95.0%
Total	96.8%	98.8%

D Clinical Cut-Off:

Not Applicable. The candidate device is a qualitative assay with a visually read binary result without numeric raw data.

E Expected Values/Reference Range:

A patient sample is expected to be negative for SARS-CoV-2, influenza A and influenza B. This section is therefore not applicable.

Other Supportive Performance Characteristics Data: F

1. Variant Monitoring Plan:

To determine whether the Healgen Rapid Check COVID-19/Flu A/B Antigen Test can detect newly emerging variants, and/or to assess whether new mutations are impacting analytical sensitivity of the test performance, the sponsor provided the variant monitoring plan as described below:

a. Monitoring SARS-CoV-2, Influenza A and B Sequence Data in GISAID database, WHO, NIH and other public health entities:
The updated sequence data for SARS-CoV-2, influenza a and influenza B variants from GISAID database, WHO, NIH and other public health entities will be downloaded and analyzed bimonthly for variant mutations in the target proteins with an allele frequency of ≥5%

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b. In silico analysis of antigenicity of the N proteins: In silico monitoring of antigen variations caused by changes in aminoacid residues will be performed by analyzing linear epitopes through sequence alignments and structure prediction methods.
c. Wet lab test on recombinant N proteins: Full length recombinant mutated N proteins will be used for wet lab testing for VOC monitoring, including VOC/VOI/VUM SARS-CoV-2 strains as well as other strains with higher frequency mutation.
d. Evaluation with SARS-CoV-2, Flu A and B variant viruses with External Quality Assessment schemes or commercially available viruses:

External quality assessment schemes or commercial resources will be used to determine possible performance effect of our test affected by mutations in the testing proteins. Additional validation tests using clinical samples will be conducted involving a comparative analysis of the results obtained through the candidate device, an FDA-approved PCR test and next-generation sequencing techniques to ensure the utmost accuracy and reliability of our testing device's performance assessment.

2. Frequently Asked Questions:

To improve user label comprehension, the labeling includes a Frequently Asked Questions (FAQ) section. The FAQ section was created to provide users information to adequately understand the meaning of the test results and test types as well as the accuracy of the test. The concepts covered in the FAO section include:

. Meaning of the test results.
. When to re-test (e.g., following an invalid result). Difference between antigen and molecular test
Accuracy of the test ●
. Follow-up for appropriate health management.

3. Hazard Analysis:

A comprehensive hazard analysis of the Healgen Rapid Check COVID-19/Flu A/B Antigen Test included identification of the potential hazard, likelihood of occurrence, severity of potential harm, hazard control measure(s), hazard control verification, and assignment of pre- and postcontrol risk levels. The elements considered included operator errors (i.e., human factors), sample and device handling and storage, and environmental factors.

Potential sources of errors that could adversely affect system performance were identified and mitigated through cautions in the labeling. The identified risks which could result in erroneous test results were evaluated in flex studies that evaluated the functionality of fail-safe mechanisms and stressed the functional limits of the test system (see below).

4. Fail-Safe Features:

The device features an internal control to minimize false results due to user error. The internal control monitors for grossly insufficient sample volume, adequate membrane wicking, sample flow, and the integrity of the detection reagents.

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5. Flex Studies:

To assess the robustness and risk for false results of the test when deviating from the IFU/QRI test steps, flex studies were conducted that assessed all major aspects of the test procedure (sample volume, reading time, swab extraction time and procedure [delay in mixing and addition of the sample], sample hold time before and during processing) and variability of environmental test conditions that the test may be subjected to when in use (lighting, disturbance during use, temperature and humidity stress conditions). Testing was performed with negative PNW samples and low positive samples co-spiked with SARS-CoV-2, Flu A and Flu B virus into negative PNW at 2xLoD.

The results demonstrated that the test system is robust and that false results can be expected to be reasonably mitigated through labeling.

VII. Proposed Labeling:

The labeling supports the decision to grant the De Novo request for this device.

Risks to Health	Mitigation Measures
False Results	Certain labeling information including limitations,
device descriptions, explanations of procedures, and
performance information.
	Certain design verification and validation including
documentation of device descriptions, certain analytical
and clinical studies, and risk analysis.
	Certain strain monitoring and reporting strategies.
Failure to correctly interpret test results	Certain labeling information including limitations,
device descriptions, explanations of procedures, and
performance information.
	Certain design verification and validation including
documentation of device descriptions, certain analytical
and clinical studies, and risk analysis.
	Certain labeling information including limitations,
device descriptions, explanations of procedures, and
performance information.
Failure to correctly operate the device	Certain labeling information including limitations,
device descriptions, explanations of procedures, and
performance information.

VIII. Identified Risks and Mitigations:

IX. Benefit/Risk Assessment:

A Summary of the Assessment of Benefit:

The primary probable benefit of the Healgen Rapid Check COVID-19/Flu A&B Antigen Test device is to facilitate easy-to-use rapid detection of respiratory viruses by the lay user in the

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home environment using an adequately validated, well-performing, multi-analyte respiratory test, with the potential to test an entire household within the first 5 days of onset of symptoms. Homebased testing for the detection and differentiation of these viruses is addressing an unmet public health need because it offers several important advantages, foremost shortening the time to diagnosis and therefore treatment and isolation, as necessary. This reduces the risk of severe illness as well as virus transmission. Indirectly, this may lessen demand on overburdened public health and clinical laboratories during times of increased transmission. For persons facing particular barriers to accessing care outside the home, whether due to physical, cognitive, or demographic factors, at-home over-the-counter (OTC) tests may potentially help improve health equity in early diagnosis and treatment, and thereby improve outcomes of COVID-19, influenza A (Flu A), and influenza B (Flu B).

B Summary of the Assessment of Risk:

The risks associated with the device, when used as intended, are related to the risk of false results as a consequence of either performance issues of the test, and/or a failure of the operator to correctly operate the test or interpret the test results correctly.

False positive SARS-CoV-2, influenza A virus, and influenza B virus test results may lead to initiation of improper patient management (such as administration of unnecessary or wrong antiviral drugs with unintended complications) or failure to recognize and appropriately treat other respiratory illnesses that may resemble COVID-19 or influenza but are not caused by these viruses.

False negative SARS-CoV-2, influenza A virus, and influenza B virus test results may lead to unnecessary additional diagnostic evaluation or treatment, and delays in correct diagnosis possibly leading to missed opportunities to initiate time-sensitive treatment, and/or infection control measures to avoid transmission to additional persons. This may consequently lead to an increase in patient morbidity and mortality.

Visually interpreted tests also have a risk to be falsely interpreted as invalid when the actual result is a valid positive or valid negative result. In instances where a valid test result is falsely interpreted as invalid, this may lead to unnecessary delays while additional testing is sought, possibly leading to missed opportunities to initiate time-sensitive treatment, and/or infection control measures to avoid transmission to additional persons.

C Summary of the Assessment of Benefit-Risk:

The most serious risk of the device is a false negative result, leading to a missed or delayed diagnosis of SARS-CoV-2, influenza A virus or influenza B virus infection, or a false positive result, leading to improper patient management or failure to recognize and appropriately treat other respiratory illnesses resembling COVID-19 and influenza. However, the risk of false results is low, as evident from the results of the required device validation and verification, including analytical and clinical studies. In addition, this risk can be mitigated by the clear guidance provided in the labeling limitations, the clear procedural instructions in the labeling written at a 7th grade reading level, and the repeated and clear guidance in the labeling for the user to contact their healthcare provider if there is evidence of worsening disease or risk factors for severe disease are present.

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The risk of failure to correctly interpret the test results of a visually read device is mitigated by clear pictorial and descriptive instructions in the result interpretation section of the labeling with important warning statements being emphasized in bolded text.

Equally, the risk of failure to correctly operate the device is also mitigated by certain labeling information, including the presence and emphasis (bolded text) of warnings and limitations within the procedural instructions, device descriptions, and performance sections.

Clinical and usability data suggest that errors will be uncommon and that the test can be accurately performed and results accurately interpreted. While general controls are not sufficient to mitigate the risks of this device, in light of the special controls, the benefits outweigh the risks for this device.

X. Conclusion:

The De Novo request is granted and the device is classified under the following and subject to the special controls identified in the letter granting the De Novo request:

Product Code(s): SCA Device Type: Multi-analyte respiratory virus antigen detection test Class: II Regulation: 866.3987