Acucy® Influenza A&B Test with the Acucy® 2 System

K241188 · SEKISUI Diagnostics, LLC · PSZ · Apr 18, 2025 · Microbiology

Device Facts

• Record ID: K241188
• Device Name: Acucy® Influenza A&B Test with the Acucy® 2 System
• Applicant: SEKISUI Diagnostics, LLC
• Product Code: PSZ · Microbiology
• Decision Date: Apr 18, 2025
• Decision: SESE
• Submission Type: Dual Track
• Regulation: 21 CFR 866.3328
• Device Class: Class 2

Intended Use

The Acucy Influenza A&B Test is a rapid chromatographic immunoassay for the qualitative detection and differentiation of influenza A and B viral nucleoprotein antigens directly from anterior nasal and nasopharyngeal swabs from patients with signs and symptoms of respiratory infection. The test is intended for use with the Acucy or Acucy 2 Reader as an aid in the diagnosis of influenza A and B viral infections. The test is not intended for the detection of influenza C viruses. Negative test results are presumptive and should be confirmed by viral culture or an FDA-cleared influenza A and B molecular assay. Negative test results do not preclude influenza viral infection and should not be used as the sole basis for treatment or other patient management decisions. Performance characteristics for influenza A were established during the 2017-2018 influenza season when influenza A/H3N2 and A/H1N1pdm09 were the predominant influenza A viruses in circulation. When other influenza A viruses are emerging, performance characteristics may vary. 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 department for testing. Viral culture should not be attempted in these cases unless a BSL 3+ facility is available to receive and culture specimens.

Device Story

Device is a rapid chromatographic immunoassay for influenza A and B detection. Input: anterior nasal or nasopharyngeal swab samples. Process: sample extraction in buffer to expose viral nucleoproteins; migration on test cassette membrane; binding to colloidal gold-conjugated monoclonal antibodies; capture by membrane-bound antibodies. Reader: Acucy 2 Reader scans cassette to measure absorbance intensity; processes results via method-specific algorithms; displays POS/NEG/INVALID results. Used in clinical settings by healthcare professionals. Two modes: 'WALK AWAY' (automatically timed) or 'READ NOW' (manually timed). Output aids diagnosis; negative results require confirmatory testing. Benefits: rapid, automated, objective interpretation of lateral flow results.

Clinical Evidence

No new clinical studies performed; clinical performance leveraged from K182001. Analytical performance validated via bench testing: precision/reproducibility (n=90 replicates per panel member), LoD (confirmed for 4 strains), carry-over, and method comparison (n=60 samples) against predicate, showing 100% PPA for Flu A/B and 98.3% NPA for Flu A.

Technological Characteristics

Lateral flow immunochromatographic assay; sandwich format. Materials: nitrocellulose membrane, colloidal gold-conjugated mouse monoclonal antibodies. Reader: optoelectronic reflectance-based measurement using CMOS camera. Connectivity: USB for data transfer. Sterilization: Ethylene oxide (ISO 11135, ISO 10993-7). Software: algorithm-based qualitative result determination.

Indications for Use

Indicated for qualitative detection and differentiation of influenza A and B viral nucleoprotein antigens in symptomatic patients using anterior nasal or nasopharyngeal swabs. Not for influenza C detection. Negative results require confirmation by culture or molecular assay.

Regulatory Classification

Identification

An influenza virus antigen detection test system is a device intended for the qualitative detection of influenza viral antigens directly from clinical specimens in patients with signs and symptoms of respiratory infection. The test aids in the diagnosis of influenza infection and provides epidemiological information on influenza. Due to the propensity of the virus to mutate, new strains emerge over time which may potentially affect the performance of these devices. Because influenza is highly contagious and may lead to an acute respiratory tract infection causing severe illness and even death, the accuracy of these devices has serious public health implications.

Special Controls

*Classification.* Class II (special controls). The special controls for this device are:(1) The device's sensitivity and specificity performance characteristics or positive percent agreement and negative percent agreement, for each specimen type claimed in the intended use of the device, must meet one of the following two minimum clinical performance criteria: (i) For devices evaluated as compared to an FDA-cleared nucleic acid based-test or other currently appropriate and FDA accepted comparator method other than correctly performed viral culture method: (A) The positive percent agreement estimate for the device when testing for influenza A and influenza B must be at the point estimate of at least 80 percent with a lower bound of the 95 percent confidence interval that is greater than or equal to 70 percent. (B) The negative percent agreement estimate for the device when testing for influenza A and influenza B must be at the point estimate of at least 95 percent with a lower bound of the 95 percent confidence interval that is greater than or equal to 90 percent. (ii) For devices evaluated as compared to correctly performed viral culture method as the comparator method: (A) The sensitivity estimate for the device when testing for influenza A must be at the point estimate of at least 90 percent with a lower bound of the 95 percent confidence interval that is greater than or equal to 80 percent. The sensitivity estimate for the device when testing for influenza B must be at the point estimate of at least 80 percent with a lower bound of the 95 percent confidence interval that is greater than or equal to 70 percent. (B) The specificity estimate for the device when testing for influenza A and influenza B must be at the point estimate of at least 95 percent with a lower bound of the 95 percent confidence interval that is greater than or equal to 90 percent. (2) When performing testing to demonstrate the device meets the requirements in paragraph (b)(1) of this section, a currently appropriate and FDA accepted comparator method must be used to establish assay performance in clinical studies. (3) Annual analytical reactivity testing of the device must be performed with contemporary influenza strains. This annual analytical reactivity testing must meet the following criteria: (i) The appropriate strains to be tested will be identified by FDA in consultation with the Centers for Disease Control and Prevention (CDC) and sourced from CDC or an FDA-designated source. If the annual strains are not available from CDC, FDA will identify an alternative source for obtaining the requisite strains. (ii) The testing must be conducted according to a standardized protocol considered and determined by FDA to be acceptable and appropriate. (iii) By July 31 of each calendar year, the results of the last 3 years of annual analytical reactivity testing must be included as part of the device's labeling. If a device has not been on the market long enough for 3 years of annual analytical reactivity testing to have been conducted since the device received marketing authorization from FDA, then the results of every annual analytical reactivity testing since the device received marketing authorization from FDA must be included. The results must be presented as part of the device's labeling in a tabular format, which includes the detailed information for each virus tested as described in the certificate of authentication, either by: (A) Placing the results directly in the device's § 809.10(b) of this chapter compliant labeling that physically accompanies the device in a separate section of the labeling where the analytical reactivity testing data can be found; or (B) In the device's label or in other labeling that physically accompanies the device, prominently providing a hyperlink to the manufacturer's public Web site where the analytical reactivity testing data can be found. The manufacturer's home page, as well as the primary part of the manufacturer's Web site that discusses the device, must provide a prominently placed hyperlink to the Web page containing this information and must allow unrestricted viewing access. (4) If one of the actions listed at section 564(b)(1)(A)-(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 (HHS) 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 viral samples in accordance with a standardized protocol considered and determined by FDA to be acceptable and appropriate. The procedure and location of testing may depend on the nature of the emerging virus. (ii) Within 60 days from the date that FDA notifies manufacturers that characterized 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 § 809.10(b) of this chapter compliant labeling that physically 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 physically accompanies the device, prominently providing a hyperlink to the manufacturer's public Web site where the analytical reactivity testing data can be found. The manufacturer's home page, as well as the primary part of the manufacturer's Web site that discusses the device, must provide a prominently placed hyperlink to the Web page containing this information and must allow unrestricted viewing access.

Predicate Devices

• Acucy Influenza A&B Test with the Acucy System (K182001)

Submission Summary (Full Text)

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FDA

U.S. FOOD & DRUG

ADMINISTRATION

# 510(k) SUBSTANTIAL EQUIVALENCE DETERMINATION DECISION SUMMARY

ASSAY AND INSTRUMENT

# I. Background Information:

A 510(k) Number

K241188

B Applicant

Sekisui Diagnostics, LLC

C Proprietary and Established Names

Acucy Influenza A&B Test with the Acucy 2 System

D Regulatory Information

| Product Code(s) | Classification | Regulation Section | Panel |
| --- | --- | --- | --- |
| PSZ | Class II | 21 CFR 866.3328 - Influenza Virus Antigen Detection Test System | MI - Microbiology |

# II. Submission/Device Overview:

A Purpose for Submission:

To obtain 510(k) clearance of the Acucy Influenza A&B Test with the Acucy 2 System

B Measurand:

Influenza A and influenza B viral nucleoprotein antigens

C Type of Test:

Qualitative immunoassay using an optoelectronic reader

Food and Drug Administration

10903 New Hampshire Avenue

Silver Spring, MD 20993-0002

www.fda.gov

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III. Intended Use/Indications for Use:

A Intended Use(s):
See Indications for Use below.

B Indication(s) for Use:
The Acucy Influenza A&B Test is a rapid chromatographic immunoassay for the qualitative detection and differentiation of influenza A and B viral nucleoprotein antigens directly from anterior nasal and nasopharyngeal swabs from patients with signs and symptoms of respiratory infection. The test is intended for use with the Acucy or Acucy 2 Reader as an aid in the diagnosis of influenza A and B viral infections. The test is not intended for the detection of influenza C viruses. Negative test results are presumptive and should be confirmed by viral culture or an FDA-cleared influenza A and B molecular assay. Negative test results do not preclude influenza viral infection and should not be used as the sole basis for treatment or other patient management decisions.

Performance characteristics for influenza A were established during the 2017-2018 influenza season when influenza A/H3N2 and A/H1N1pdm09 were the predominant influenza A viruses in circulation. When other influenza A viruses are emerging, performance characteristics may vary.

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 department for testing. Viral culture should not be attempted in these cases unless a BSL 3+ facility is available to receive and culture specimens.

C Special Conditions for Use Statement(s):
Rx - For Prescription Use Only

D Special Instrument Requirements:
Acucy 2 Reader

IV. Device/System Characteristics:

A Device Description:
The Acucy Influenza A&B Test is a lateral flow immunochromatographic assay in the sandwich immunoassay format for the direct and qualitative detection and differentiation of influenza A and B viral nucleoprotein antigens from anterior nasal and nasopharyngeal swabs of symptomatic patients. The test is automatically analyzed on the Acucy 2 Reader and should not be visually read. The Acucy Influenza A&B Test components/reagents have not been modified from those originally cleared in K182001.

B Principle of Operation:
The Acucy Influenza A&B Test allows for the differential detection of influenza A and influenza B antigens, when used with the Acucy 2 Reader. The patient swab sample is placed in the Extraction Buffer vial, during which time the virus particles in the sample are disrupted,

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exposing internal viral nucleoproteins. After disruption, the sample is dispensed into the Test Cassette's sample well where it solubilizes reagents necessary for the detection of the viral antigens. From the sample well, the sample migrates along the membrane surface. If influenza A or B viral antigens are present, they will form a complex with the respective solubilized colloidal gold conjugated mouse monoclonal antibody to influenza A and/or B nucleoproteins. The complex will then continue to migrate until it is bound by rat anti-influenza A and/or mouse anti-influenza B antibodies that are immobilized at the test lines of the cassette's nitrocellulose membrane.

Depending upon the operator's choice, the Test Cassette is either immediately placed inside the Acucy 2 Reader for automatically timed development mode (WALK AWAY Mode) or placed on the counter or bench top for a manually timed development and then placed into the Acucy 2 Reader to be scanned (READ NOW mode). In either mode, upon completion of the development time, the Acucy 2 Reader scans the Test Cassette to measure the absorbance intensity at the test and control lines. These absorbance units are then processed to generate a final qualitative result using a method-specific algorithms.

The Acucy 2 Reader then displays the test results on the screen as either POS (+) or NEG (-) for each viral analyte, or as INVALID. The results can also be automatically printed on the optional printer.

There are five possible results:

- FLU A POS (+)/FLU B NEG (-)
- FLU A NEG (-)/FLU B POS (+)
- FLU A NEG (-)/FLU B NEG (-)
- FLU A POS (+)/FLU B POS (+)
- INVALID

Any INVALID result should be retested with a new patient sample and a new Test Kit.

FLU A and FLU B single positive results, and FLU A and B dual positive results do not rule out co-infection with other pathogens, nor do they identify any specific Influenza A or B virus subtypes.

Co-infection with Influenza A and B (i.e., FLU A and B dual positive results) is rare and it is therefore recommended that a FLU A and FLU B dual positive sample be re-tested with a new sample and a new test kit. Repeatable Influenza A and B dual positive results should be confirmed by cell culture or PCR testing before reporting results.

A FLU A and B negative result does not exclude influenza viral infection. Negative results should be confirmed by viral culture or an FDA-cleared Influenza A and Influenza B molecular assay.

## Materials Provided

- 25 Test Cassettes: individually foil pouched with desiccant
- 25 Sterile anterior nasal swabs
- 25 Extraction Buffer vials each containing 0.4 mL phosphate buffered salt solution with 0.09% sodium azide as a preservative
- 25 Extraction Vial Dropper Tips
- 1 Influenza A+/B- Control Swab (Formalin inactivated Influenza A containing 0.05% sodium azide. Inactivity confirmed by inability of virus to infect cell culture)

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- 1 Influenza A-/B+ Control Swab (Formalin inactivated Influenza A containing 0.05% sodium azide. Inactivity confirmed by inability of virus to infect cell culture)
- 2 separate Instructions for Use (IFU) documents for Acucy 2 Reader and Acucy Reader
- 2 separate Quick Reference Guides for Acucy 2 Reader and Acucy Reader (each containing instructions for READ NOW and WALK AWAY Modes)
- 2 separate External Quality Control (QC) Quick Reference Guides for the Acucy and the Acucy 2 Readers
- Workstation

## Materials Required but Not Provided

- Acucy 2 Reader
- Timer or watch
- If needed, sterile nasopharyngeal swabs (Copan Catalog # 534CS01)
- If needed, additional external quality controls may be purchased separately (Acucy Influenza A&B Control Kit # 1011)

## C Instrument Description Information:

### 1. Acucy 2 Reader Description:

The Acucy 2 Reader is an optoelectronic reader that detects the insertion of an Acucy test cassette and reads the cassette barcode to detect the test information, prior to the drawer closing. Once it recognizes the test type, the user is prompted to close the drawer manually and the Acucy 2 Reader illuminates the cassette in the drawer and measures the intensity of the control and test lines.

The Acucy 2 Reader uses a reflectance-based measurement method to evaluate the line signal intensities on the test strip, and a specific algorithm to determine the presence or absence of any target analyte(s). The Acucy 2 Reader determines the line intensity at each of the spatially-defined test and control line positions, interprets the results using a scoring algorithm, and reports a positive, negative, or invalid result based on pre-set thresholds. A liquid crystal display (LCD) on the Acucy 2 Reader communicates the testing progress and the final sample's test results to the operator.

The Acucy 2 Reader supports the use of different assays by reading an assay-specific 2D barcode (2D) on the Test Cassette that identifies the test type (currently only Influenza A&B).

The Reader has the ability to store and transfer test results via a USB memory drive.

### 2. Acucy 2 Reader Modes:

Similar to the Acucy Reader, the Acucy 2 Reader has two modes of operation, the WALK AWAY and the READ NOW modes, described below:

#### a. WALK AWAY Mode:

In the WALK AWAY mode, Test Cassettes are inserted into the reader. Then, after the addition of the sample to the Test Cassette, the development and timing of the test occur within the reader. This allows the user to "walk away" until the test has been completed.

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The following changes have been made to the "WALK AWAY" mode in the user interface:

- The user can now select WALK AWAY button from MAIN MENU
- All screens in WALK AWAY test mode have a blue progress bar at the top.
- The user does not log in to access this mode unlike Acucy 1.
- The user now enters the Patient ID or can, on the same screen, instruct the Acucy 2 to scan the barcode on the test cassette.
- The user now has a reference to the instructions for use (IFU) on the screen to access more details for sample preparation (if needed).
- The Analyzing Sample screen now has a progress bar indicating time to result (TTR).
- The user can run another test in WALK AWAY mode or return to MAIN MENU.

b. READ NOW Mode:

In the READ NOW mode, the reader analyzes the results after manually timing the development of the Test Cassette on a flat surface for the full 15 minutes.

The following changes have been made on the "READ NOW" mode in the user interface.

- The user can now select READ NOW button from MAIN MENU
- All screens in READ NOW test mode have a blue progress bar at the top.
- The user does not log in to access this mode unlike the Acucy 1.

3. Calibration:

The calibration test is a required function for Acucy 2 Reader that checks the Reader optics and calculation systems using the calibration zone label affixed to the drawer. The calibration procedure is performed upon installation to activate the quality control and READ NOW/WALK AWAY functionality and is required every 30-days. The operator is prompted by the Reader to conduct calibration after 30-days has elapsed. The calibration procedure may also be performed as directed during troubleshooting or whenever the Reader date and time has been changed.

4. Quality Control:

There are three types of quality controls for the Acucy 2 and Acucy reader and Influenza A&B Test systems:

- Acucy 2 Reader calibration (described above)
- Internal control line integrated into the test strip of the Test Cassette (unchanged from K182001)
- External quality controls: Influenza A+/B- and A-/B+ Control Swabs (unchanged from K182001)

V. Substantial Equivalence Information:

A Predicate Device Name(s):

Acucy Influenza A&B Test with the Acucy System

B Predicate 510(k) Number(s):

K182001

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C Comparison with Predicate(s):

| Device & Predicate Device(s): | K241188
(Candidate Device) | K182001
(Predicate Device) |
| --- | --- | --- |
| Device Trade Name | Acucy Influenza A&B Test with the Acucy 2 Reader | Acucy Influenza A&B Test with the Acucy System |
| General Device Characteristic Similarities | | |
| Intended Use/Indications For Use | The Acucy Influenza A&B Test is a rapid chromatographic immunoassay for the qualitative detection and differentiation of influenza A and B viral nucleoprotein antigens directly from anterior nasal and nasopharyngeal swabs from patients with signs and symptoms of respiratory infection. The test is intended for use with the Acucy or Acucy 2 Reader as an aid in the diagnosis of influenza A and B viral infections. The test is not intended for the detection of influenza C viruses. Negative test results are presumptive and should be confirmed by viral culture or an FDA-cleared influenza A and B molecular assay. Negative test results do not preclude influenza viral infection and should not be used as the sole basis for treatment or other patient management decisions.

Performance characteristics for influenza A were established during the 2017-2018 influenza season when influenza A/H3N2 and A/H1N1pdm09 were the predominant influenza A viruses in circulation. When other influenza A viruses are emerging, performance characteristics may vary. | The Acucy Influenza A&B Test for the rapid qualitative detection of influenza A and B is composed of a rapid chromatographic immunoassay for the direct and qualitative detection of influenza A and B viral nucleoprotein antigens from nasal and nasopharyngeal swabs of symptomatic patients that is automatically analyzed on the Acucy Reader. The Acucy Influenza A&B Test is a differentiated test, such that influenza A viral antigens can be distinguished from influenza B viral antigens from a single processed sample using a single Test Cassette. The test is intended for use with the Acucy System as an aid in the diagnosis of influenza A and B viral infections. The test is not intended for the detection of influenza C viruses. Negative test results are presumptive and should be confirmed by viral culture or an FDA-cleared influenza A and B molecular assay. Negative test results do not preclude influenza viral infection and should not be used as the sole basis for treatment or other patient management decisions.

Performance characteristics for influenza A were established during the 2017-2018 influenza season when influenza A/H3N2 and A/H1N1pdm09 were the predominant influenza A viruses in circulation. When other influenza A viruses are emerging, performance characteristics may vary. |

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| Device & Predicate Device(s): | K241188
(Candidate Device) | K182001
(Predicate Device) |
| --- | --- | --- |
| | 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 department for testing. Viral culture should not be attempted in these cases unless a BSL 3+ facility is available to receive and culture specimens | 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 department for testing. Viral culture should not be attempted in these cases unless a BSL 3+ facility is available to receive and culture specimens. |
| Assay Results | Qualitative | Same |
| Assay Targets | Influenza A and B nucleoprotein antigens | Same |
| Assay Format | Lateral flow test cassette | Same |
| Sample Types | Anterior nasal swab and nasopharyngeal swab | Same |
| Assay Antibodies | Monoclonal antibodies to influenza A and B nucleoproteins. | Same |
| Additional Reagents | Extraction buffer with detergents | Same |
| Sample Transfer Method | Dropper tip applied to extraction vial to transfer extracted sample onto test cassette. | Same |
| Reporting of Results | Reader displays results on screen; or may be printed | Same |
| Time to Results | 15 Minutes | Same |
| Instrument Modes | READ NOW or WALK AWAY | Same |
| Storage Temperature | Room Temperature | Same |
| Assay Controls | Internal procedural control | Same |
| Instrument Quality Control Features | • Scanning procedural control zone for adequate flow
• Reader prevents use of used cassettes.
• Reader prevents use of expired cassettes.
• Reader prevents improper cassette insertion | Same |
| General Device Characteristic Differences | | |
| Calibrator | External Calibration Device | Internal Calibration device. |
| Imaging Mode | Photodiode | CMOS Camera |
| Printer | Included | Optional |

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| Device & Predicate Device(s): | K241188
(Candidate Device) | K182001
(Predicate Device) |
| --- | --- | --- |
| LCD Display/Graphic User Interface | | |
| Cybersecurity | • Removal of Master Password Card replaced with FORGOT PIN.
• Allows adjustment of time of Auto-logout at supervisor level access. | • Operator/supervisor passwords
• Unique Master Password Card
• Auto-logout after 60 minutes |

VI. Standards/Guidance Documents Referenced:

| Document | Title | Publisher | Applicable Study |
| --- | --- | --- | --- |
| Special Controls 21 CFR 862.2160 | Special Controls for Influenza virus antigen detection test systems | FDA/CDRH | All Studies |
| CLSI EP17-A2 | Evaluation of Detection Capability for Clinical Laboratory Measurement Procedures, 2nd Edition | CLSI | Detection Limit |
| CLSI EP12-A2 | Evaluation of Qualitative, Binary Output Examination Performance - 3rd Edition | CLSI | Analytical Studies |
| IEC/EN 61326-1Ed.3.0 b:2020 | Electrical equipment for measurement, control, and laboratory use - EMC requirements - Part 1: General requirements | IEC/EN | EMC |
| ISO 11135:2014 | Sterilization of health care products - Ethylene oxide Requirements for | ISO | Sterility |

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| Document | Title | Publisher | Applicable Study |
| --- | --- | --- | --- |
| | development, validation and routine control of a sterilization process for medical devices. | | |
| ISO 10993-7:2008 | Biological evaluation of medical devices - Part 7: Ethylene oxide sterilization residuals | ISO | Sterility |

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

### A Analytical Performance:

#### 1. Precision/Reproducibility

##### a. Precision/Reproducibility Panel:

In three different studies, the same seven-member sample panel was used to evaluate the Acucy Influenza A&B Test with the Acucy 2 Reader for repeatability, instrument-to-instrument precision, and between site reproducibility. Each panel member was made with two viral strains: Influenza A/Michigan/45/2015 Strain and Influenza B/Colorado/06/2017. Each strain was diluted into negative clinical matrix (NCM) at three different target levels (see Table 1 below):

Simulated nasal swabs were prepared by pipetting 50 µL of sample (virus dilution) onto the head of the swab. Each swab was allowed to air dry for one (1) minute before testing per IFU was performed.

For MP and LP samples, the expected result is detection of virus (positive result). For HN and N samples, the expected result is the absence of virus (negative result) is, as summarized in Table 1 below:

Table 1. Sample Panel Tested in the Precision Study with Expected Detection Rates

| Panel Member | Target Concentration | | Target Positive Detection Rate |
| --- | --- | --- | --- |
| Negative | (N) | 0X LoD | 0% |
| High Negative Flu A or Flu B | (HN) | 0.05X LoD | 0-5% |
| Low Positive Flu A or Flu B | (LP) | 0.95X LoD | ≥95 % |
| Moderate Positive Flu A or Flu B | (MP) | 2X LoD | 100% |

##### b. Repeatability (Within-Laboratory Precision):

The seven-member panel was tested at one internal site two (2) times per day (2 runs), each with two (2) replicates per run and for twenty (20) non-consecutive days. The testing was conducted by one (1) operator using one (1) lot of the Acucy Influenza A&B Test and one Acucy 2 Reader. All samples produced expected results as summarized in Table 2 below:

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Table 2. Within-Laboratory Repeatability Study Results

| Sample | Count | Agreement | 95% CI |
| --- | --- | --- | --- |
| Flu A MP | 80/80 | 100% | 95.4% - 100% |
| Flu A LP | 80/80 | 100% | 95.4% - 100% |
| Flu A HN | 80/80 | 100% | 95.4% - 100% |
| Flu B MP | 80/80 | 100% | 95.4% - 100% |
| Flu B LP | 80/80 | 100% | 95.4% - 100% |
| Flu B HN | 80/80 | 100% | 95.4% - 100% |
| Negative | 80/80 | 100% | 95.4% - 100% |

c. Instrument-to-Instrument Precision:

For between-instrument precision, the seven-member panel was tested on three (3) different Acucy 2 Readers over the course of five (5) days at the manufacturer's internal site. Testing was conducted by one operator using one lot of the Acucy Influenza A&B Test testing five (5) replicates per panel member and run. Twenty-five (25) replicates were obtained per panel member per instrument (1 operator x 5 replicates x 5 days), for a total of 75 replicates per panel member (1 operator x 5 replicates x 3 instruments x 5 days). All samples produced expected results as summarized in Table 3 below:

Table 3. Instrument-to-Instrument Precision Study Results

| Sample | Reader #1 | Reader #2 | Reader #3 | Overall (All Readers) | |
| --- | --- | --- | --- | --- | --- |
| | %Agreement (Count) | %Agreement (Count) | %Agreement (Count) | Agreement (Count) | 95% CI |
| Flu A (MP) | 100% (25/25) | 100% (25/25) | 100% (25/25) | 100% (75/75) | 95.1% - 100% |
| Flu A (LP) | 100% (25/25) | 100% (25/25) | 100% (25/25) | 100% (75/75) | 95.1% - 100% |
| Flu A (HN) | 100% (25/25) | 100% (25/25) | 100% (25/25) | 100% (75/75) | 95.1% - 100% |
| Flu B (MP) | 100% (25/25) | 100% (25/25) | 100% (25/25) | 100% (75/75) | 95.1% - 100% |
| Flu B (LP) | 100% (25/25) | 100% (25/25) | 100% (25/25) | 100% (75/75) | 95.1% - 100% |
| Flu B (HN) | 100% (25/25) | 100% (25/25) | 100% (25/25) | 100% (75/75) | 95.1% - 100% |
| Negative | 100% (25/25) | 100% (25/25) | 100% (25/25) | 100% (75/75) | N/A |

d. Multi-Site Reproducibility Study:

To assess between-site reproducibility, the seven-member panel was tested at three testing sites for five (5) non-consecutive days over a period of two (2) weeks. At each site, testing was conducted by two (2) operators using the same Acucy Influenza A&B Test lot and testing three (3) replicates per panel member and run. Each site had their own unique Acucy 2 Reader. A total of ninety (90) replicates for each panel member were tested across three (3) sites to evaluate reproducibility (2 operators x 3 replicates x 5 days x 3 sites).

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This study was used to supplement the original reproducibility study used to support the clearance of the Acucy Influenza A&B Test with the Acucy System (K182001) to demonstrate that performance remains equivalent. Results were acceptable, and are summarized in Table 4 below:

Table 4. Multi-Site Reproducibility Study Results

| Sample | Site #1 | Site #2 | Site #3 | Overall (All Sites) | |
| --- | --- | --- | --- | --- | --- |
| | Agreement (Count) | Agreement (Count) | Agreement (Count) | Agreement (Count) | 95% CI |
| Flu A (MP) | 100% (30/30) | 100% (30/30) | 100% (30/30) | 100% (90/90) | 95.9-100.0% |
| Flu A (LP) | 100% (30/30) | 100% (30/30) | 100% (30/30) | 100% (90/90) | 95.9-100.0% |
| Flu A (HN) | 100% (30/30) | 100% (30/30) | 100% (30/30) | 100% (90/90) | 95.9-100.0% |
| Flu B (MP) | 100% (30/30) | 100% (30/30) | 100% (30/30) | 100% (90/90) | 95.9-100.0% |
| Flu B (LP) | 100% (30/30) | 100% (30/30) | 100% (30/30) | 100% (90/90) | 95.9-100.0% |
| Flu B (HN) | 100% (30/30) | 100% (30/30) | 100% (30/30) | 100% (90/90) | 95.9-100.0% |
| Negative | 100% (30/30) | 100% (30/30) | 100% (30/30) | 100% (90/90) | 95.9-100.0% |

2. Analytical Specificity/Interference:

The analytical specificity and interference testing (Inclusivity, Cross Reactivity, Microbial Interference, Endogenous/Exogenous Interference, Competitive Interference) were conducted in support of K182001 and were leveraged for this submission. Refer to the Decision Summary of K182001 for further details.

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

a. Instrument Calibration

The Acucy 2 Reader's optics and calculation systems are calibrated using the calibration zone label affixed to the drawer. This calibration is performed upon installation to activate the instrument's functionality and is required every 30-days. The operator is prompted by the Reader to conduct calibration after 30-days has elapsed. The calibration procedure may also be performed as directed during troubleshooting or whenever the Reader date and time has been changed.

b. Internal Quality Control

The Acucy Influenza A&B Test Cassette contains a built-in internal control feature. Each time a test is run in the Acucy 2 Reader, the internal control zone is scanned by the reader. A

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"VALID" test result displayed by the reader indicates that the internal control was present, demonstrates that the test flowed correctly, and that the functional integrity of the Test Cassette and reagents was maintained. An "INVALID" test result displayed by the reader indicates that the internal control was not present, demonstrates that the test did not flow correctly, and/or that the functional integrity of the test cassette and reagents were not maintained. Should this occur, the end user is instructed to review the testing procedure and repeat the test using a new patient sample, test cassette, and reagents.

## c. External Quality Controls

The Acucy Influenza A&B Test with Acucy 2 Reader includes one Influenza A+/B- control swab (Red Label) and one Influenza A-/B+ control swab, each of which contains inactivated virus, for external quality control testing. These controls are identical to those used with the Acucy Reader and were validated in K182001.

## d. Swab Equivalency

The test is intended for use with anterior nasal (ANS) and nasopharyngeal (NP) swabs. A swab equivalency test was performed in K182001 that demonstrated equivalent performance between ANS and NP swabs and was also used in support of this submission.

## e. Test Mode Equivalency:

This study was performed to validate equivalent performance of the READ NOW and WALK AWAY test modes when reading Acucy Influenza A&B Test results on the Acucy 2 Reader.

Contrived positive Influenza A (A/Michigan/45/2015) and Influenza B (B/Colorado/06/2017) samples were prepared at 2X LoD, and negative samples were prepared in NCM. For each sample replicate, 50 μL of contrived sample was pipetted onto a swab and tested per the IFU of the test. Testing was conducted with one lot of the Acucy Influenza A&B Test and two unique Acucy 2 Readers.

Twenty (20) replicates of each contrived sample were tested using the READ NOW mode and 20 replicates of each contrived sample were also tested using the WALK AWAY mode. Results of the test mode equivalency study are shown in Table 5 below. The samples showed 100% results concurrence in both modes and similar measured mean absorbance (mAbs) values for all test lines. This data supports use of either test mode for analytical testing.

Table 5. Test Mode Equivalency Study Results

| | READ NOW | WALK AWAY |
| --- | --- | --- |
| Flu A | | |
| Percent Agreement | 100% (20/20) | 100% (20/20) |
| Mean mAbs | 39.0 mAbs | 39.4 mAbs |
| % CV | 9.6% | 9.8% |
| Flu B | | |
| Percent Agreement | 100% (20/20) | 100% (20/20) |
| Mean mAbs | 45.2 mAbs | 44.8 mAbs |
| % CV | 18.2% | 18.4% |

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| | READ NOW | WALK AWAY |
| --- | --- | --- |
| Negative | | |
| Percent Agreement | 100% (20/20) | 100% (20/20) |
| Mean mAbs | 0 mAbs | 0 mAbs |
| % CV | NA | NA |

# 4. Limit of Detection (LoD):

The LoD of the test for two strains of Influenza A (A/Michigan/45/2015 (H1N1) and A/Singapore/INFIMH-16-0019/2016 (H3N2) and two strains of Influenza B (B/Phuket/3073/2013 (Yamagata Lineage) and B/Colorado/06/2017 (Victoria Lineage) was determined using two lots of Acucy Influenza A and B test cassettes on each, the Acucy reader and the Acucy 2 reader. The LoD was determined as the lowest virus concentration that was detected $\geq 95\%$ of the time (i.e., concentration at which at least 19 out of 20 replicates tested positive). The limit of detection was established in two (2) phases: a range finding and a confirmatory LoD study.

# a. Range Finding LoD Study:

Contrived samples were prepared by diluting Influenza A or B virus in PCR negative NCM. Five (5) replicates of a 1:10 dilution series were tested on two lots of Acucy cassettes, using both Acucy 1 and Acucy 2 readers. For each test, $50~\mu \mathrm{L}$ was pipetted onto the head of an anterior nares swab (ANS). Swabs were then processed per the IFU, using the READ NOW mode. The lowest concentration of each lot with 4/5 positive results was selected as a preliminary LoD for confirmation. The preliminary LoD study results are shown in Table 6.

Table 6: Preliminary Testing Results for LoD Study

| Viral Strain | Concentration (TCID50/mL) | Acucy 1 Reader | | Acucy 2 Reader | |
| --- | --- | --- | --- | --- | --- |
| | | Test Lot 1 | Test Lot 2 | Test Lot 1 | Test Lot 2 |
| Influenza A/Michigan/45/2015 (H1N1) | 1.41E+04 | 5/5 | 5/5 | 5/5 | 5/5 |
| | 1.41E+03 | 5/5 | 5/5 | 5/5 | 5/5 |
| | 1.41E+02 | 5/5 | 5/5 | 5/5 | 5/5 |
| | 1.41E+01 | 0/5 | 0/5 | 0/5 | 0/5 |
| | 1.41E+00 | 0/5 | 0/5 | 0/5 | 0/5 |
| | 1.41E-01 | 0/5 | 0/5 | 0/5 | 0/5 |
| Influenza A/Singapore/INFIMH-16-0019/2016 (H3N2) | 3.16E+05 | 5/5 | 5/5 | 5/5 | 5/5 |
| | 3.16E+04 | 5/5 | 5/5 | 5/5 | 5/5 |
| | 3.16E+03 | 5/5 | 5/5 | 5/5 | 5/5 |
| | 3.16E+02 | 0/5 | 0/5 | 0/5 | 0/5 |
| | 3.16E+01 | 0/5 | 0/5 | 0/5 | 0/5 |
| | 3.16E+00 | 0/5 | 0/5 | 0/5 | 0/5 |
| Influenza B/Phuket/3073/2013 (Yamagata Lineage) | 4.17E+04 | 5/5 | 5/5 | 5/5 | 5/5 |
| | 4.17E+03 | 5/5 | 5/5 | 5/5 | 5/5 |
| | 4.17E+02 | 5/5 | 5/5 | 5/5 | 5/5 |
| | 4.17E+01 | 0/5 | 0/5 | 0/5 | 0/5 |
| | 4.17E+00 | 0/5 | 0/5 | 0/5 | 0/5 |
| | 4.17E-01 | 0/5 | 0/5 | 0/5 | 0/5 |

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| Viral Strain | Concentration (TCID50/mL) | Acucy 1 Reader | | Acucy 2 Reader | |
| --- | --- | --- | --- | --- | --- |
| | | Test Lot 1 | Test Lot 2 | Test Lot 1 | Test Lot 2 |
| Influenza B/Colorado/06/2017 (Victoria Lineage) | 1.41E+04 | 5/5 | 5/5 | 5/5 | 5/5 |
| | 1.41E+03 | 5/5 | 5/5 | 5/5 | 5/5 |
| | 1.41E+02 | 4/5 | 0/5 | 4/5 | 1/5 |
| | 1.41E+01 | 0/5 | 0/5 | 0/5 | 0/5 |
| | 1.41E+00 | 0/5 | 0/5 | 0/5 | 0/5 |
| | 1.41E-01 | 0/5 | 0/5 | 0/5 | 0/5 |

b. Confirmatory LoD Study

After determining the target dilution for each strain, serial 2-fold dilutions were made corresponding to five (5) low level samples around the LoD ranges determined in the preliminary studies. These concentrations were tested over three (3) days for a total of 20 replicates per concentration and the lowest concentration with $\geq 95\%$ of samples generating positive results was designated as the LoD for that lot for the strain tested, as shown in Table 7 below.

Table 7. Final Testing for LoD Study

| Viral Strain | Preliminary LoD (TCID50/mL) | Tested Concentration (TCID50/mL) | Acucy 1 Reader | | Acucy 2 Reader | |
| --- | --- | --- | --- | --- | --- | --- |
| | | | Test Lot 1 | Test Lot 2 | Test Lot 1 | Test Lot 2 |
| Influenza A/Michigan/45/2015 (H1N1) | 1.41E+02 | 2.82E+02 | 20/20 | 20/20 | 20/20 | 20/20 |
| | | 1.41E+02 | 17/20 | 18/20 | 17/20 | 18/20 |
| | | 7.05E+01 | 5/20 | 2/20 | 4/20 | 4/20 |
| | | 3.53E+01 | 2/20 | 0/20 | 0/20 | 0/20 |
| | | 1.76E+01 | 0/20 | 0/20 | 0/20 | 0/20 |
| Influenza A/Singapore/IN FIMH-16-0019/2016 (H3N2) | 3.16E+03 | 6.32E+03 | 20/20 | 20/20 | 20/20 | 20/20 |
| | | 3.16E+03 | 20/20 | 20/20 | 20/20 | 20/20 |
| | | 1.58E+03 | 14/20 | 9/20 | 15/20 | 10/20 |
| | | 7.90E+02 | 0/20 | 0/20 | 0/20 | 0/20 |
| | | 3.95E+02 | 0/20 | 0/20 | 0/20 | 0/20 |
| Influenza B/Phuket/3073/2013 (Yamagata Lineage) | 4.17E+02 | 8.34E+02 | 20/20 | 20/20 | 20/20 | 20/20 |
| | | 4.17E+02 | 20/20 | 20/20 | 20/20 | 20/20 |
| | | 2.09E+02 | 19/20 | 13/20 | 20/20 | 17/20 |
| | | 1.04E+02 | 14/20 | 0/20 | 18/20 | 5/20 |
| | | 5.02E+01 | 0/20 | 0/20 | 0/20 | 0/20 |
| Influenza B/Colorado/06/2017 (Victoria Lineage) | 1.41E+02 (Lot 1) | 2.82E+02 | 19/20 | NA | 20/20 | NA |
| | | 1.41E+02 | 9/20 | NA | 17/20 | NA |
| | | 7.05E+01 | 1/20 | NA | 3/20 | NA |
| | | 3.53E+01 | 0/20 | NA | 0/20 | NA |
| | | 1.76E+01 | 0/20 | NA | 0/20 | NA |
| | 1.41E+03 (Lot 2) | 2.82E+03 | NA | 20/20 | NA | 20/20 |
| | | 1.41E+03 | NA | 20/20 | NA | 20/20 |
| | | 7.05E+02 | NA | 20/20 | NA | 20/20 |
| | | 3.53E+02 | NA | 17/20 | NA | 17/20 |
| | | 1.76E+02 | NA | 3/20 | NA | 5/20 |

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The established LoDs for the Acucy and Acucy 2 Reader are identical for each cassette lot, indicating that the performance of the predicate and candidate instruments is similar.

# 5. Assay Cut-Off:

The initial assay threshold of 5.0 milli-absorbance units (mAbs) was established as a default setting for the Acucy 2 Reader by the instrument developer and validated with a limit of blank study.

The Limit of Blank (LoB) of the Acucy Influenza A&B Assay on the Acucy 2 System was determined by testing sixty (60) replicates of a blank sample (pooled NCM negative for Influenza A and B) on 2 lots of Acucy Influenza A/B cartridges. All blank samples tested on Acucy 2 Reader showed $0\mathrm{mAbs}$ for both the Influenza A and Influenza B lines.

The analytical instrument cut-offs for influenza A and B that were original established for the Acucy system were determined to be 6.4 and $5.4\mathrm{mAbs}$ , respectively. As such, the Acucy Influenza A and B Test specific cut-offs for the Acucy 2 Reader are not significantly different from those previously established for the Acucy reader. Since the Acucy 2 reader uses similar detection technology than the Acucy reader, the assay specific cut-off for the Acucy 2 reader were set to the same values previously established for the Acucy reader.

# 6. Carry-Over:

This study evaluated the potential of sample carry-over between test cassettes when high-titer samples were run in an alternating fashion with negative samples.

Thirty (30) high titer influenza A samples were prepared using Influenza A/Michigan and Influenza B/Colorado strains diluted in NCM to a concentration of $1 \times 105$ TCID50/mL. Thirty (30) negative samples were prepared using NCM. For each contrived sample, $50~\mu \mathrm{L}$ was applied to nasal swabs, allowed to dry for up to one minute, and tested according to the device's IFU using the READ NOW mode. Positive and negative samples were alternated so that a negative sample was always run immediately after a positive sample. A false positive result for a negative sample would suggest contamination from the previous test.

All influenza A and influenza B positive samples gave positive results as expected. All negative samples gave negative results, as summarized in Table 8 below:

Table 8. Summary of Carry-Over Study Results

| Sample | Flu A Positive | Flu B Positive | Negative |
| --- | --- | --- | --- |
| Influenza A High Positive | 30/30 (100%) | 0/30 (0%) | 0/30 (0%) |
| Influenza B High Positive | 0/30 (0%) | 30/30 (100%) | 0/30 (0%) |
| Negative | 0/60 (0%) | 0/60 (0%) | 60/60 (100%) |

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# B Comparison Studies:

Method Comparison with Predicate Device:

This study evaluated the systemic differences between the Acucy and the Acucy 2 Reader using the same samples.

Aliquots of $50\mu \mathrm{L}$ of known Influenza A or Influenza B positive or Influenza A and B negative nasopharyngeal swab (NPS) remnant samples were spiked onto sterile swabs and tested in parallel on both instruments per the IFU of the test. These samples were representative of a range of virus levels with RT-PCR values ranging from Ct 11.5 - 32.4.

Testing was performed as per Acucy Influenza A&B instructions for use. Thirty each of Influenza A and Influenza B samples and 30 Influenza A/B negative clinical samples were tested using one lot of Acucy Influenza A&B test kits. Each cassette was read on both the Acucy Reader and an Acucy 2 Reader. The Influenza A positive samples are considered as negatives for Influenza B and those positive for Influenza B are considered negative for Influenza A. Thus, together with the double negative samples, for each group, there are a total of sixty (60) negative results. All the positive samples showed up as positive on both readers. Results for the performance of the Acucy Influenza A&B Test for Influenza A and Influenza B stratified by the two Acucy Readers are in Tables 9 and 10 below:

Table 9. Influenza A performance with Archived Nasopharyngeal Swab Specimens Using READ NOW Mode on Acucy 2 Reader Compared to Acucy Reader.

| Flu A by Acucy 2 Reader | Flu A by Acucy Reader | |
| --- | --- | --- |
| | Positive | Negative |
| Positive | 30 | 1* |
| Negative | 0 | 59 |
| PPA = 100.0 % (95% CI: 88.7% - 100.0 %) NPA = 98.3 % (95% CI: 91.1% - 99.7 %) | | |
| *One of the Flu B positive samples returned a double positive result (positive for Flu A and Flu B) on Acucy 2 Reader but not on the Acucy Reader. | | |

For Influenza A, the results of the method comparison with the predicate demonstrated $100\%$ PPA (95% CI: 88.7% -100%) and 98.3% NPA (95% CI: 91.1% - 99.7%) One of the Flu B positive samples returned a positive result for Flu A as well (double positive) when read on Acucy 2 Reader.

Table 10. Influenza B performance with Archived Nasopharyngeal Swab Specimens Using READ NOW Mode on Acucy 2 Reader Compared to Acucy Reader

| Flu B by Acucy 2 Reader | Flu B by Acucy Reader | |
| --- | --- | --- |
| | Positive | Negative |
| Positive | 30 | 0 |
| Negative | 0 | 60 |
| PPA = 100.0 % (95% CI: 88.7% - 100.0 %) NPA = 100.0 % (95% CI: 94.0.% - 100.0 %) | | |

For Influenza B, the results of the method comparison with predicate device demonstrated $100.0\%$ PPA (95% score CI: $88.7\% - 100.0\%$ ) and $100\%$ NPA (95% score CI: $94.0\% - 100.0\%$ ) as compared to the Acucy 2 reader.

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C Clinical Studies:

The clinical performance was previously reviewed under K182001 and leveraged to support performance for the Acucy Influenza A&B Test with the Acucy 2 System.

D Other Supportive Instrument Performance Characteristics Data:

Not applicable.

VIII. Proposed Labeling:

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

IX. Conclusion:

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

The submitted information in this CLIA waiver application supports a CLIA waiver approval decision.

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