CLEARinse CTS is intended to collect and transport clinical nasal wash aspirate specimens that may contain influenza A or influenza B virus, by health care professionals, from the collection site to a laboratory or testing site. CLEARinse CTS specimens are suitable for use with legally marketed lateral flow immunoassays and molecular assays. CLEARinse CTS is for professional in-vitro diagnostic use.

Device Description

The product is an extension of the CLEARinse Pro device, a 510(k)-cleared (K082762) professional use medical device with indications for nasal lavage as well as mucus sample collection for subsequent testing. The CLEARinse Pro Handle and Instructions for Use are required for use of the CLEARinse CTS.

The CLEARinse CTS is comprised of two component assemblies: a revision to the 510(k)cleared Wash Head and a Transport Container to protect the Wash Head and specimen during transit. The CLEARinse CTS. used with the CLEARinse Pro Handle, will allow a nasal wash aspirate specimen to be collected by a medical professional, protected for transit, and ground shipped to a testing site. The user mounts the CLEARinse CTS Wash Head onto the CLEARinse Pro Handle and adds sterile saline as directed. The tip of the Wash Head is inserted into the patient's nostril and sterile saline is introduced. The irrigated saline and nasal secretions are then aspirated back into the Wash Head, collecting the specimen to be tested. The CLEARinse CTS Wash Head is removed from the Handle and placed into the Transport Container assembly (Container Cup, Filter Seal, Silicone Seal, and Container Lid) and then back into the CLEARinse CTS box for ground transportation of the specimen to a laboratory for testing.

The Wash Head is manufactured from inert, biocompatible plastics and is supplied as a sterile, single use device. The Transport Container is also manufactured from inert, biocompatible plastics and is a single use device but is not supplied sterile.

AI/ML Overview

Here's a breakdown of the acceptance criteria and the study details for the CLEARinse CTS Specimen Collection and Transport System, based on the provided FDA 510(k) summary:

1. Table of Acceptance Criteria and Reported Device Performance:

Acceptance Criteria Category	Specific Criteria	Reported Device Performance/Results
Limit of Detection (LoD) - PCR	Recovery of target analyte from transport media > 95% accuracy. Ct values within +/- 3 Ct of the mean for confirmatory tests.	Influenza A (H1N1): Preliminary: 1X LoD (4.0E-03 TCID50/mL) showed 100% positive (3 of 3 replicates). Confirmatory: 1X LoD (4.0E-03 TCID50/mL) showed 19 out of 20 results within acceptance range for FluA1. Influenza A (H3N2): Preliminary: 1X LoD (8.7E-02 TCID50/mL) showed 100% positive (3 of 3 replicates). Confirmatory: 1X LoD (2.6E-01 TCID50/mL) showed 20 out of 20 results within acceptance range for FluA1. Influenza B: Preliminary: 0.1X LoD (4.0E-03 TCID50/mL) showed 100% positive (3 of 3 replicates). Confirmatory: 1X LoD (4.0E-03 TCID50/mL) showed 20 out of 20 results within acceptance range for FluB. Established PCR LoDs: FluA (H1N1) 4.0 x10^-3 TCID50/mL, FluA (H3N2) 2.6 x10^-1 TCID50/mL, FluB 4.0 x10^-3 TCID50/mL.
Limit of Detection (LoD) - LFA	Lowest concentration producing >= 19 positives out of 20 samples (95%) in confirmatory testing.	Influenza A (H1N1): Confirmatory: 0.25X (2.63E+01 TCID50/mL) showed 100% positive (20 of 20). Influenza A (H3N2): Confirmatory: 40X (3.98E+03 TCID50/mL) showed 100% positive (20 of 20). Influenza B: Confirmatory: 20X (3.98E+02 TCID50/mL) showed 100% positive (20 of 20). Established LFA LoDs: FluA (H1N1) 2.6 x10^1 TCID50/mL, FluA (H3N2) 4.0 x10^3 TCID50/mL, FluB 4.0 x10^2 TCID50/mL.
Sample Stability - PCR	Deviation of no more than +/- 3 Ct for each target at a given time point from T=0.	Samples for viral nucleic acid testing are stable when stored for 48 hours at room (27°C) and refrigerated (4°C) temperatures. All average Ct values across time points (0-hr, 4-hr, 6-hr, 24-hr, 48-hr) for FluA H1N1, FluA H3N2, and FluB at both temperatures were within +/- 3 Ct of the baseline.
Sample Stability - LFA	Sufficient positive results maintained over time (implied by 3 of 3 positive replicates for 2.5X LoD samples).	Samples for antigen testing are stable when stored for 24 hours at room (27°C) temperature or 48 hours refrigerated (4°C). All replicates (3 of 3) were positive for FluA H1N1, FluA H3N2, and FluB at 2.5X LoD across tested time points (0-hr, 4-hr, 6-hr, 24-hr, 48-hr). Note: "24-hr" for room temp for LFA stability is implied as stability is reported as "24 hours at room temperature or 48 hours refrigerated."

2. Sample size used for the test set and the data provenance:

LoD Testing (PCR and LFA):
- Preliminary Test Set: Various concentrations were tested, typically with 3 replicates for each concentration.
- Confirmatory Test Set: 20 replicates were used for each influenza strain (H1N1, H3N2, FluB) at their respective 1X LoD (for PCR) or determined LoD (for LFA).
- Data Provenance: The data appears to be prospective as it involves controlled spiking of viral strains into a negative clinical matrix and subsequent testing with the device. The origin of the clinical nasal wash matrix (NWA) is not explicitly stated but is described as "pooled negative clinical nasal wash matrix." The tests were conducted using commercially available assays (Cepheid Xpert Xpress SARS-CoV-2/Flu/RSV and PBM BioSign Flu A+B).
Sample Stability Testing (PCR and LFA):
- Test Set: Each stability study evaluated triplicate samples for each influenza strain (H1N1, H3N2, FluB) per time point (0-hr, 4-hr, 6-hr, 24-hr, 48-hr) and per storage condition (4°C, 27°C). This means, for PCR, 3 strains * 5 time points * 2 temperatures * 3 replicates = 90 tests. Similarly for LFA, though the 0-hr was a baseline, and some time points appear a little different in the table headers.
- Data Provenance: Similar to LoD, this is prospective testing using spiked contrived samples in a negative clinical nasal wash matrix.

3. Number of experts used to establish the ground truth for the test set and the qualifications of those experts:

The study in this 510(k) summary focuses on analytical performance (Limit of Detection and Stability) rather than diagnostic accuracy against a clinical ground truth established by experts.
The "ground truth" for the test sets was derived from known concentrations of spiked viral strains (TCID50/mL) determined by established laboratory methods, and then assessed using commercially available and validated diagnostic assays (Cepheid Xpert Xpress SARS-CoV-2/Flu/RSV for PCR and PBM BioSign Flu A+B for LFA).
No human experts (e.g., radiologists) were used to establish the ground truth for these particular analytical studies.

4. Adjudication method (e.g., 2+1, 3+1, none) for the test set:

For the analytical studies described, no adjudication method (like 2+1 or 3+1) was used or required.
The results for LoD and stability were primarily quantitative (Ct values for PCR) or categorical (positive/negative for LFA) based on the output of the reference diagnostic assays. The "ground truth" for the spiked samples was the known concentration of the viral load.

5. If a multi reader multi case (MRMC) comparative effectiveness study was done, If so, what was the effect size of how much human readers improve with AI vs without AI assistance:

No Multi-Reader Multi-Case (MRMC) comparative effectiveness study was conducted or presented in this 510(k) summary.
The CLEARinse CTS is a specimen collection and transport system, not an AI-powered diagnostic imaging device or an AI-assisted detection tool where human reader performance would be a relevant metric. The studies focus on the ability of the device to preserve and recover viral nucleic acids and antigens for downstream testing.

6. If a standalone (i.e. algorithm only without human-in-the-loop performance) was done:

The performance described in this 510(k) is essentially standalone performance of the collection and transport system.
The device itself (CLEARinse CTS) is not an "algorithm" in the typical sense of AI/ML. Its performance is evaluated by its ability to maintain the integrity of spiked samples, which are then analyzed by already cleared in vitro diagnostic assays (the Cepheid PCR and PBM BioSign LFA). These assays operate independently to detect the target. The studies demonstrate the device's contribution to maintaining sample quality for these standalone diagnostic tests.

7. The type of ground truth used (expert consensus, pathology, outcomes data, etc.):

The ground truth for both the LoD and stability studies was based on contrived samples with known concentrations of viral pathogens. These concentrations were quantified using standard laboratory methods (TCID50/mL).
The "true" positivity or negativity of a sample at a given concentration was determined by the performance of the established reference nucleic acid (PCR) and antigen (LFA) diagnostic assays, and compared against their expected LoDs.

8. The sample size for the training set:

As this submission pertains to an in vitro diagnostic specimen collection and transport device and not an AI/ML algorithm that requires training, there is no "training set" in the context of machine learning.
The performance data provided is for verification and validation of the device's analytical capabilities.

9. How the ground truth for the training set was established:

Again, since this is not an AI/ML device, the concept of a "training set" and establishing its ground truth for machine learning does not apply.
The ground truth for the test samples used in the analytical studies was established by spiking known concentrations of influenza strains into a negative clinical matrix (as described in point 7).

Summary

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

July 3, 2023

Aardvark Medical, Inc. Steven Bacich CEO 204 Cardinal Drive, Suite 300 Denton, TX 94960 United States

Re: K221664

Trade/Device Name: CLEARinse CTS Specimen Collection and Transport System Regulation Number: 21 CFR 866.2950 Regulation Name: Microbial Nucleic Acid Storage And Stabilization Device Regulatory Class: Class II Product Code: OBD Dated: June 7, 2022 Received: June 8, 2022

Dear Steven Bacich:

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 (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 located 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.

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

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

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

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

Sincerely,

Noel J. Gerald -S

Noel J. Gerald, Ph.D. Branch Chief Bacterial Respiratory and Medical Countermeasures Branch Division of Microbiology Devices OHT7: Office of In Vitro Diagnostics Office of Product Evaluation and Ouality Center for Devices and Radiological Health

Enclosure

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Indications for Use

510(k) Number (if known) K221664

Device Name CLEARinse CTS

Indications for Use (Describe)

Type of Use (Select one or both, as applicable)

☑ Prescription Use (Part 21 CFR 801 Subpart D)
☐ Over-The-Counter Use (21 CFR 801 Subpart C)

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510(K) SUMMARY

Submitter's Name and Address:

Aardvark Medical Company. 204 Cardinal Drive, Suite 300 Denton. TX 94960 United States

Contact Name and Information:

Steven Bacich 650-218-5934 stevenbacich(@gmail.com

Date prepared:

June 7, 2023

Device Information:

Trade/Proprietary Name:

System Common or Usual Name: Classification Name: Classification Number: OBD Product Code:

CLEARinse™ CTS Specimen Collection and Transport Microbial Nucleic Acid Storage and Stabilization Device Microbial Nucleic Acid Storage and Stabilization Device Class II, 21 CFR 866.2950

Predicate Device:

510(k)Number	Device	Product Code
DEN170029	PrimeStore® MTM	QBD

Device Description:

CLEARinse CTS Specimen Collection and Transport System

The product is an extension of the CLEARinse Pro device, a 510(k)-cleared (K082762) professional use medical device with indications for nasal lavage as well as mucus sample collection for subsequent testing. The CLEARinse Pro device has two main components: a pump-containing Handle and a disposable collection Wash Head. The CLEARinse Pro Handle and Instructions for Use are required for use of the CLEARinse CTS.

The CLEARinse CTS is comprised of two component assemblies: a revision to the 510(k)cleared Wash Head and a Transport Container to protect the Wash Head and specimen during

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transit. The CLEARinse CTS. used with the CLEARinse Pro Handle, will allow a nasal wash aspirate specimen to be collected by a medical professional, protected for transit, and ground shipped to a testing site. The user mounts the CLEARinse CTS Wash Head onto the CLEARinse Pro Handle and adds sterile saline as directed. The tip of the Wash Head is inserted into the patient's nostril and sterile saline is introduced. The irrigated saline and nasal secretions are then aspirated back into the Wash Head, collecting the specimen to be tested. The CLEARinse CTS Wash Head is removed from the Handle and placed into the Transport Container assembly (Container Cup, Filter Seal, Silicone Seal, and Container Lid) and then back into the CLEARinse CTS box for ground transportation of the specimen to a laboratory for testing.

Indications for Use/Intended Use:

Device & PredicateDevice(s):	Subject: K221664	Predicate: DEN170029
Device Trade Name	CLEARinse CTS:Specimen Collectionand Transport System	PrimeStore MTM
General DeviceCharacteristic Similarities
Intended Use/IndicationsFor Use	CLEARinse CTS isintended to collect andtransport clinical nasalwash aspiratespecimens that maycontain influenza A orinfluenza B virus, byhealth careprofessionals, from thecollection site to alaboratory or testingsite. CLEARinse CTS	PrimeStore MTM isintended for thestabilization,transportation andinactivation ofinfectious unprocessednasal washes suspectedof containing Influenzaa virus RNA.PrimeStore MTM isalso intended for thestabilization,

Comparison of Technological Characteristics:

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	specimens are suitablefor use with legallymarketed lateral flowimmunoassays andmolecular assays.CLEARinse CTS is forprofessional in-vitrodiagnostic use.	transportation andinactivation ofinfectious unprocessedsputum samplessuspected of containingMycobacteriumtuberculosis DNA fromhuman samples.
Material	Polypropylene	Same
General DeviceCharacteristic Differences
Specimen Type	Nasal Wash Aspiratesuspected of containinginfluenza A or B viruses	Nasal wash suspectedof containing InfluenzaA virus. Sputumsamples suspected ofcontaining MTB.
Transport MediaFormulation	0.9% saline from nasalwash	Nucleic AcidStabilization solution
Device Material Supplied	Wash Head andTransport Container	Molecular TransportMedium provided incryogenic tube
Sterility	Sterile	Not sterile
Inactivation	No inactivation ofviruses or bacteria	Inactivation ofinfluenza A and MTBby Guanidinethiocyanate
Specimen Collection	Collection using theCLEARinse Pro Handleand Instructions for Use	Collection usingstandard clinicalprocedures

The Aardvark Medical Company CLEARinse CTS Device is substantially equivalent to the legally marketed predicate device. The CLEARinse CTS Device has the same intended use, similar indications for use, and similar technological characteristics for transporting nasal wash specimens to the laboratory as the predicate device. Performance data demonstrates that the CLEARinse CTS Device is as safe and effective.

Performance Data:

Detection Limit

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Limit of Detection (LoD) testing was conducted to determine the lowest concentration of influenza A and influenza B that contains measurable nucleic acids or measurable antigens. Recovery of target analyte from the transport media, with a greater than 95% accuracy, was conducted by spiking pooled negative clinical nasal wash matrix (NWA and 0.9% saline solution) with various concentrations of influenza nucleic acid or antigen. The contrived samples were then placed into and removed from the Wash Head. The recovered samples are then placed into the workflow for the Cepheid® Xpert® Xpress SARS-CoV-2/Flu/RSV and Princeton BioMeditech Corporation (PBM) BioSign® Flu A+B assays. The results are compared to established LoD for the reference assays, found in their respective Package Inserts (PI).

A. PCR LoD Test Results:

Preliminary LoD testing was initially performed by spiking multiple concentrations of three different strains of influenza into NWA. The three viral concentrations used were 3X, 1X, and 0.5X of the established LoDs of similar influenza strains (Table 1) in the PI for the reference assay. The assay used was the EUA authorized, real-time reverse transcription polymerase chain reaction (PCR) Cepheid Xpert Xpress SARS-CoV-2/Flu/RSV Cartridge using the GeneXpert® Xpress System. Positive and negative results were determined as explained in the PI for the device.

Virus Type	Strain tested with CTS device	Reported LoD of Similar Strain*
Influenza A, H1N1	A/Netherlands/602/2009	$4.0 x 10^{-3}$ TCID50/mL for A/California/7/2009(H1N1)
Influenza A, H3N2	A/Indiana/10/2011	$8.7 x 10^{-2}$ TCID50/mL for A/Victoria/361/2011(H3N2)
Influenza B	B/Florida/07/2004	$4.0 x 10^{-2}$ TCID50/mL for B/Massachusetts/2/2012

Table 1. Proposed test levels for Cepheid PCR testing for preliminary LoD study

*Note: strains used to determine analytical sensitivity (LoDs) for validation of the Cepheid Xpert Xpress SARS-CoV-2/Flu/RSV Cartridge

For the Cepheid Xpert Xpress SARS-CoV-2/Flu/RSV PCR test, LoD range-finding results indicated that the influenza A (FluA) samples contrived by the sponsor performed similarly to those used by Cepheid to establish the reported LoDs. For both FluA viruses, all three replicates were positive at 1X and 3X the reported LoD (Table 2). At 0.5X the reported LoD, each virus was negative for one or more replicates, indicating that the 1X level was truly near the LoD using the viral stocks listed in Table 1. For the influenza B (FluB), testing at 10-fold lower levels (0.05X - 0.3X) resulted in similar qualitative results: all three replicates positive at 0.1X and 0.3X, and only two of three replicates positive at 0.05X (Table 2).

Table 2. Preliminary PCR LoD estimation results with individual LoDs highlighted in grey.

Flu Type	Test Level	Qualitative Results	Mean Ct*
----------	------------	---------------------	----------

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	Description	TCID50/mL	# Positive	% Positive
FluA H1N1	3X	1.2E-02	3 of 3	100%	34.9
	1X	4.0E-03	3 of 3	100%	36.8
	0.5X	2.0E-03	2 of 3	67%	37.4
FluA H3N2	3X	2.6E-01	3 of 3	100%	35.6
	1X	8.7E-02	3 of 3**	100%	36.9
	0.5X	4.3E-02	0 of 3	0%	N/A
FluB	0.3X	1.2E-02	3 of 3	100%	36.0
	0.1X	4.0E-03	3 of 3	100%	38.8
	0.05X	2.0E-03	2 of 3	67%	38.1

*Note: Mean Ct for FluA viruses are calculated based on the FluA1 analyte, which produced consistent Ct values.

**One replicate was positive on FluA1 and was negative on FluA2 and was considered below the LoD.

Confirmatory LoD testing was conducted with 20 replicates using contrived samples. The samples for the confirmatory testing were contrived using the sample procedure that was used for the preliminary study. The Ct values for the 20 replicates of the confirmatory tests for each of the three influenza strains are shown in Tables 3-5. Influenza A H1N1 had one Ct value out of acceptance range for analyte FluA1 resulting in 19 out of 20 results within the acceptance range of +/- 3 Ct of the mean. Both influenza A H3N2 and B had 20 out of 20 Ct values for either analyte FluA1 or FluB, respectively, within the acceptance range.

Table 3. PCR LoD confirmation results for influenza A H1N1 with out of acceptance range FluA1 Ct values highlighted in grey.

Concentration(1X LoD)	Replicate	FluA1Result	FluA1 Ct	FluA2 Ct
4.00E-03	1	Positive	35.8	36.7
4.00E-03	2	Positive	35.2	36.4
4.00E-03	3	Positive	35.3	37
4.00E-03	4	Positive	38	41.1
4.00E-03	5	Positive	35.1	36.5
4.00E-03	6	Positive	36	37.8
4.00E-03	7	Positive	35.7	38.2
4.00E-03	8	Positive	35.9	40
4.00E-03	9	Positive	36.5	37.8
4.00E-03	10	Positive	35.6	37.2
4.00E-03	11	Positive	36.9	38.6

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4.00E-03	12	Positive	35.4	36.8
4.00E-03	13	Positive	32.2	34.3
4.00E-03	14	Positive	35.2	36.1
4.00E-03	15	Positive	34.4	35.3
4.00E-03	16	Positive	36	38.6
4.00E-03	17	Positive	34.9	36.7
4.00E-03	18	Positive	36.3	37.8
4.00E-03	19	Positive	35.3	37.2
4.00E-03	20	Positive	35.5	37.3

Table 4. PCR LoD confirmation results for influenza A H3N2 with out of acceptance range
FluA1 Ct values highlighted in grey.

Concentration(1X LoD)	Replicate	FluA1Result	FluA1 Ct	FluA2 Ct
2.60E-01	1	Positive	37.1	ND
2.60E-01	2	Positive	36.1	40
2.60E-01	3	Positive	37.7	39
2.60E-01	4	Positive	35.1	37
2.60E-01	5	Positive	35.4	43
2.60E-01	6	Positive	34.3	37
2.60E-01	7	Positive	35.3	ND
2.60E-01	8	Positive	35.5	38
2.60E-01	9	Positive	35.3	39
2.60E-01	10	Positive	35	37
2.60E-01	11	Positive	35.2	39
2.60E-01	12	Positive	35.8	38
2.60E-01	13	NA*	NA*	NA*
2.60E-01	14	Positive	35.2	38
2.60E-01	15	Positive	35.6	38
2.60E-01	16	Positive	36	39
2.60E-01	17	Positive	35.3	39
2.60E-01	18	Positive	35.6	39
2.60E-01	19	Positive	35.5	38
2.60E-01	20	Positive	34.9	37
2.60E-01	21	Positive	35.5	41

*Note: Error. Probe check failed with the output of "NO RESULT – REPEAT TEST."

Table 5. PCR LoD confirmation results for influenza B with out of acceptance range FluB Ct values highlighted in grey.

Concentration(1X LoD)	Replicate	FluBResult	FluB Ct
4.00E-03	1	Positive	36.1

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4.00E-03	2	Positive	36.1
4.00E-03	3	Positive	36.1
4.00E-03	4	Positive	36.3
4.00E-03	5	Positive	36.5
4.00E-03	6	Positive	36.7
4.00E-03	7	Positive	36.3
4.00E-03	8	Positive	36.2
4.00E-03	9	Positive	36.9
4.00E-03	10	Positive	36.2
4.00E-03	11	Positive	35.8
4.00E-03	12	Positive	35.9
4.00E-03	13	Positive	35.7
4.00E-03	14	NA*	NA*
4.00E-03	15	Positive	36.3
4.00E-03	16	Positive	36.6
4.00E-03	17	Positive	39.2
4.00E-03	18	Positive	36.4
4.00E-03	19	Positive	35.9
4.00E-03	20	Positive	37.1
4.00E-03	21	Positive	36.4

*Note: Error. Probe check failed with the output of "NO RESULT – REPEAT TEST."

In summary, the testing performed established that for PCR assays 4.0 x10-3 TCIDsolmL is the LoD for both influenza A(H1N1) and B, and that 2.6 x10'' TCID50/mL is the LoD for influenza A(H3N2).

B. LFA LoD Test Results:

Preliminary LoD testing was initially performed by spiking multiple concentrations of three different strains of influenza into NWA. The three viral concentrations used were 3X, 1X, and 0.5X of the established LoDs of similar influenza strains (Table 6) in the PI for the reference assay. The preliminary results indicated that additional antigen concentrations needed to be tested beyond the initially planned levels. The assay used was the PBM BioSign® Flu A+B lateral flow assay (LFA) (K182157). Positive and negative results were determined as explained in the PI for the device.

Virus Type	Strain tested with CTSdevice	Reported LoD of SimilarStrain*
Influenza A,H1N1	A/Netherlands/602/2009	1.1 x 102 TCID50/mL forA/PR/8/34(H1N1)
Influenza A,H3N2	A/Indiana/10/2011	10 x 101 TCID50/mL forA/Victoria/3/75(H3N2)

Table 6. Proposed test levels for PBM BioSign LFA testing for preliminary LoD study

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Influenza B	B/Florida/07/2004	$2.0 x 101 TCID50/mL for$B/Maryland/1/59
-------------	-------------------	----------------------------------------------

*Note: strains used to determine analytical sensitivity (LoDs) for validation of the PBM BioSign Flu A+B LFA

For influenza A H1N1, all replicates produced positive results at all test levels, so additional lower levels (0.25X and 0.125X) were tested. For influenza A H3N2 and influenza B, all levels initially tested produced negative results for all three replicates. Thus, higher levels were tested until a test level produced three of three positive results. A summary of levels tested and the results at each level are presented in Table 7.

	Test Level		Qualitative Results
Flu Type	Description	TCID50/mL	# Positive	% Positive
FluA H1N1	0.5X	5.25E+01	3 of 3	100%
FluA H1N1	0.25X	2.63E+01	3 of 3	100%
FluA H1N1	0.125X	1.32E+01	0 of 3	0%
FluA H3N2	27X*	2.65E+03	3 of 3	100%
FluA H3N2	20X	1.99E+03	0 of 3	0%
FluA H3N2	10X	9.95E+02	0 of 3	0%
FluB	20X	3.98E+02	3 of 3	100%
FluB	10X	1.99E+02	0 of 3	0%
FluB	3X	5.97E+01	0 of 3	67%

Table 7. Preliminary LFA LoD estimation results with individual LoDs highlighted in grey.

*Note: LoD is lower than for confirmatory studies below (Table 8) because of an accidental miscalculation in preparation of H3N2 analyte for the range-finding study.

Confirmatory LoD testing was conducted with 20 replicates using contrived samples. The samples for the confirmatory testing were contrived using the sample procedure that was used for the preliminary study. For all three viral test strains the lowest concentration that produced at least 19 positives out of 20 samples (95%) was determined to be the LoD for each test strain (Table 8).

Flu Type	Test Level		Qualitative Results
	Description	TCID50/mL	# Positive	% Positive
FluA H1N1	0.25X	2.63E+01	20 of 20	100%
FluA H3N2	40X*	3.98E+03	20 of 20	100%
FluB	20X	3.98E+02	20 of 20	100%

Table 8. LFA LoD confirmation results

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*Note: Confirmatory testing for FluA strain H3N2 was performed at a higher LoD than the range-finding LoD study.

In summary, the testing performed established that 2.6 x101 TCIDs0/mL is the LoD for influenza A(H3N2), .4.0 x103 TCID50/mL is the LoD for influenza A(H1N1) and 4.0 x102 TCID50/mL is the LoD for influenza B.

Sample Stability

A. Viral Nucleic Acid Stability

Contrived samples containing influenza strains at a concentration of 2.5X LoD were prepared in pooled NWA matrix. Sample stability was evaluated in a study in which each device contained 2 mL of virus inoculum in NWA. The spiked NWA was placed in a CLEARinse CTS Wash Head, sealed in a CLEARinse CTS Transport Container and incubated at 4ºC and 27°C. Testing of the contrived samples was conducted at time point zero to establish a baseline and again at 4 hours, 6 hours, 24 hours, and 48 hours. An aliquot of each contrived sample was tested for influenza using a Cepheid Xpert Xpress SARS-CoV-2/Flu/RSV Cartridge. Each PCR assay was performed in triplicate for each time point and for each storage condition. Samples were not batched, and all testing was completed immediately upon removal of sample from the Wash Head following the instructions in the PI. The acceptance criteria for the stability study included a deviation of no more than +/- 3 Ct for each target at given time point from T = 0

Results of stability testing for each of the three influenza strains are shown below in Table 9.

Virus and LoD	StorageTemp	Average Ct value
		0-hr	4-hr	6-hr	24-hr	48-hr
FluA H1N1	4°C	NT	35.6	36	36	36.7
2.5X LoD	27°C	34.6	35.8	34.8	34.9	36.9
FluA H3N2	4°C	NT	35.1	35.6	35.4	36
2.5X LoD	27°C	35.4	35.6	35.8	36.1	36.4
FluB	4°C	NT	36.7	36.1	37.8	37.0
5X LoD	27°C	36.3	36.4	37.6	37.2	38.5

Table 9. Specimen stability study quantitative PCR results from CTS devices

In summary, these data support the claim that samples that will be used for viral nucleic acid testing are stable when stored for 48 hours at room and refrigerated temperature.

B. Viral Antigen Stability

Contrived samples containing Influenza strains at a concentration of 2.5X LoD were prepared in pooled NWA matrix. Sample stability was evaluated in a study in which each

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device contained 2 mL of virus inoculum in NWA. The spiked NWA was placed in a CLEARinse CTS Wash Head, sealed in a CLEARinse CTS Transport Container and incubated at 4°C and 27°C. Testing of the contrived samples was conducted at time point zero to establish a baseline and again at 4 hours, 6 hours, and 48 hours. An aliquot of each contrived sample was tested for influenza using a PBM BioSign Flu A + B test kit. Each LFA assay was performed in triplicate for each time point and for each storage condition. Samples were not batched, and all testing was completed immediately upon removal of sample from the Wash Head following the instructions in the PI.

Results of stability testing for each of the three influenza strains are shown below in Table 10.

Virus	StorageTemp	Number of positive samples out of replicates with valid results
		0-hr	4-hr	6-hr	24-hr	48-hr
FluA	4°C	NT	3 of 3	3 of 3	3 of 3	3 of 3
H1N1	27°C	3 of 3	3 of 3	3 of 3	3 of 3	3 of 3
FluA	4°C	NT	3 of 3	3 of 3	3 of 3	3 of 3
H3N2	27°C	3 of 3	3 of 3	3 of 3	3 of 3	3 of 3
FluB	4°C	NT	3 of 3	3 of 3	3 of 3	3 of 3
	27°C	3 of 3	3 of 3	3 of 3	3 of 3	3 of 3

Table 10. LFA Results from CTS Devices at 2.5X LoD

In summary, these data support the claim that samples that will be used for antigen testing are stable when stored for 24 hours at room temperature or 48 hours refrigerated.

Conclusion:

The CLEARinse CTS device does not raise different questions regarding safety and effectiveness as compared to predicate device. The proposed device is as safe, as effective, and performs as well as or better than the predicate device. The information submitted in this premarket notification supports a substantial equivalence decision.

Regulation Number and Section

§ 866.2950 Microbial nucleic acid storage and stabilization device.

(a)
Identification. A microbial nucleic acid storage and stabilization device is a device that consists of a container and reagents intended to stabilize microbial nucleic acids in human specimens for subsequent isolation and purification of nucleic acids for further molecular testing. The device is not intended for preserving morphology or viability of microorganisms.(b)
Classification. Class II (special controls). The special controls for this device are:(1) The intended use for the labeling required under § 809.10 of this chapter must include a detailed description of microorganisms and types of human specimens intended to be preserved.
(2) The labeling required under § 809.10(b) of this chapter must include the following:
(i) A detailed device description, including all device components;
(ii) Performance characteristics from applicable analytical studies, including nucleic acid stability and microorganism inactivation;
(iii) A limiting statement that erroneous results may occur when the transport device is not compatible with molecular testing; and
(iv) A limiting statement that the device has only been validated to preserve the representative microorganisms used in the analytical studies.
(3) Design verification and validation must include the following:
(i) Overall device design, including all device components and all control elements incorporated into the analytical validation procedures;
(ii) Thorough description of the microorganisms and methodology used in the validation of the device including, extraction platforms and assays used for the detection of preserved nucleic acids; and
(iii) The limit of detection (LoD) of the molecular test used to establish microorganism nucleic acid stability.