The VELO Respiratory Test is an automated rapid multiplex real-time, reverse transcriptase polymerase chain reaction (RT-PCR) test performed on the VELO Instrument and is intended for the simultaneous qualitative detection and differentiation of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), influenza A, and influenza B virus nucleic acid in anterior nasal swab (ANS) specimens from individuals with signs and symptoms of respiratory tract infection. Clinical signs and symptoms of respiratory tract infection due to SARS-CoV-2 and influenza can be similar.

The VELO Respiratory Test is intended for use as an aid in the differential diagnosis of SARS-CoV-2, influenza A, and/or influenza B infection if used in conjunction with other clinical and epidemiological information, and laboratory findings. SARS-CoV-2, influenza A and influenza B viral nucleic acid are generally detectable in ANS specimens during the acute phase of infection.

Positive results do not rule out co-infection with other organisms. The agent(s) detected by the VELO Respiratory Test may not be the definite cause of disease.

Negative results do not preclude SARS-CoV-2, influenza A, and/or influenza B infection. The results of this test should not be used as the sole basis for diagnosis, treatment, or other patient management decisions.

Device Description

The VELO System is comprised of a single-use VELO Respiratory Test, and a reusable VELO Instrument:

The Test contains individually packaged consumables: a "Test Cartridge" and an anterior nasal swab.
The Instrument is a small, benchtop device with preloaded software for running tests and viewing the results. For a full description of the VELO Instrument, please refer to the VELO Instrument Manual.

Each Test Cartridge contains all necessary reagents for the detection of Flu A, Flu B and SARS-CoV-2 viral RNA from anterior nasal swab specimens. Primers and probes in the VELO Respiratory Test are designed to amplify and detect unique sequences within the following regions of each target pathogen: influenza A genome (matrix protein gene), influenza B genome (non-structural protein gene), and SARS-CoV-2 genome (ORF 1a/b non-structural region, and membrane protein gene). Each Test Cartridge also contains an endogenous Sample and Process Control (SPC). The SPC acts as an Internal Control (IC), to control for adequate sample collection and processing, the detection of failures in the reaction resulting from PCR inhibition, or a failure of the reagents. The SPC also ensures that the RT-PCR reaction conditions (temperature and time) are appropriate for the amplification reaction and that the resulting signal can be detected and processed by the system. Test outcomes are reported to the operator in real-time via the Instrument viewscreen with 'Not Detected' results available in under 10 minutes, when all cycles have completed. When the Test ends, all results can be viewed via the Instrument viewscreen, and the Test Cartridge may be removed for disposal.

AI/ML Overview

N/A

Summary

FDA 510(k) Clearance Letter - VELO Respiratory Test

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U.S. Food & Drug Administration
10903 New Hampshire Avenue
Silver Spring, MD 20993
www.fda.gov

Doc ID # 04017.08.03

February 13, 2026

LEX Diagnostics Limited
Joanne Channon
Director of Program Management
Melbourn Science Park
Melbourn, SG8 6EE
United Kingdom

Re: K251742
Trade/Device Name: VELO Respiratory Test
Regulation Number: 21 CFR 866.3981
Regulation Name: Device To Detect And Identify Nucleic Acid Targets In Respiratory Specimens From Microbial Agents That Cause The Sars-Cov-2 Respiratory Infection And Other Microbial Agents When In A Multi-Target Test
Regulatory Class: Class II
Product Code: QOF
Dated: June 6, 2025
Received: June 6, 2025

Dear Joanne Channon:

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

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

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K251742 - Joanne Channon Page 2

Additional information about changes that may require a new premarket notification are provided in the FDA guidance documents entitled "Deciding When to Submit a 510(k) for a Change to an Existing Device" (https://www.fda.gov/media/99812/download) and "Deciding When to Submit a 510(k) for a Software Change to an Existing Device" (https://www.fda.gov/media/99785/download).

Your device is also subject to, among other requirements, the Quality Management System Regulation (QMSR) (21 CFR Part 820), which includes, but is not limited to, ISO 13485 clause 7.3 (Design controls), ISO 13484 clause 8.3 (Nonconforming product), and ISO 13485 clause 8.5 (Corrective and preventative action). Please note that regardless of whether a change requires premarket review, the QMSR requires device manufacturers to review and approve changes to device design and production (ISO 13485 clause 7.3 and 21 CFR 820.70) and document changes and approvals in the device master record (21 CFR 820.181).

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

All medical devices, including Class I and unclassified devices and combination product device constituent parts are required to be in compliance with the final Unique Device Identification System rule ("UDI Rule"). The UDI Rule requires, among other things, that a device bear a unique device identifier (UDI) on its label and package (21 CFR 801.20(a)) unless an exception or alternative applies (21 CFR 801.20(b)) and that the dates on the device label be formatted in accordance with 21 CFR 801.18. The UDI Rule (21 CFR 830.300(a) and 830.320(b)) also requires that certain information be submitted to the Global Unique Device Identification Database (GUDID) (21 CFR Part 830 Subpart E). For additional information on these requirements, please see the UDI System webpage at https://www.fda.gov/medical-devices/device-advice-comprehensive-regulatory-assistance/unique-device-identification-system-udi-system.

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

For comprehensive regulatory information about medical devices and radiation-emitting products, including information about labeling regulations, please see Device Advice (https://www.fda.gov/medical-devices/device-advice-comprehensive-regulatory-assistance) and CDRH Learn (https://www.fda.gov/training-and-continuing-education/cdrh-learn). Additionally, you may contact the Division of Industry and Consumer Education (DICE) to ask a question about a specific regulatory topic. See the DICE website (https://www.fda.gov/medical-devices/device-advice-comprehensive-regulatory-

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K251742 - Joanne Channon Page 3

assistance/contact-us-division-industry-and-consumer-education-dice) for more information or contact DICE by email (DICE@fda.hhs.gov) or phone (1-800-638-2041 or 301-796-7100).

Sincerely,

JOSEPH BRIGGS -S

Joseph Briggs, Ph.D.
Deputy Director
Division of Microbiology Devices
OHT7: Office of In Vitro Diagnostics
Office of Product Evaluation and Quality
Center for Devices and Radiological Health

Enclosure

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DEPARTMENT OF HEALTH AND HUMAN SERVICES
Food and Drug Administration

Form Approved: OMB No. 0910-0120
Expiration Date: 06/30/2023
See PRA Statement below.

Indications for Use

510(k) Number (if known)
K251742

Device Name
VELO Respiratory Test

Indications for Use (Describe)

Positive results do not rule out co-infection with other organisms. The agent(s) detected by the VELO Respiratory Test may not be the definite cause of disease.

Type of Use (Select one or both, as applicable)
☒ Prescription Use (Part 21 CFR 801 Subpart D) ☐ Over-The-Counter Use (21 CFR 801 Subpart C)

CONTINUE ON A SEPARATE PAGE IF NEEDED.

This section applies only to requirements of the Paperwork Reduction Act of 1995.
DO NOT SEND YOUR COMPLETED FORM TO THE PRA STAFF EMAIL ADDRESS BELOW.

The burden time for this collection of information is estimated to average 79 hours per response, including the time to review instructions, search existing data sources, gather and maintain the data needed and complete and review the collection of information. Send comments regarding this burden estimate or any other aspect of this information collection, including suggestions for reducing this burden, to:

Department of Health and Human Services
Food and Drug Administration
Office of Chief Information Officer
Paperwork Reduction Act (PRA) Staff
PRAStaff@fda.hhs.gov

"An agency may not conduct or sponsor, and a person is not required to respond to, a collection of information unless it displays a currently valid OMB number."

FORM FDA 3881 (6/20) Page 1 of 1 PSC Publishing Services (301) 443-6740 EF

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LEX Diagnostics Limited
Melbourn Science Park
Melbourn, SG8 6EE, UK
+44 1223 752144
enquiries@lexdiagnostics.com
lexdiagnostics.com

Registered office: TTP Campus, Melbourn, SG8 6HQ, United Kingdom.
Registered number 12455716

VELO Respiratory Test 510(k) Summary

Date Prepared: 04 February 2026
Submitter's Name: Heather Danks
Submitter's email: Heather.Danks@LEXDiagnostics.com
Submitter's phone: +44 1223 752144

Device Trade Name: VELO Respiratory Test

Common Name: VELO Respiratory Test

Classification Name: Device to detect and identify nucleic acid targets in respiratory specimens from microbial agents that cause the SARS-CoV-2 respiratory infection and other microbial agents when in a multi-target test. (21 CFR § 866.3981)

Product Code: QOF

Predicate Device: cobas® SARS-CoV-2 & Influenza A/B for use on the cobas Liat System (K223591)

DEVICE DESCRIPTION

The VELO System is comprised of a single-use VELO Respiratory Test, and a reusable VELO Instrument:

The Test contains individually packaged consumables: a "Test Cartridge" and an anterior nasal swab.
The Instrument is a small, benchtop device with preloaded software for running tests and viewing the results. For a full description of the VELO Instrument, please refer to the VELO Instrument Manual.

INTENDED USE

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influenza A, and influenza B virus nucleic acid in anterior nasal swab (ANS) specimens from individuals with signs and symptoms of respiratory tract infection. Clinical signs and symptoms of respiratory tract infection due to SARS-CoV-2 and influenza can be similar.

Positive results do not rule out co-infection with other organisms. The agent(s) detected by the VELO Respiratory Test may not be the definite cause of disease.

INTENDED OPERATORS AND USE ENVIRONMENTS (CLIA STATEMENT):

The VELO Respiratory Test is intended for use by non-laboratory trained healthcare professionals in healthcare facilities, operating under a CLIA Certificate of Waiver.

PRINCIPLES OF THE PROCEDURE

Each VELO Respiratory Test Cartridge contains all necessary reagents for the detection of influenza A (Flu A), influenza B (Flu B) and SARS-CoV-2 viral RNA from anterior nasal swab specimens. Reverse Transcription-Polymerase Chain Reaction (RT-PCR) is used with specific primers and probes within the Cartridge to amplify and detect sequences unique to each target pathogen. Specifically, the matrix protein gene of Flu A, the non-structural gene of Flu B and the ORF1a/b non-structural region and membrane protein gene of SARS-CoV-2.

Each Test Cartridge also contains an endogenous Sample Process Control (SPC). The SPC serves as an Internal Control (IC) mechanism to ensure adequate sample collection and processing. It also monitors for under performance of the RT-PCR reaction, resulting from sample mediated inhibition or failure of the reagents. Additionally, the SPC verifies that the RT-PCR reaction conditions (temperature and duration) are optimal for the amplification process. This ensures that the resulting signal can be accurately detected and processed by the system.

Within the Test Cartridge, reagents are prepackaged and require no special handling. The anterior nasal specimen is directly entered into the Test Cartridge with the swab shaft removed at a pre-specified breakpoint. All further operational steps are automatically executed, eliminating the need for transfer media, reagent preparation, pipetting, temperature monitoring or timing. No complex nucleic acid extraction or purification is performed, rather, thermal lysis is used with the lysed materials directly rehydrating lyophilized RT-PCR reagents. Each target amplification reaction proceeds in an independent PCR chamber.

The VELO System is based upon proprietary technology that allows ultra-fast thermal cycling. This is achieved by combining the rapid transfer of heat into and out of the reaction with rapid thermalization from low volume and a high surface area to volume ratio of the reaction chamber. This ultra-fast thermal cycling enables rapid amplification, with typical thermal cycle durations of less than 10 seconds (per cycle), while still retaining the high sensitivity and specificity associated with hydrolysis probe detection.

In the event of amplification, a fluorescence signal is generated through the degradation of oligonucleotide probes modified with 5' fluorophores and 3' quenchers. Fluorescence is monitored by the Instrument with every thermal cycle and reports as "Detected" once meeting pre-determined criteria. Test outcomes are

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reported to the operator in real-time via the Instrument view screen with 'Not Detected' results available in under 10 minutes with the completion of all cycles. When the Test ends, all results can be viewed via the Instrument view screen, and the Test Cartridge may be removed for disposal.

PREDICATE

The VELO Respiratory Test is predicated on the cobas® SARS-CoV-2 & Influenza A/B for use on the cobas Liat System (K223591, Roche Molecular Systems).

TECHNOLOGICAL CHARACTERISTICS

A comparison of the VELO Respiratory Test and VELO Instrument to the predicate is provided in the table below.

Differences in technological characteristics between the VELO Respiratory Test and VELO Instrument and the predicate are limited to the following:

VELO employs the use of direct swab elution within the Test Cartridge.
VELO employs the use of an endogenous (RNaseP) Sample Process Control
VELO employs thermal lysis followed by direct RT-PCR amplification
VELO's second SARS-CoV-2 viral target is within the Membrane protein gene.
VELO provides test results in 10 minutes or less

Table 1 Comparison Table

Parameter	VELO Respiratory Test	cobas® SARS-CoV-2 & Influenza A/B for use on the cobas Liat System
510(k) Number	K251742	K223591
510(k) Manufacturer	LEX Diagnostics Limited	Roche Molecular Systems, Inc
Regulation	Same	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
Regulatory Class	Same	Class II (Special Controls)
Classification Product Code	Same	QOF
Panel	Same	Microbiology
Assay Technology	Same	Multiplexed PCR amplification with qualitative fluorescence detection
Indications for Use	The VELO Respiratory Test is an automated rapid multiplex real-time, reverse transcriptase polymerase chain reaction (RT-PCR) test performed on the VELO	The cobas SARS-CoV-2 & Influenza A/B nucleic acid test for use on the cobas Liat System (cobas SARS-CoV-2 & Influenza A/B) is an automated rapid multiplex real-

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Parameter	VELO Respiratory Test	cobas® SARS-CoV-2 & Influenza A/B for use on the cobas Liat System
	Instrument and is intended for the simultaneous qualitative detection and differentiation of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), influenza A, and/or influenza B virus nucleic acid in anterior nasal swab (ANS) specimens from individuals with signs and symptoms of respiratory tract infection. Clinical signs and symptoms of respiratory tract infection due to SARS-CoV-2 and influenza can be similar. The VELO Respiratory Test is intended for use as an aid in the differential diagnosis of SARS-CoV-2, influenza A, and/or influenza B infection if used in conjunction with other clinical and epidemiological information, and laboratory findings. SARS-CoV-2, influenza A, and influenza B viral nucleic acid are generally detectable in ANS specimens during the acute phase of infection. Positive results do not rule out co-infection with other organisms. The agent(s) detected by the VELO Respiratory Test may not be the definitive cause of disease. Negative results do not preclude SARS-CoV-2, influenza A, and/or influenza B infection. The results of this test should not be used as the sole basis for diagnosis, treatment, or other patient management decisions.	time, reverse transcriptase polymerase chain reaction (RT-PCR) test intended for the simultaneous qualitative detection and differentiation of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), influenza A, and/or influenza B virus nucleic acid in nasopharyngeal swab (NPS) and anterior nasal swab (ANS) specimens from individuals with signs and symptoms of respiratory tract infection. Clinical signs and symptoms of respiratory tract infection due to SARS-CoV-2 and influenza can be similar. cobas SARS-CoV-2 & Influenza A/B is intended for use as an aid in the differential diagnosis of SARS-CoV-2, influenza A, and/or influenza B infection if used in conjunction with other clinical and epidemiological information, and laboratory findings. SARS-CoV-2, influenza A, and influenza B viral nucleic acid are generally detectable in NPS and ANS specimens during the acute phase of infection. Positive results do not rule out co-infection with other organisms. The agent(s) detected by the cobas SARS-CoV-2 & Influenza A/B may not be the definitive cause of disease. Negative results do not preclude SARS-CoV-2, influenza A, and/or influenza B infection. The results of this test should not be used as the sole basis for diagnosis, treatment, or other patient management decisions.
Intended Users and Use Locations	Same	Healthcare facilities operating under a CLIA Certificate of Waiver
CLIA Complexity	Same	Waived
Analyte Targets	SARS-CoV-2: ORF1a/b nonstructural regionSARS-CoV-2: membrane protein geneInfluenza A: matrix geneInfluenza B: non-structural protein gene	SARS-CoV-2: ORF1a/b nonstructural regionSARS-CoV-2: nucleocapsid protein geneInfluenza A: matrix geneInfluenza B: non-structural protein gene

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Parameter	VELO Respiratory Test	cobas® SARS-CoV-2 & Influenza A/B for use on the cobas Liat System
Sample Types	Anterior Nasal Swabs (ANS)	Anterior Nasal Swabs (ANS), and Nasopharyngeal Swabs (NPS)
Instrumentation	VELO Instrument	cobas Liat System
Reagents / Kit Components	Included in VELO Respiratory Test Cartridge, with no user involvement required	Included in Liat assay tube, with no user involvement required
Ancillary Collection Kits	Copan FLOQSwabs™	Copan FLOQSwabs with UTM™, UVT,0.9% and other swabs with other VTM
Elution	Direct elution within the VELO Respiratory Test Cartridge	Swab eluted in transfer media (VTM/UTM)
Sample Preparation	Thermal lysis for nucleic acid release from cells / viral particles	Automated silica magnetic particle-based nucleic acid extraction
Detection	Multiplexed assay using fluorescent probes, detected automatically by the VELO Instrument, not reliant on user judgement	Multiplexed assay using different reporter dyes for target and control, detected automatically by the cobas® Liat instrument, not reliant on user judgement
Internal Controls	Yes – Endogenous RNaseP Sample Process Control	Yes – Exogenous Sample Process Control ('Internal Process control')
External Control	Yes – commercially available	Yes – sold separately as the cobas® SARS-CoV-2 & Influenza A/B Quality Control Kit
Assay Results	Qualitative – as determined by an algorithm within the instrument (VELO)	Qualitative – as determined by an algorithm within the instrument (Liat)
Results Interpretation	Same	Visual read of automated results on a viewscreen
Time to Result	In 10 minutes or less	In approximately 20 minutes

Material differences are provided in bold typeface for ease of reference

A rigorous analytical performance assessment demonstrated comparable performance of the VELO Respiratory Test used on the VELO Instrument against the predicate device. No safety or efficacy risks have been identified in relation to the similarities and/or differences identified herein.

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PERFORMANCE CHARACTERISTICS

Analytical Sensitivity (Limit of Detection)

The Limit of Detection (LoD) of the VELO Respiratory Test in pooled anterior nasal matrix was evaluated by testing limiting dilutions of two (2) strains of Influenza A H1N1 (A/PR/8/34 and A/New Cal/20/99), two (2) strains of Influenza A H3N2 (A/Hong Kong/8/68 and A/South Australia/55/14), Influenza B Yamagata Lineage (B/Wisconsin/1/2010), Influenza B Victoria Lineage (Malaysia/2506/04), the 2nd WHO International Standard for SARS-CoV-2 (Beta CoV/Australia/VIC01/2020) and an inactivated Omicron (BA.5) strain of SARS-CoV-2 (hCoV-19/USA/COR-22-06-3113/2022).

The preliminary LoD for each virus strain per lot was established as the minimum concentration for which 3/3 replicates returned a 'Detected' result for the VELO Respiratory Test. The confirmatory LoD for each virus strain was determined by retesting the preliminary LoD and at least one (1) higher concentration and one (1) lower concentration for each viral target with at least 20 replicates of each concentration/batch across a minimum of three (3) days. Confirmatory LoD was determined as the lowest concentration of each virus strain that was consistently detected (defined as ≥95% of samples) by both lots of tests. The highest (least sensitive) LoD value for the two (2) lots was reported as the final LoD. These LoD values were then verified with one (1) additional batch for each strain. The verified LoD values for each virus tested are shown in the table below.

Table 2 LoD Determination of influenza A, influenza B and SARS-CoV-2

Virus	Strain	Limit of Detection
SARS-CoV-2	BetaCoV/Australia/VIC01/2020	3,000 IU/swab
	hCoV-19/USA/COR-22-06-3113/2022	0.5 TCID50/swab
Influenza A H1N1	A/PR/8/34	3,000 cp/swab
	A/New Cal/20/99	2.5 TCID50/swab
Influenza A H3N2	A/Hong Kong/8/68	3,000 cp/swab
	A/South Australia/55/14	2.5 TCID50/swab
Influenza B (Yamagata)	B/Wisconsin/1/2010	3,000 cp/swab
Influenza B (Victoria)	Malaysia/2506/04	1.0 TCID50/swab

Analytical Reactivity (Inclusivity) - Inclusivity wet testing

The inclusivity of the VELO Respiratory Test on the VELO instrument was established through wet testing by evaluating ten (10) strains of SARS-CoV-2, ten (10) strains of influenza A H1N1, ten (10) strains of influenza H3N2 and eight (8) strains of influenza B. Each strain was tested individually initially at 3x LoD in pooled negative nasal matrix (NNM) in replicates of three (3) in a blinded and randomized fashion. Inclusivity was determined if the virus was detected in 3/3 replicates. If the initial test did not return 3/3 detected results, template concentration was increased until 3/3 was detected. The lowest concentration of template returning 3/3 detected results is detailed below.

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Inclusivity of SARS-CoV-2

Table 3 Inclusivity results for SARS-CoV-2 strains

Virus	Strain	Concentration	Number detected/number tested (% detected)
SARS-CoV-2	2019-nCoV/USA-WA1/2020	9,000 cp/swab	3/3 (100%)
SARS-CoV-2 (Alpha-Variant)	Alpha (B.1.1.7) VOC202012/01	9,000 cp/swab	3/3 (100%)
SARS-CoV-2 (Beta-Variant)	Beta (B.1.351) VOC202012/02	9,000 cp/swab	3/3 (100%)
SARS-CoV-2 (Gamma-Variant)	Gamma (P.1)	9,000 cp/swab	3/3 (100%)
SARS-CoV-2 (Delta-Variant)	Delta (B.1.617.2) VOC21APR-02	9,000 cp/swab	3/3 (100%)
SARS-CoV-2 (Omicron-Variant)	BA.2.12.1; USA/NY-Wadsworth-22014351-01/2022	1.5 TCID50/swab	3/3 (100%)
SARS-CoV-2 (Omicron-Variant)	BA.4.6; USA/MDHP35538/2022	1.5 TCID50/swab	3/3 (100%)
SARS-CoV-2 (Omicron-Variant)	BF.7; USA/NY-Wadsworth22042128-01/2022	1.5 TCID50/swab	3/3 (100%)
SARS-CoV-2 (Omicron-Variant)	XBB; USA/CA-Stanford109_S21/2022	1.5 TCID50/swab	3/3 (100%)
SARS-CoV-2 (Omicron-Variant)	XBB.1.5; USA/NY-Wadsworth-22061020-01/2022	1.5 TCID50/swab	3/3 (100%)

These SARS-CoV-2 strains are in addition to the BetaCoV/Australia/VIC01/2020 and hCoV-19/USA/COR-22-06-3113/2022 used in the analytical sensitivity study.

In addition, the two (2) SARS-CoV-2 assay targets (Orf1ab and M) within the VELO Respiratory Test were subject to in silico analysis to ensure coverage of the assay as a whole. In silico analysis of 137,277 SARS-CoV-2 sequences (as of May 2025) indicate that 100% of sequences for SARS-CoV-2 have no changes within the primer and/or probe binding regions that would be predicted to affect the performance of both assay targets. Therefore, all known variants are predicted to be detected.

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Influenza A inclusivity

Table 4 Inclusivity results for influenza A H1N1 strains

Virus	Strain	Concentration	Number detected/number tested (% detected)
Influenza A (H1N1pdm)	A/California/07/09	7.5 TCID50/swab	3/3 (100%)
Influenza A (H1N1pdm)	A/NY/01/09	10 TCID50/swab ᵃ	3/3 (100%)
Influenza A (H1N1pdm)	A/Victoria/4897/2022	9,000 cp/swab	3/3 (100%)
Influenza A (H1N1pdm)	A/Wisconsin/67/2022	9,000 cp/swab	3/3 (100%)
Influenza A (H1N1pdm)	A/California/08/2009	168 CEID50/swab ᵇ	3/3 (100%)
Influenza A H1N1	A/Denver/1/1957	112 CEID50/swab ᵃ	3/3 (100%)
Influenza A H1N1	A/New Jersey/8/1976	168 CEID50/swab ᶜ	3/3 (100%)
Influenza A H1N1	A/NWS/33	84 CEID50/swab	3/3 (100%)
Influenza A H1N1	A/Solomon Island/3/2006	84 CEID50/swab	3/3 (100%)
Influenza A H1N1	A/Swine/Iowa/15/30	140 CEID50/swab ᵈ	3/3 (100%)

ᵃ Results for 4x LoD, results at 3x LoD were 5/6 detected
ᵇ Results for 6x LoD as inclusivity at 3x LoD could not be determined (4/6 detected), results at 5x LoD were 5/6 detected.
ᶜ Results for 6x LoD as inclusivity at 3x LoD could not be determined (0/3 detected), results at 5x LoD were 5/6 detected
ᵈ Results for 5x LoD as inclusivity at 3x LoD could not be determined (3/6 detected)

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Table 5 Inclusivity results for influenza A H3N2 strains

Virus	Strain	Concentration	Number detected/number tested (% detected)
Influenza A (H3N2)	A/Aichi/2/68	236.4 CEID50/swab	3/3 (100%)
Influenza A (H3N2)	A/Brisbane/10/07	236.4 CEID50/swab	3/3 (100%)
Influenza A (H3N2)	A/California/122/2022	9,000 cp/swab	3/3 (100%)
Influenza A (H3N2)	A/Switzerland/9715293/13	7.5 TCID50/swab	3/3 (100%)
Influenza A (H3N2)	A/Sydney/5/1997	236.4 CEID50/swab	3/3 (100%)
Influenza A (H3N2)	A/Texas/50/12	7.5 TCID50/swab	3/3 (100%)
Influenza A (H3N2)	A/Thailand/8/2022	9,000 cp/swab	3/3 (100%)
Influenza A (H3N2)	A/Uruguay/716/2007	236.4 CEID50/swab	3/3 (100%)
Influenza A (H3N2)	A/Victoria/361/2011	236.4 CEID50/swab	3/3 (100%)
Influenza A (H3N2)	A/Wisconsin/67/05	7.5 TCID50/swab	3/3 (100%)

These influenza A strains are in addition to the A/PR/8/34, A/New Cal/20/99, A/Hong Kong/8/68 and A/South Australia/55/14 strains used in the analytical sensitivity study.

In addition, in silico analysis (May 2025) of 57,399 available influenza A H1N1 and H3N2 sequences predicts that the VELO Respiratory Test would detect ≥99.8% of the isolates/variants available for analysis.

Influenza B inclusivity

Table 6 Inclusivity results for influenza B strains

Virus	Strain	Concentration	Number detected/number tested (% detected)
Influenza B	B/Lee/1940	26.4 CEID50/swab	3/3 (100%)
Influenza B (Victoria Lineage)	B/Michigan/1/2021	9,000 cp/swab	3/3 (100%)
Influenza B (Victoria Lineage)	B/Brisbane/60/2008	26.4 CEID50/swab	3/3 (100%)
Influenza B	B/Nevada/03/2011	26.4 CEID50/swab	3/3 (100%)

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Virus	Strain	Concentration	Number detected/number tested (% detected)
Influenza B (Victoria Lineage)	B/Hong Kong/330/2001	26.4 CEID50/swab	3/3 (100%)
Influenza B (Yamagata Lineage)	B/Guangdong/120/00	3 TCID50/swab	3/3 (100%)
Influenza B (Yamagata Lineage)	B/Massachusetts/2/12	3 TCID50/swab	3/3 (100%)
Influenza B (Yamagata Lineage)	B/Texas/6/11	3 TCID50/swab	3/3 (100%)

These influenza B strains are in addition to the B/Wisconsin/1/2010 and Malaysia/2506/04 strains used in the analytical sensitivity study.

In addition, in silico analysis (as of May 2025) of 12,876 influenza B sequences predicts that the VELO Respiratory Test would detect ≥98.4% of the isolates/variants available for analysis.

Cross-reactivity and Microbial Interference

Exclusivity (cross-reactivity) and microbial interference of the VELO Respiratory Test on the VELO Instrument were evaluated by wet testing 52 different viruses, bacteria and fungi that are common in respiratory infections. All samples were prepared in pooled negative nasal matrix in the presence of influenza A H3N2, influenza B and SARS-CoV-2 at 3x LoD (to determine microbial interference) or in the absence of viral targets (to determine exclusivity). Each sample type was tested in replicates of three (3). Exclusivity was determined if 0/3 replicates returned a detected result for each of the three (3) viral targets. Absence of microbial interference was determined if 3/3 replicates returned a detected result for each of the three (3) viral targets. None of the 52 microorganisms tested were found to show any interference or cross-reactivity with the VELO Respiratory Test at the concentrations tested.

Table 7 Exclusivity (cross reactivity) wet testing results

Microorganism	Concentration (per mL)	% negative agreement
Adenovirus Type 1	3.45 x10⁶ TCID50	100% (0/3 detected)
Adenovirus Type 7	3.37 x10⁶ TCID50	100% (0/3 detected)
Adenovirus Type 10	9.6 x10⁵ TCID50	100% (0/3 detected)
Adenovirus Type 21	2 x10⁵ TCID50	100% (0/3 detected)
Human Coronavirus OC43	2 x10⁵ TCID50	100% (0/3 detected)
Human Coronavirus 229E	2 x10⁵ copies	100% (0/3 detected)
Human Coronavirus NL63	2.13x10⁵ TCID50	100% (0/3 detected)
Human Coronavirus HKU1 ᵃ	2 x10⁵ copies	100% (0/3 detected)

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Microorganism	Concentration (per mL)	% negative agreement
Cytomegalovirus	2.02 x10⁵ TCID50	100% (0/3 detected)
Enterovirus Coxsackievirus CV-A16	2 x10⁵ TCID50	100% (0/3 detected)
Enterovirus D68	2 x10⁵ TCID50	100% (0/3 detected)
Enterovirus Type 71	2 x10⁵ TCID50	100% (0/3 detected)
Epstein Barr Virus	2 x10⁵ TCID50	100% (0/3 detected)
Human parainfluenza Type 1	1.6 x10⁵ TCID50	100% (0/3 detected)
Human parainfluenza Type 2	2 x10⁵ TCID50	100% (0/3 detected)
Human parainfluenza Type 3	2 x10⁵ TCID50	100% (0/3 detected)
Human parainfluenza Type 4	2 x10⁵ TCID50	100% (0/3 detected)
Measles	1.9 x10⁵ TCID50	100% (0/3 detected)
MERS-CoV ᵇ	NA ᵈ	100% (0/3 detected)
Human Metapneumovirus Type 1A	2 x10⁵ TCID50	100% (0/3 detected)
Mumps virus	1.1 x10⁵ TCID50	100% (0/3 detected)
Respiratory syncytial virus A1998/3-2	2 x10⁵ TCID50	100% (0/3 detected)
Respiratory syncytial virus A Long	2 x10⁵ TCID50	100% (0/3 detected)
Respiratory syncytial virus B	2 x10⁵ TCID50	100% (0/3 detected)
Rhinovirus A50, A2	1.33 x10⁵ TCID50	100% (0/3 detected)
Rhinovirus 20, 15-CV19	2 x10⁵ TCID50	100% (0/3 detected)
Aspergillus fumigatus	4.65 x10⁵ CFU	100% (0/3 detected)
Aspergillus niger	2 x10⁶ CFU	100% (0/3 detected)
Bordatella parapertussis	2 x10⁶ CFU	100% (0/3 detected)
Bordetella pertussis	2 x10⁶ CFU	100% (0/3 detected)
Candida albicans	2 x10⁶ CFU	100% (0/3 detected)
Chlamydia pneumoniae	2 x10⁶ IFU	100% (0/3 detected)
Corynebacterium xerosis	2 x10⁶ CFU	100% (0/3 detected)
Escherichia coli	2 x10⁶ CFU	100% (0/3 detected)
Fusobacterium necrophorum	2 x10⁶ CFU	100% (0/3 detected)
Hemophilus influenzae	2 x10⁶ CFU	100% (0/3 detected)
Klebsiella pneumoniae	2 x10⁶ CFU	100% (0/3 detected)

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Microorganism	Concentration (per mL)	% negative agreement
Lactobacillus acidophilus	5.79 x10⁶ CFU	100% (0/3 detected)
Legionella pneumophila	2 x10⁶ CFU	100% (0/3 detected)
Moraxella catarrhalis	2 x10⁶ CFU	100% (0/3 detected)
Mycoplasma genitalium ᵇ	2 x10⁶ CFU	100% (0/3 detected)
Mycobacterium tuberculosis ᶜ	2 x10⁶ copies	100% (0/3 detected)
Mycoplasma pneumoniae	2 x10⁶ CCU	100% (0/3 detected)
Neisseria meningitidis	2 x10⁶ CFU	100% (0/3 detected)
Neisseria mucosa	2 x10⁶ CFU	100% (0/3 detected)
Pneumocystis jirovecii (PJP) ᵃ	2 x10⁶ copies	100% (0/3 detected)
Pseudomonas aeruginosa	2 x10⁶ CFU	100% (0/3 detected)
Staphylococcus aureus	2 x10⁶ CFU	100% (0/3 detected)
Staphylococcus epidermis	2 x10⁶ CFU	100% (0/3 detected)
Streptococcus pneumoniae	2 x10⁶ CFU	100% (0/3 detected)
Streptococcus pyogenes	2 x10⁶ CFU	100% (0/3 detected)

ᵃ Synthetic nucleic acid
ᵇ Inactivated whole organism
ᶜ Genomic nucleic acid
ᵈ Swabs contrived with 8µl of NATrol™ MERS-CoV Stock (Ct 25.7)

Table 8 Microbial interference wet testing results

Microorganism	Concentration (per mL)	3x LoD SARS-CoV-2/Influenza A/Influenza B % positive agreement
Adenovirus Type 1	3.45 x10⁶ TCID50	100% (3/3 detected)
Adenovirus Type 7	3.37 x10⁶ TCID50	100% (3/3 detected)
Adenovirus Type 10	9.6 x10⁵ TCID50	100% (3/3 detected)
Adenovirus Type 21	2 x10⁵ TCID50	100% (3/3 detected)
Human Coronavirus OC43	2 x10⁵ TCID50	100% (3/3 detected)
Human Coronavirus 229E	2 x10⁵ copies	100% (3/3 detected)
Human Coronavirus NL63	2.13x10⁵ TCID50	100% (3/3 detected)
Human Coronavirus HKU1 ᵃ	2 x10⁵ copies	100% (3/3 detected)
Cytomegalovirus	2.02 x10⁵ TCID50	100% (3/3 detected)

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Microorganism	Concentration (per mL)	3x LoD SARS-CoV-2/Influenza A/Influenza B % positive agreement
Enterovirus Coxsackievirus CV-A16	2 x10⁵ TCID50	100% (3/3 detected)
Enterovirus D68	2 x10⁵ TCID50	100% (3/3 detected)
Enterovirus Type 71	2 x10⁵ TCID50	100% (3/3 detected)
Epstein Barr Virus	2 x10⁵ TCID50	100% (3/3 detected)
Human parainfluenza Type 1	1.6 x10⁵ TCID50	100% (3/3 detected)
Human parainfluenza Type 2	2 x10⁵ TCID50	100% (3/3 detected)
Human parainfluenza Type 3	2 x10⁵ TCID50	100% (3/3 detected)
Human parainfluenza Type 4	2 x10⁵ TCID50	100% (3/3 detected)
Measles	1.9 x10⁵ TCID50	100% (3/3 detected)
MERS-CoV ᵇ	NA ᵈ	100% (3/3 detected)
Human Metapneumovirus Type 1A	2 x10⁵ TCID50	100% (3/3 detected)
Mumps virus	1.1 x10⁵ TCID50	100% (3/3 detected)
Respiratory syncytial virus A1998/3-2	2 x10⁵ TCID50	100% (3/3 detected)
Respiratory syncytial virus A Long	2 x10⁵ TCID50	100% (3/3 detected)
Respiratory syncytial virus B	2 x10⁵ TCID50	100% (3/3 detected)
Rhinovirus A50, A2	1.33 x10⁵ TCID50	100% (3/3 detected)
Rhinovirus 20, 15-CV19	2 x10⁵ TCID50	100% (3/3 detected)
Aspergillus fumigatus	4.65 x10⁵ CFU	100% (3/3 detected)
Aspergillus niger	2 x10⁶ CFU	100% (3/3 detected)
Bordatella parapertussis	2 x10⁶ CFU	100% (3/3 detected)
Bordetella pertussis	2 x10⁶ CFU	100% (3/3 detected)
Candida albicans	2 x10⁶ CFU	100% (3/3 detected)
Chlamydia pneumoniae	2 x10⁶ IFU	100% (3/3 detected)
Corynebacterium xerosis	2 x10⁶ CFU	100% (3/3 detected)
Escherichia coli	2 x10⁶ CFU	100% (3/3 detected)
Fusobacterium necrophorum	2 x10⁶ CFU	100% (3/3 detected)
Hemophilus influenzae	2 x10⁶ CFU	100% (3/3 detected)
Klebsiella pneumoniae	2 x10⁶ CFU	100% (3/3 detected)
Lactobacillus acidophilus	5.79 x10⁶ CFU	100% (3/3 detected)

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Microorganism	Concentration (per mL)	3x LoD SARS-CoV-2/Influenza A/Influenza B % positive agreement
Legionella pneumophila	2 x10⁶ CFU	100% (3/3 detected)
Moraxella catarrhalis	2 x10⁶ CFU	100% (3/3 detected)
Mycoplasma genitalium ᵇ	2 x10⁶ CFU	100% (3/3 detected)
Mycobacterium tuberculosis ᶜ	2 x10⁶ copies	100% (3/3 detected)
Mycoplasma pneumoniae	2 x10⁶ CCU	100% (3/3 detected)
Neisseria meningitidis	2 x10⁶ CFU	100% (3/3 detected)
Neisseria mucosa	2 x10⁶ CFU	100% (3/3 detected)
Pneumocystis jirovecii (PJP) ᵃ	2 x10⁶ copies	100% (3/3 detected)
Pseudomonas aeruginosa	2 x10⁶ CFU	100% (3/3 detected)
Staphylococcus aureus	2 x10⁶ CFU	100% (3/3 detected)
Staphylococcus epidermis	2 x10⁶ CFU	100% (3/3 detected)
Streptococcus pneumoniae	2 x10⁶ CFU	100% (3/3 detected)
Streptococcus pyogenes	2 x10⁶ CFU	100% (3/3 detected)
Streptococcus salivaris	2 x10⁶ CFU	100% (3/3 detected)

ᵃ Synthetic nucleic acid
ᵇ Inactivated whole organism
ᶜ Genomic nucleic acid
ᵈ Swabs contrived with 8µl of NATrol™ MERS-CoV Stock (Ct 25.7)

Competitive Interference

The impact of competitive interference, caused by co-infections with on-target analytes, was evaluated for the VELO Respiratory Test on the VELO Instrument by testing contrived samples consisting of individual SARS-CoV-2, influenza A or influenza B strains at high concentrations in the presence of either one or both of the other target viruses at 3x LoD prepared in pooled negative nasal matrix. For this study, competitive interference was assessed using one strain each of SARS-CoV-2 (USA-WA1/2020) and influenza B (B/Wisconsin/1/2010) and two strains of influenza A (A/ PR/8/34 -H1N1 strain and A/Hong Kong/8/68 -H3N2 strain). Testing was performed in triplicate. Absence of competitive interference was determined if all replicates for the low concentration (3x LoD) target(s) yielded positive results.

The study showed that influenza B at 1x10⁶ copies/swab inhibited detection of influenza A (H1N1) at 3x LoD, in the presence of SARS-CoV-2 at 3x LoD. Subsequent testing using samples consisting of influenza A (H1N1) at 3x LoD, in the presence of influenza B at concentrations ranging from 1x10⁵ – 1.5x10⁶ copies/swab demonstrated no competitive interference. No competitive interference was observed for the other potential co-infections evaluated at the concentrations tested.

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Table 9 Competitive Interference Results Summary

Viral Targets in Sample			Detection Rate
Influenza A	Influenza B	SARS-CoV-2	Influenza A	Influenza B	SARS-CoV-2
H3N2 1x10⁶ cp/swab*	3xLoD	3x LoD	3/3	3/3	3/3
H1N1 1x10⁶ cp/swab	3xLoD	3x LoD	3/3	3/3	3/3
H3N2 3x LoD	N/A	1x10⁶ cp/swab	3/3	0/3	3/3
H1N1 3x LoD	3x LoD	1x10⁶ cp/swab	3/3	3/3	3/3
H3N2 3x LoD	1x10⁶ cp/swab	N/A	3/3	3/3	0/3
H1N1 3x LoD	1x10⁶ cp/swab	3x LoD	2/3	3/3	3/3
H1N1 3x LoD	1.5x10⁶ cp/swab	N/A	3/3	3/3	0/3
	1x10⁶ cp/swab	N/A	3/3	3/3	0/3
	9x10⁵ cp/swab	N/A	3/3	3/3	0/3
	1x10⁵ cp/swab	N/A	3/3	3/3	0/3

cp/swab = copies/swab

Carry-over

An analytical study was performed to assess potential carry-over or cross contamination of the VELO Respiratory Test by running alternating no template (negative) and high positive samples on five (5) VELO instruments. The high positive samples consisted of a single viral target (influenza A H1N1, influenza A H3N2, influenza B or SARS-CoV-2) at 1x10⁶ copies/swab prepared in pooled negative nasal matrix. Samples were tested across five (5) VELO instruments, where a no template control (NTC) sample was followed by a positive sample alternating eight (8) times before running a final negative sample to give 17 runs per VELO instrument. All high positive samples gave detected results for the viral target present in the sample, while 93% of the NTC samples gave the expected Invalid result. These results demonstrated that there is an acceptable, low likelihood of cross-contamination between samples when the VELO Respiratory Test Cartridge is performed on the VELO instrument according to the instructions for use.

Endogenous and Exogenous Interference

Endogenous and exogenous substances that may be found in the upper respiratory tract were evaluated to determine if they interfere with the performance of the VELO Respiratory Test on the VELO Instrument. All samples were prepared in negative nasal matrix and tested in replicates of three (3) positive and three (3) negative samples. Positive samples contained an individual exogenous or endogenous substance plus 3x LoD of SARS-CoV-2, influenza A and influenza B. Negative samples contained an individual exogenous or endogenous substance in negative nasal matrix. No interference was seen at the concentrations indicated.

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Table 10 Summary of results for Interfering substance testing

Interfering Substance	Product	Active Ingredient	Concentration at which the interference is not observed
Blood (human)	N/A	None Specified	10% v/v ᵃ
Leukocytes	N/A	None Specified	1 x 10⁶ cells/swab
Mucin: bovine submaxillary gland, type I-S		Purified mucin protein	2% v/v ᵇ
Nasal spray or drops	Zicam Intense Sinus Relief	Oxymetazoline HCl (0.05% w/v)	15% v/v
		Menthol
	Phenylephrine	Phenylephrine	0.03 µg/ml
	Calpol saline nasal spray	Sodium chloride (0.9%) with preservatives	15% v/v
Nasal corticosteroids	Pirinase Hayfever once daily spray	Fluticasone propionate (50 µg/spray)	15% v/v
	Boots Adult Hay fever relief	Beclomethasone (50 µg/spray)	15% v/v
	Dexamethasone	Dexamethasone	12 µg/ml
	Flunisolide	Flunisolide	16 µg/swab
	Triamcinolone	Triamcinolone	10 µg/swab ᶜ
	Benacort Hayfever Relief Nasal Spray	Budesonide (64 µg/spray)	15% v/v
	Clarinaze Allergy Control 0.05% Nasal Spray	Mometasone furoate (50 µg/spray)	15% v/v

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Interfering Substance	Product	Active Ingredient	Concentration at which the interference is not observed
Nasal gel	Zicam, Powerful Allergy Relief	Sulfur	15% v/v
		Luffa opperculata,
		Galphimia glauca
		Histaminum hydrochloricum
Sore throat and cough lozenges	Ultra Chloraseptic Spray	Benzocaine (0.71%)	15% v/v
Anti-viral drugs	N/A	Zanamivir	6.0 mg/ml ᵈ
	N/A	Oseltamivir phosphate	0.4 µg/ml
Antibiotics	N/A	Mupirocin	1.5 µg/ml
	N/A	Tobramycin	33 µg/ml
Zinc (common ingredient in nasal sprays)	Zinc chloride	Zinc	0.1 mg/ml
Nicotine or Tobacco	Nicorette (Nicotine 0.5 mg/spray)	Nicotine	15% v/v
Decongestant	Otravine Extra Dual Relief Nasal Spray	Xylometazoline hydrochloride	13.5% v/v ᵃ
		Ipratropium bromide

ᵃ Potential interference observed with 15% v/v.
ᵇ Potential interference observed with 5 and 2.5% w/v.
ᶜ Potential interference observed 11 µg/swab.
ᵈ Potential interference observed with 7.5 and 6.75 mg/ml.

Note: FluMist Quadrivalent was not evaluated as cross-reactivity with targets is expected.

Precision (within-lab repeatability)

The repeatability of the VELO Respiratory Test to detect SARS-CoV-2, influenza A and influenza B was evaluated over a 12-day period at a single site with two (2) operators conducting the testing. A 3-member panel was evaluated consisting of a true negative (no analyte), low positive (2x LoD of all three [3] targets), and a moderate positive (4x LoD of all three [3] targets). The negative samples were contrived using simulated respiratory matrix and the positive samples were contrived using simulated respiratory matrix co-spiked with SARS-CoV-2, influenza A, and influenza B. The test samples were randomized and blinded to the operator running the VELO Instrument. The study was conducted using one (1) lot of VELO Respiratory Test cartridges tested by two (2) operators each performing two (2) replicates per run and two (2) runs per day. The study was performed over 12 days, totalling 96 replicates per panel member and 288 tests in total. Four (4) VELO Instruments were utilized during this study. The VELO Respiratory Test reported the expected

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positive results for panel members in 97.9%-100% of samples at 2x LoD, 99.0%-100% of samples at 4x LoD and the expected negative results in 100% of negative samples.

Table 11 Summary of repeatability results by operator

Analyte	Panel member	% Agreement with expected results (95% CI)
		Operator 1	Operator 2	Overall
SARS-CoV-2	Negative	100% (48/48) (92.6-100%)	100% (48/48) (92.6-100%)	100% (96/96) (96.2-100%)
	Low positive (2x LoD)	97.9% (47/48) (89.1-99.6%)	97.9% (47/48) (89.1-99.6%)	97.9% (94/96) (92.7-99.4%)
	Moderate positive (4x LoD)	97.9% (47/48) (89.1-99.6%)	100% (48/48) (92.6-100%)	99.0% (95/96) (94.3-99.8%)
Influenza A	Negative	100% (48/48) (92.6-100%)	100% (48/48) (92.6-100%)	100% (96/96) (96.2-100%)
	Low positive (2x LoD)	100% (48/48) (92.6-100%)	97.9% (47/48) (89.1-99.6%)	99.0% (95/96) (94.3-99.8%)
	Moderate positive (4x LoD)	100% (48/48) (92.6-100%)	100% (48/48) (92.6-100%)	100% (96/96) (96.2-100%)
Influenza B	Negative	100% (48/48) (92.6-100%)	100% (48/48) (92.6-100%)	100% (96/96) (96.2-100%)
	Low positive (2x LoD)	100% (48/48) (92.6-100%)	97.9% (47/48) (89.1-99.6%)	99.0% (95/96) (94.3-99.8%)
	Moderate positive (4x LoD)	100% (48/48) (92.6-100%)	100% (48/48) (92.6-100%)	100% (96/96) (96.2-100%)

Table 12 Summary of repeatability results by VELO Instrument

Analyte	Panel member	% Agreement with expected results (95% CI)
		Instrument 1	Instrument 2	Instrument 3	Instrument 4	Overall
SARS-CoV-2	Negative	100% (25/25) (86.7-100%)	100% (22/22) (85.1-100%)	100% (24/24) (86.2-100%)	100% (25/25) (86.7-100%)	100% (96/96) (96.2-100%)
	Low positive (2x LoD)	95.8% (23/24) (79.8-99.3%)	95.8% (23/24) (79.8-99.3%)	100% (25/25) (86.7-100%)	100% (23/23) (85.7-100%)	97.9% (94/96) (92.7-99.4%)
	Moderate positive (4x LoD)	100% (23/23) (85.7-100%)	100% (23/23) (85.7-100%)	96.2% (25/26) (81.1-99.3%)	100% (24/24) (86.2-100%)	99.0% (95/96) (94.3-99.8%)
Influenza A	Negative	100% (25/25) (86.7-100%)	100% (22/22) (85.1-100%)	100% (24/24) (86.2-100%)	100% (25/25) (86.7-100%)	100% (96/96) (96.2-100%)
	Low positive (2x LoD)	95.8% (23/24) (79.8-99.3%)	100% (24/24) (86.2-100%)	100% (25/25) (86.7-100%)	100% (23/23) (85.7-100%)	99.0% (95/96) (94.3-99.8%)

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Analyte	Panel member	% Agreement with expected results (95% CI)
		Instrument 1	Instrument 2	Instrument 3	Instrument 4	Overall
	Moderate positive (4x LoD)	100% (23/23) (85.7-100%)	100% (23/23) (85.7-100%)	100% (26/26) (87.1-100%)	100% (24/24) (86.2-100%)	100% (96/96) (96.2-100%)
Influenza B	Negative	100% (25/25) (86.7-100%)	100% (22/22) (85.1-100%)	100% (24/24) (86.2-100%)	100% (25/25) (86.7-100%)	100% (96/96) (96.2-100%)
	Low positive (2x LoD)	100% (24/24) (86.2-100%)	100% (24/24) (86.2-100%)	96.0% (24/25) (80.5-99.3%)	100% (23/23) (85.7-100%)	99.0% (95/96) (94.3-99.8%)
	Moderate positive (4x LoD)	100% (23/23) (85.7-100%)	100% (23/23) (85.7-100%)	100% (26/26) (87.1-100%)	100% (24/24) (86.2-100%)	100% (96/96) (96.2-100%)

Clinical Performance

The clinical performance of the VELO Respiratory Test for the detection of influenza A, influenza B, and SARS-CoV-2 was evaluated using paired prospective clinical anterior nasal swab (ANS) specimens collected from individuals with signs and symptoms of upper respiratory viral infection. Testing of clinical samples was performed with the VELO Respiratory Test in nine (9) CLIA waived healthcare facilities (e.g., physician offices, primary care / outpatient clinics, and urgent care centers). The results of all three (3) viral targets (Instrument software v7.2.0) were compared to results from an FDA-cleared, CLIA waived RT-PCR assay (Comparator).

Prospective clinical specimens were collected and tested December 2024–March 2025. Initial enrollment in the prospective clinical study included 1815 anterior nasal swab specimens. Of these, 97 specimens were excluded from the performance analysis for major protocol deviations. Table 13 provides a summary of the demographic information for the remaining 1718 subjects enrolled in the clinical study.

Table 13 Subject Demographics- Prospective Symptomatic Population

Characteristics	Symptomatic Subjects
Total, N	1718
Age (years)
Mean	41.0
Standard Deviation	17.97
Median	41
Range (minimum – maximum)	0 – 92
Age Group (Years), n (%)
≤ 12	78 (4.5%)
13 to ≤18	96 (5.6%)
19 to <55	1130 (65.8%)
≥55	414 (24.1%)

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Characteristics	Symptomatic Subjects
Sex, n (%)
Male	677 (39.4%)
Female	1041 (60.6%)
Ethnicity, n (%)
Hispanic / Latino	191 (11.1%)
Not Hispanic / Not Latino	1516 (88.2%)
Note Reported	10 (0.6%)
Unknown	1 (0.1%)
Race, n (%)
American Indian / Alaskan Native	30 (1.7%)
Asian	20 (1.2%)
Black / African-American	389 (22.6%)
Native Hawaiian / Pacific Islander	3 (0.2%)
White	1225 (71.3%)
Other	12 (0.7%)
More than one race	32 (1.9%)
Not Reported	7 (0.4%)

In addition, of the 1718 evaluable specimens collected in the prospective study a further six (6) specimens were excluded either due to invalid comparator tests results for all three (3) target analytes or the patient subsequently withdrew from the study. Of the remaining 1712 evaluable specimens from the prospective clinical study, additional exclusions due to invalid VELO Respiratory Test results upon retest resulted in the following final sample sizes for the performance evaluation, influenza A 1,677 (35 excluded), influenza B 1,665 (47 excluded), and SARS-CoV-2 1,670 (42 excluded). Three (3) influenza A / SARS-CoV-2 coinfections were detected by the Comparator.

Influenza A

For prospective symptomatic subjects, 342 ANS specimens tested positive for influenza A with both the VELO Respiratory Test and Comparator. A total of 1,295 ANS specimens tested negative for influenza A with both the VELO Respiratory Test and Comparator. Discrepant results were forwarded on to a central reference laboratory for testing on a second FDA-cleared, CLIA waived RT-PCR assay (secondary reference test). Results of discrepant analysis are included as a footnote to the results table.

For influenza A, the results of the clinical performance evaluation using ANS specimens from prospective symptomatic subjects demonstrated 92.4% positive percent agreement (PPA) (342/370; 95% score CI: 89.3% - 94.7%) and 99.1% negative percent agreement (NPA) (1,295/1,307; 95% score CI: 98.4% - 99.5%) as compared to the Comparator.

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Table 14 Influenza A Results: Prospective, Paired ANS Specimens

Influenza A Prospective, Paired ANS Specimens	Comparator
	Positive	Negative	Total
VELO Respiratory Test
Positive	342 ᵃ	12 ᵇ	354
Negative	28 ᶜ	1,295	1,323
Total	370	1,307	1,677

PPA 92.4% 95% CI: 89.3 – 94.7%
NPA 99.1% 95% CI: 98.4 – 99.5%

ᵃ Includes three (3) Influenza A / SARS-CoV-2 coinfections.
ᵇ Three (3) discrepant specimens tested positive on secondary reference testing.
ᶜ Two (2) discrepant specimens tested negative on secondary reference testing.

Influenza B

For prospective symptomatic subjects, 14 ANS specimens tested positive for influenza B with both the VELO Respiratory Test and Comparator. A total of 1,646 ANS specimens tested negative for influenza B with both the VELO Respiratory Test and Comparator. Discrepant results were forwarded on to a central reference laboratory for testing on a second FDA-cleared, CLIA waived RT-PCR assay. Results of discrepant analysis are included as a footnote to the results table.

For influenza B, the results of the clinical performance evaluation using ANS specimens from prospective symptomatic subjects demonstrated 93.3% PPA (14/15; 95% score CI: 70.2% - 98.8%) and 99.8% NPA (1,646/1,650; 95% score CI: 99.4% - 99.9%) as compared to the Comparator.

Table 15 Influenza B Results: Prospective, Paired ANS Specimens

Influenza B Prospective, Paired ANS Specimens	Comparator
	Positive	Negative	Total
VELO Respiratory Test
Positive	14	4	18
Negative	1 ᵃ	1,646	1,647
Total	15	1,650	1,665

PPA 93.3% 95% CI: 70.2 – 98.8%
NPA 99.8% 95% CI: 99.4 – 99.9%

ᵃ One (1) discrepant specimen tested negative on secondary reference testing.

Due to the observed low prevalence of circulating influenza B virus, prospective influenza B testing was supplemented with paired retrospective clinical ANS specimens collected from individuals with signs and symptoms of influenza infection during the 2023-2024 North American respiratory season. Frozen paired positive and negative ANS (n=98) specimens prospectively obtained during the 2023-2024 flu season were distributed to a single site and tested during their daily workflow over a period of five (5) days.

For retrospective, paired specimens, 15 ANS specimens tested positive for influenza B with both the VELO Respiratory Test and a comparator RT-PCR assay. A total of 82 ANS specimens tested negative for influenza B with both the VELO Respiratory Test and comparator assay.

For influenza B, the results of the clinical performance evaluation using frozen, paired positive and negative ANS specimens from retrospective symptomatic subjects demonstrated 93.8% PPA (15/16; 95% score CI: 71.7% - 98.9%) and 100% NPA (82/82; 95% score CI: 95.5-100%) as compared to the comparator assay.

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Table 16 Influenza B Results: Retrospective, Paired ANS Specimens

Influenza B Retrospective, Paired ANS Specimens	Comparator
	Positive	Negative	Total
VELO Respiratory Test
Positive	15	0	15
Negative	1	82	83
Total	16	82	98

PPA 93.8% 95% CI: 71.7 – 98.9%
NPA 100% 95% CI: 95.5 – 100%

SARS-CoV-2

For prospective symptomatic subjects, 150 ANS specimens tested positive for SARS-CoV-2 with both the VELO Respiratory Test and Comparator; six (6) SARS-CoV-2-positive specimens tested negative for SARS-CoV-2 with the VELO Respiratory Test. A total of 1,507 ANS specimens tested negative for SARS-CoV-2 with both the VELO Respiratory Test and Comparator; seven (7) SARS-CoV-2-negative specimens tested

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positive for SARS-CoV-2 with the VELO Respiratory Test. Discrepant results were forwarded on to a central reference laboratory for testing on a second FDA-cleared, CLIA waived RT-PCR assay. Results of discrepant analysis are included as a footnote to the results table.

For SARS-CoV-2, the results of the clinical performance evaluation using ANS specimens from prospective symptomatic subjects demonstrated 96.2% PPA (150/156; 95% score CI: 91.9% - 98.2%) and 99.5% NPA (1,507/1,514; 95% score CI: 99.1% - 99.8%) as compared to the Comparator.

Table 17 SARS-CoV-2 Results: Prospective, Paired ANS Specimens

SARS-CoV-2 Prospective, Paired ANS Specimens	Comparator
	Positive	Negative	Total
VELO Respiratory Test
Positive	150 ᵃ	7	157
Negative	6 ᵇ	1,507	1,513
Total	156	1,514	1,670

PPA 96.2% 95% CI: 91.9 – 98.2%
NPA 99.5% 95% CI: 99.1 – 99.8%

ᵃ Includes three (3) Influenza A / SARS-CoV-2 coinfections.
ᵇ One (1) discrepant specimen tested negative on secondary reference testing.

Expected Values/Reference Range

For the prospective clinical performance evaluation of VELO Respiratory Test for use on the VELO Instrument ANS specimens from 1,718 evaluable subjects were freshly collected and tested at nine (9) point-of-care clinical sites in the United States between December 2024 – March 2025. Expected value (as determined by VELO Respiratory Test) summaries for prospective specimens, stratified by specimen collection/testing site are presented for influenza A, influenza B and SARS-CoV-2 targets:

Table 18 Expected value summary by clinical site for prospective clinical evaluation for influenza A from December 2024 – March 2025

Clinical Site ID	Site Location	ANS Specimens
		Total No.	No. Positive for Influenza A	Expected Value
Overall		1,677	354	21.1%
1	Riverside, CA	138	26	18.8%

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Clinical Site ID	Site Location	ANS Specimens
		Total No.	No. Positive for Influenza A	Expected Value
2	Orange City, FL	73	14	19.2%
3	DeLand, FL	20	1	5.0%
4	Brooklyn, NY	5	0	0.0%
5	Gulfport, MS	232	13	5.6%
6	Birmingham, AL	380	121	31.8%
7	Birmingham, AL	600	157	26.2%
8	Tulsa, OK	215	19	8.8%
9	Salt Lake City, UT	14	3	21.4%

Table 19 Expected value summary by clinical site for prospective clinical evaluation for influenza B from December 2024 – March 2025

Clinical Site ID	Site Location	ANS Specimens
		Total No.	No. Positive for Influenza B	Expected Value
Overall		1,665	18	1.1%
1	Riverside, CA	138	6	4.3%
2	Orange City, FL	73	1	1.4%
3	DeLand, FL	20	0	0.0%
4	Brooklyn, NY	5	0	0.0%
5	Gulfport, MS	232	0	0.0%
6	Birmingham, AL	377	5	1.3%
7	Birmingham, AL	592	6	1.0%
8	Tulsa, OK	215	0	0.0%
9	Salt Lake City, UT	13	0	0.0%

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Table 20 Expected value summary by clinical site for prospective clinical evaluation for SARS-CoV-2 from December 2024 – March 2025

Clinical Site ID	Site Location	ANS Specimens
		Total No.	No. Positive for SARS-CoV-2	Expected Value
Overall		1,670	157	9.4%
1	Riverside, CA	138	1	0.7%
2	Orange City, FL	73	3	4.1%
3	DeLand, FL	20	2	10.0%
4	Brooklyn, NY	5	0	0.0%
5	Gulfport, MS	232	5	2.2%
6	Birmingham, AL	379	49	12.9%
7	Birmingham, AL	595	90	15.1%
8	Tulsa, OK	215	6	2.8%
9	Salt Lake City, UT	13	1	7.7%

Invalid rates

Of the 1716 tests performed for these subjects, there were a total of 90 (5.24%, 95% CI: 4.29 - 6.40%) invalid results obtained on initial testing with the VELO Respiratory Test run on the VELO Instrument. Of these, 34 were invalid upon retesting, for a final invalid rate of 1.98% (34/1716) with 95% CI: 1.42 – 2.76%.

Reproducibility

The reproducibility of the VELO Respiratory Test to detect SARS-CoV-2, Influenza A and Influenza B was established at three (3) external CLIA-waived sites using contrived nasal swabs and a 3-member panel consisting of a true negative (no analyte-matrix only), low positive (2x LoD of all three [3] targets), and a moderate positive (4x LoD of all three [3] targets). The negative samples were contrived using simulated respiratory matrix and the positive samples were contrived using simulated respiratory matrix co-spiked with SARS-CoV-2, influenza A and influenza B. The study was conducted using three (3) lots of VELO Respiratory Test cartridges tested by nine (9) untrained operators over three (3) sites (3 operators per site). Each panel member was tested in one (1) replicate/run with two (2) runs per day by each of the nine (9) operators. The study was performed over five (5) non-consecutive days, totalling 90 replicates per panel member and 270 tests in total. The VELO Respiratory Test reported the expected positive results for panel members in 97.8%-100% of samples at 2x LoD, 100% of samples at 4x LoD and the expected negative results in 100% of negative samples.

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Table 21 Summary of reproducibility results by site

Analyte	Panel member	% Agreement with expected results (95% CI)
		Site A	Site B	Site C	Overall
SARS-CoV-2	Negative	100% (30/30) (88.7-100%)	100% (30/30) (88.7-100%)	100% (30/30) (88.7-100%)	100% (90/90) (95.9-100%)
	Low positive (2x LoD)	100% (30/30) (88.7-100%)	96.7% (29/30) (83.3-99.4%)	96.7% (29/30) (83.3-99.4%)	97.8% (88/90) (92.3-99.4%)
	Moderate positive (4x LoD)	100% (30/30) (88.7-100%)	100% (30/30) (88.7-100%)	100% (30/30) (88.7-100%)	100% (90/90) (95.9-100%)
Influenza A	Negative	100% (30/30) (88.7-100%)	100% (30/30) (88.7-100%)	100% (30/30) (88.7-100%)	100% (90/90) (95.9-100%)
	Low positive (2x LoD)	100% (30/30) (88.7-100%)	100% (30/30) (88.7-100%)	100% (30/30) (88.7-100%)	100% (90/90) (95.9-100%)
	Moderate positive (4x LoD)	100% (30/30) (88.7-100%)	100% (30/30) (88.7-100%)	100% (30/30) (88.7-100%)	100% (90/90) (95.9-100%)
Influenza B	Negative	100% (30/30) (88.7-100%)	100% (30/30) (88.7-100%)	100% (30/30) (88.7-100%)	100% (90/90) (95.9-100%)
	Low positive (2x LoD)	100% (30/30) (88.7-100%)	100% (30/30) (88.7-100%)	100% (30/30) (88.7-100%)	100% (90/90) (95.9-100%)
	Moderate positive (4x LoD)	100% (30/30) (88.7-100%)	100% (30/30) (88.7-100%)	100% (30/30) (88.7-100%)	100% (90/90) (95.9-100%)

Table 22 Summary of reproducibility results by operator

Analyte	Panel member	% Agreement with expected results (95% CI)
		A1	A2	A3	Overall
SARS-CoV-2	Negative	100% (10/10) (72.3 -100%)	100% (10/10) (72.3 -100%)	100% (10/10) (72.3 -100%)	100% (30/30) (88.7-100%)
	Low positive (2x LoD)	100% (10/10) (72.3 -100%)	100% (10/10) (72.3 -100%)	100% (10/10) (72.3 -100%)	100% (30/30) (88.7-100%)
	Moderate positive (4x LoD)	100% (10/10) (72.3 -100%)	100% (10/10) (72.3 -100%)	100% (10/10) (72.3 -100%)	100% (30/30) (88.7-100%)
Influenza A	Negative	100% (10/10) (72.3 -100%)	100% (10/10) (72.3 -100%)	100% (10/10) (72.3 -100%)	100% (30/30) (88.7-100%)
	Low positive (2x LoD)	100% (10/10) (72.3 -100%)	100% (10/10) (72.3 -100%)	100% (10/10) (72.3 -100%)	100% (30/30) (88.7-100%)
	Moderate positive (4x LoD)	100% (10/10) (72.3 -100%)	100% (10/10) (72.3 -100%)	100% (10/10) (72.3 -100%)	100% (30/30) (88.7-100%)
Influenza B	Negative	100% (10/10) (72.3 -100%)	100% (10/10) (72.3 -100%)	100% (10/10) (72.3 -100%)	100% (30/30) (88.7-100%)

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Panel member	% Agreement with expected results (95% CI)
	A1	A2	A3	Overall
Low positive (2x LoD)	100% (10/10) (72.3 -100%)	100% (10/10) (72.3 -100%)	100% (10/10) (72.3 -100%)	100% (30/30) (887-100%)
Moderate positive (4x LoD)	100% (10/10) (72.3 -100%)	100% (10/10) (72.3 -100%)	100% (10/10) (72.3 -100%)	100% (30/30) (88.7-100%)

Analyte	Panel member	B1	B2	B3	Overall
SARS-CoV-2	Negative	100% (10/10) (72.3 -100%)	100% (10/10) (72.3 -100%)	100% (10/10) (72.3 -100%)	100% (30/30) (88.7-100%)
	Low positive (2x LoD)	100% (10/10) (72.3 -100%)	100% (10/10) (72.3 -100%)	90% (9/10) (59.6-98.2%)	96.67% (29/30) (88.3-99.4%)
	Moderate positive (4x LoD)	100% (10/10) (72.3 -100%)	100% (10/10) (72.3 -100%)	100% (10/10) (72.3-100%)	100% (30/30) (88.7-100%)
Influenza A	Negative	100% (10/10) (72.3 -100%)	100% (10/10) (72.3 -100%)	100% (10/10) (72.3 -100%)	100% (30/30) (88.7-100%)
	Low positive (2x LoD)	100% (10/10) (72.3 -100%)	100% (10/10) (72.3 -100%)	100% (10/10) (72.3 -100%)	100% (30/30) (88.7-100%)
	Moderate positive (4x LoD)	100% (10/10) (72.3 -100%)	100% (10/10) (72.3 -100%)	100% (10/10) (72.3 -100%)	100% (30/30) (88.7-100%)
Influenza B	Negative	100% (10/10) (72.3 -100%)	100% (10/10) (72.3 -100%)	100% (10/10) (72.3-100%)	100% (30/30) (88.7-100%)
	Low positive (2x LoD)	100% (10/10) (72.3 -100%)	100% (10/10) (72.3 -100%)	100% (10/10) (72.3 -100%)	100% (30/30) (88.7-100%)
	Moderate positive (4x LoD)	100% (10/10) (72.3 -100%)	100% (10/10) (72.3 -100%)	100% (10/10) (72.3 -100%)	100% (30/30) (88.7-100%)

Analyte	Panel member	C1	C2	C3	Overall
SARS-CoV-2	Negative	100% (10/10) (72.3 -100%)	100% (10/10) (72.3 -100%)	100% (10/10) (72.3 -100%)	100% (30/30) (88.7-100%)
	Low –positive (2x LoD)	100% (10/10) (72.3 -100%)	100% (10/10) (72.3 -100%)	90% (9/10) (59.6 -98.2%)	96.7% (29/30) (83.3-99.4%)
	Moderate positive (4x LoD)	100% (10/10) (72.3 -100%)	100% (10/10) (72.3 -100%)	100% (10/10) (72.3 -100%)	100% (30/30) (88.7-100%)
Influenza A	Negative	100% (10/10) (72.3 -100%)	100% (10/10) (72.3 -100%)	100% (10/10) (72.3 -100%)	100% (30/30) (88.7-100%)
	Low –positive (2x LoD)	100% (10/10) (72.3 -100%)	100% (10/10) (72.3 -100%)	100% (10/10) (72.3 -100%)	100% (30/30) (88.7-100%)
	Moderate positive (4x LoD)	100% (10/10) (72.3 -100%)	100% (10/10) (72.3 -100%)	100% (10/10) (72.3 -100%)	100% (30/30) (88.7-100%)
Influenza B	Negative	100% (10/10) (72.3 -100%)	100% (10/10) (72.3 -100%)	100% (10/10) (72.3 -100%)	100% (30/30) (88.7-100%)
	Low –positive (2x LoD)	100% (10/10) (72.3 -100%)	100% (10/10) (72.3 -100%)	100% (10/10) (72.3 -100%)	100% (30/30) (88.7-100%)
	Moderate positive (4x LoD)	100% (10/10) (72.3 -100%)	100% (10/10) (72.3 -100%)	100% (10/10) (72.3 -100%)	100% (30/30) (88.7-100%)

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Table 23 Summary of reproducibility results by cartridge lot

Analyte	Panel member	% Agreement with expected results (95% CI)
		Lot 1	Lot 2	Lot 3	Overall
SARS-CoV-2	Negative	100% (25/25) (86.7-100%)	100% (30/30) (88.7-100%)	100% (35/35) (90.1-100%)	100% (90/90) (95.9-100%)
	Low –positive (2x LoD)	96.7% (29/30) (83.3-99.4%)	96.7% (29/30) (83.3-99.4%)	100% (30/30) (88.7-100%)	97.8% (88/90) (92.3-99.4%)
	Moderate positive (4x LoD)	100% (33/33) (89.6-100%)	100% (31/31) (89.0-100%)	100% (26/26) (87.1-100%)	100% (90/90) (95.9-100%)
Influenza A	Negative	100% (25/25) (86.7-100%)	100% (30/30) (88.7-100%)	100% (35/35) (90.1-100%)	100% (90/90) (95.9-100%)
	Low –positive (2x LoD)	100% (30/30) (88.7-100%)	100% (30/30) (88.7-100%)	100% (30/30) (88.7-100%)	100% (90/90) (95.9-100%)
	Moderate positive (4x LoD)	100% (33/33) (89.6-100%)	100% (31/31) (89.0-100%)	100% (26/26) (87.1-100%)	100% (90/90) (95.9-100%)
Influenza B	Negative	100% (25/25) (86.7-100%)	100% (30/30) (88.7-100%)	100% (35/35) (90.1-100%)	100% (90/90) (95.9-100%)
	Low –positive (2x LoD)	100% (30/30) (88.7-100%)	100% (30/30) (88.7-100%)	100% (30/30) (88.7-100%)	100% (90/90) (95.9-100%)
	Moderate positive (4x LoD)	100% (33/33) (89.6-100%)	100% (31/31) (89.0-100%)	100% (26/26) (87.1-100%)	100% (90/90) (95.9-100%)

Flex Studies

Flex studies were performed to evaluate the robustness of the VELO Respiratory Test on the VELO Instrument. Variations in workflow and operating environment that may reasonably be expected to occur with untrained operators in the intended use CLIA-waived setting were evaluated. Results of the flex studies conducted demonstrate robust performance of the VELO Respiratory Test on the VELO Instrument under conditions of stress.

CONCLUSIONS

A comparison of the intended use, technological characteristics, and the results of analytical and clinical performance studies demonstrate that the VELO Respiratory Test is substantially equivalent to the predicate device.

Results of non-clinical analytical, clinical performance and usability studies demonstrate that the VELO Respiratory Test's suitability for CLIA Waived Testing environments. Results demonstrate the VELO Respiratory Test is so simple and accurate as to render the likelihood of erroneous results by the user negligible.

Regulation Number and Section

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

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