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The Verigene® Respiratory Virus Plus Nucleic Acid Test (RV+) on the Verigene® System is a qualitative nucleic acid multiplex test intended to simultaneously detect and identify multiple respiratory virus nucleic acids in nasopharyngeal (NP) swab specimens from individuals with signs and symptoms of respiratory tract infection. The following virus types and subtypes are identified using the RV+: Influenza A, Influenza A subtype H1, Influenza A subtype H3, 2009 H1N1, Influenza B, Respiratory Syncytial Virus (RSV) subtype A, and RSV subtype B. The test is not intended to detect Influenza C virus. Detecting and identifying specific viral nucleic acids from individuals exhibiting signs and symptoms of respiratory infection aids in the diagnosis of respiratory viral infection, if used in conjunction with other clinical and laboratory findings.

Negative results for Influenza A, Influenza B, or RSV do not preclude influenza virus or RSV infection and should not be used as the sole basis for diagnosis, treatment, or patient management decisions. Conversely, positive results do not rule-out bacterial infection or co-infection with other viruses. The agent detected may not be the definite cause of disease. The use of additional laboratory testing and clinical presentation must be considered in order to obtain the final diagnosis of respiratory viral infection.

Performance characteristics for Influenza A Virus were established when Influenza A/H3, A/H1, and 2009 H1N1 were the predominant Influenza A viruses circulating. These characteristics may vary when other Influenza A viruses are emerging.

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 used specifically 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.

The entire RV+ test is performed on the Verigene® System, which is a bench-top molecular diagnostics workstation that consists of two instruments, the Verigene Processor SP and the Verigene Reader. The Verigene Processor SP performs the assay steps on each sample by using a robotic pipettor to transfer and mix reagents within and between separate testing modules designed for nucleic acid extraction, target amplification, and the Verigene Hybridization Test. The Verigene Hybridization Test module is the same as in the original Verigene System with added modules for nucleic acid extraction and RT-PCR target amplification. Key functions of the Verigene Processor SP include: 1) Reading of the barcode identification label on inserted Test Consumables to maintain positive identification of patient samples throughout processing. 2) Facilitation of nucleic acid extraction, multiplex RT-PCR target amplification, and the Verigene Hybridization Test. 3) Real-time communication of test processing status to the Reader.

The Verigene Reader is the same instrument as in the FDA-cleared RVNATSP. It is a free-standing instrument with a touch screen control panel and a wand-based barcode scanner. It utilizes a graphical user interface to guide the user through the process of ordering tests and reporting results. There are no serviceable parts and no user calibration is required. Interaction with the touch screen is minimized through barcode use. This instrument also serves as the reader of the Test Cartridges using advanced optics. The key functions of the Verigene Reader include: 1) Entry and tracking of specimen identification numbers via manual keyboard input or via barcode-reader wand. 2) Test selection for each specimen. 3) Automated transfer of specimen processing instructions on Test Cartridge-specific basis to linked Processor SP unit(s). A single Reader unit can control up to 32 Processor units. 4) Automated imaging and analysis of Test Cartridges. 5) Results display. 6) Results report generation.

RV+ consumables within each single-use disposable test kit include: (i) Tip Holder Assembly; (ii) Extraction Tray; (iii) Amplification Tray; and (iv) RV+ Test Cartridge. The kit components are inserted into the corresponding module of the Verigene Processor SP prior to each test, and the sample is added to the Extraction Tray. Patient information is entered into the Reader to initiate the test procedure.

1. Tip Holder Assembly - The robotic pipettor picks up pipettes from the Tip Holder Assembly. The pipettes are used for mixing and transferring reagents within the test procedure.
2. Extraction Tray – Nucleic acids are extracted from the sample by using magnetic bead-based methods within the Extraction Tray. Each Tray contains reagents for a single extraction procedure. A robotic pipette transfers reagents to designated wells within the Extraction Tray to affect the steps of lysis, capture of nucleic acids onto the magnetic beads, washing, and eluting the isolated nucleic acids from the magnetic beads.
3. Amplification Tray – The isolated nucleic acids are amplified by using multiplex RT-PCR within the Amplification Tray. Each Tray contains reagents for a single multiplex RT-PCR procedure. A robotic pipette transfers the reagents to a specific well within the Amplification Tray. A set thermal profile is then initiated to perform all of the amplification related steps including UDG-based decontamination, reverse transcription, and multiplex PCR in a single tube. Upon completion, an aliquot of the amplified sample is mixed with hybridization buffer containing the virus specific mediator probes. The sample is then transferred to the Test Cartridge.
4. RV+ Test Cartridge for Verigene Hybridization Test – The virus-specific and subtype-specific amplicons are detected and identified within a Test Cartridge by using specific nucleic acid probes in conjunction with gold nanoparticle probe-based detection technology. Each Test Cartridge is a self-contained, laboratory consumable that consists of two parts. The upper housing of each cartridge is called the "reagent pack" and contains reservoirs filled with the detection reagents. When in place with the 'substrate holder', the reagent pack creates an air-tight hybridization chamber surrounding the region of the substrate containing a target-specific capture array. As each step of the test is completed, old reagents are moved out of the hybridization chamber and new reagents are added from the reagent pack via microfluidic channels and pumps. Once the test is complete, the Test Cartridge is removed from the Verigene Processor SP unit and the reagent pack is snapped off and discarded. The remaining slide is now ready for imaging and analysis in the Verigene Reader.
5. End-point detection on the Verigene Reader: The test slide is inserted into the Verigene Reader wherein it is illuminated along its side. The gold-silver aggregates at the test sites scatter the light, which is in turn captured by a photosensor. The relative intensity arising from each arrayed test site is tabulated. Net signals, defined as the absolute signal intensities with background signals subtracted, are compared with thresholds determined by negative controls within the slide in order to arrive at a decision regarding the presence or absence of target. These results are linked to the test and patient information entered at the beginning of each test session to provide a comprehensive results file.

Here's an analysis of the acceptance criteria and the study proving the device meets those criteria, based on the provided text:

Acceptance Criteria and Device Performance for Verigene® Respiratory Virus Plus Nucleic Acid Test (RV+)

1. Table of Acceptance Criteria and Reported Device Performance

The document does not explicitly state pre-defined acceptance criteria in terms of specific sensitivity and specificity thresholds. Instead, it presents the "Performance Characteristics" from a methods comparison study, and the acceptance is implied by the FDA's clearance of the device (K103209). Therefore, the "acceptance criteria" here are interpreted as the observed performance deemed acceptable for FDA clearance, and the "reported device performance" refers to the results from the clinical methods comparison study.

Reproducibility/Precision Study:
The document also presents reproducibility data, interpreted as another form of acceptance criteria for device performance stability.

• Total Agreement for all panel members across all 3 sites: 97.2% - 100% (95% CI range from 90.3% - 99.7% to 95.0% - 100.0%).
• "No Call" rate: 1.6% (14/864)
• "Pre-analytical error" failure rate: 0.2% (2/864)

2. Sample Size Used for the Test Set and Data Provenance

• Sample Size for Test Set: 1022 prospectively-collected specimens were used for the methods comparison study.
• Data Provenance:
  • Country of Origin: Not explicitly stated, but the study was conducted at "three collection hospital sites" and shipped to "Nanosphere" (located in Northbrook, IL, USA) for processing and then to "testing sites." Given the FDA submission from a US-based company, it is highly probable the data is from the USA.
  • Retrospective or Prospective: The samples were collected prospectively during the 2008-2009 and 2009-2010 respiratory seasons. Residual specimens were then de-identified, frozen, and shipped to Nanosphere, then shipped to testing sites.

3. Number of Experts Used to Establish the Ground Truth for the Test Set and Qualifications of Those Experts

• Number of Experts: Not explicitly stated as "experts." The ground truth was established using a combination of methods:
  • Culture-based methods confirmed with an FDA-cleared Direct Fluorescent Antibody (DFA) test for Influenza A, Influenza B, and RSV.
  • Bidirectional sequencing for Influenza A subtyping (H3, H1, 2009 H1N1) and RSV subtyping (RSV A, RSV B).
  • For discordant results between RV+ and culture/DFA, bidirectional sequencing and/or NAAT (Nucleic Acid Amplification Test) was used for resolution.
• Qualifications of Experts: Not specified. The reference methods mentioned are laboratory-based standard tests, implying qualified laboratory personnel perform these tests, but no specific "expert" role or qualifications (e.g., years of experience) are detailed.

4. Adjudication Method for the Test Set

The adjudication method for the test set involved a multi-step process for resolving discrepancies:

• Initial comparison of RV+ results against culture-based methods confirmed with FDA-cleared DFA.
• For Influenza A subtyping and RSV subtyping, bidirectional sequencing was used as the primary comparative method.
• For discordant results between RV+ and culture/DFA, further clarification was sought using bi-directional sequencing and/or NAAT.
• Specific notes for individual discrepancies (e.g., footnote comments below the tables) illustrate this process, where sequencing or repeat testing with NAAT/culture informed the final ground truth. This resembles a tie-breaker adjudication approach for discordant results.

5. If a Multi-Reader Multi-Case (MRMC) Comparative Effectiveness Study Was Done

• No, a multi-reader multi-case (MRMC) comparative effectiveness study was NOT mentioned. This study focuses on the standalone performance of the RV+ device against established laboratory methods, not on comparing human reader performance with and without AI assistance. The device itself is a molecular diagnostic test, not an AI-powered image analysis tool for human readers.

6. If a Standalone (i.e., algorithm only without human-in-the-loop performance) Was Done

• Yes, a standalone performance study was conducted. The methods comparison study evaluated the RV+ system (which includes sample preparation, target amplification, and a Verigene Hybridization Test with automated imaging and analysis, generating results automatically) as a direct comparison against standard laboratory ground truth methods. The Verigene System is described as a "bench-top molecular diagnostics workstation" that automates various steps and provides "results display" and "results report generation." This indicates a standalone performance evaluation of the device as a diagnostic tool without continuous human-in-the-loop adjustments to the test outcome.

7. The Type of Ground Truth Used

The ground truth used was a composite gold standard primarily consisting of:

• Culture-based methods confirmed with an FDA-cleared DFA test: For general detection of Influenza A, Influenza B, and RSV.
• Bidirectional sequencing: For subtyping of Influenza A (H3, H1, 2009 H1N1) and RSV (RSV A, RSV B), and for resolution of discordant results.
• NAAT (Nucleic Acid Amplification Test): Used for further clarification in cases of culture/DFA discordance.

This approach uses a combination of established laboratory diagnostic techniques.

8. The Sample Size for the Training Set

The document does not report a separate training set or its sample size for the Verigene® Respiratory Virus Plus Nucleic Acid Test (RV+). The provided information focuses on the validation studies (analytical and methods comparison) for the final trained/developed device. Molecular diagnostic tests like this one are typically developed through iterative optimization and then validated, rather than having a distinct "training set" in the machine learning sense for the final product evaluation.

9. How the Ground Truth for the Training Set Was Established

Since a distinct "training set" is not reported in the context of device development as per the provided text, the method for establishing its ground truth is also not applicable or reported. The focus is on the performance evaluation of the already developed device using the methods comparison and reproducibility studies.

Ask a specific question about this device

The Verigene® Respiratory Virus Nucleic Acid Test on the Verigene SP System (RVNATsp) is a qualitative multiplex in vitro diagnostic test for the detection and identification of Influenza A Virus, Influenza B Virus, and Respiratory Syncytial Virus (RSV) nucleic acids purified from nasopharyngeal swab specimens obtained from patients symptomatic for viral upper respiratory infection. The test is intended to be used on the Verigene SP System as an aid in the differential diagnosis of Influenza A, Influenza B, and RSV infections. The test is not intended to detect Influenza C virus.

Negative results do not preclude influenza virus or RSV infection and should not be used as the sole basis for treatment or other management decisions. It is recommended that negative test results be confirmed by culture.

Performance characteristics for Influenza A Virus were established when Influenza A/H3 and A/H1 were the predominant Influenza A viruses in circulation. As Influenza A viruses emerge, 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.

Not Found

This document is a 510(k) clearance letter for the "Verigene® Respiratory Virus Nucleic Acid Test on the Verigene SP System (RVNATSP)." It describes the device's intended use and substantial equivalence to a predicate device. However, it does not contain the detailed study information, acceptance criteria, or performance data typically found in a clinical study report or a more comprehensive 510(k) submission summary.

Therefore, many of the requested fields cannot be directly answered from this document.

Here's what can be extracted and what cannot:

1. A table of acceptance criteria and the reported device performance

• Cannot be extracted: This document does not provide acceptance criteria or detailed performance data (e.g., sensitivity, specificity, accuracy). It only mentions that performance characteristics for Influenza A Virus were established for specific subtypes.

2. Sample sized used for the test set and the data provenance (e.g. country of origin of the data, retrospective or prospective)

• Cannot be extracted: The document does not specify the sample size, data provenance, or whether the study was retrospective or prospective.

3. Number of experts used to establish the ground truth for the test set and the qualifications of those experts (e.g. radiologist with 10 years of experience)

• Cannot be extracted: The document does not mention experts, their number, or qualifications related to establishing ground truth for the test set.

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

• Cannot be extracted: The document does not describe any adjudication method.

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

• N/A: This device is a diagnostic nucleic acid test, not an AI-assisted imaging device. Therefore, an MRMC study comparing human readers with and without AI assistance is not applicable to this type of device.

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

• Applicable, but details not provided: As an in vitro diagnostic test, the "algorithm only" performance (i.e., the diagnostic test itself) is what is assessed. However, the performance metrics (sensitivity, specificity) are not provided in this letter.

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

• Implicitly standard of care/reference method, but not explicitly stated: For a nucleic acid test, the ground truth would typically be established by a highly sensitive and specific reference method, such as culture or another validated molecular test. The document mentions "Negative results do not preclude influenza virus or RSV infection and should not be used as the sole basis for treatment or other management decisions. It is recommended that negative test results be confirmed by culture," which suggests culture might be part of the ground truth or a confirmatory method. However, the specific method for establishing ground truth for the primary study is not explicitly detailed.

8. The sample size for the training set

• Cannot be extracted: This document does not provide details about a training set since it's a 510(k) clearance letter, not a full study report for an AI/machine learning device.

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

• Cannot be extracted: As above, details about a training set or its ground truth establishment are not present.

Ask a specific question about this device

The Verigene® Respiratory Virus Nucleic Acid Test is a qualitative multiplex in vitro diagnostic test for the detection and identification of Influenza A Virus. Influenza B Virus, and Respiratory Syncytial Virus (RSV) nucleic acids purified from nasopharyngeal swab specimens obtained from patients symptomatic for viral upper respiratory infection. The test is intended to be used on the Verigene® System as an aid in the differential diagnosis of Influenza A, Influenza B, and RSV infections. The test is not intended to detect Influenza C virus.

Negative results do not preclude influenza virus or RSV infection and should not be used as the sole basis for treatment or other management decisions. It is recommended that negative test results be confirmed by culture.

Performance characteristics for Influenza A Virus were established when Influenza A/H3 and A/H1 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.

Not Found

The provided text does not contain detailed information about the acceptance criteria or a comprehensive study report that proves the device meets those criteria. It is a 510(k) clearance letter from the FDA for a device called "Verigene® Respiratory Virus Nucleic Acid Test," along with its indications for use.

While it mentions that the FDA has determined the device is "substantially equivalent" to legally marketed predicate devices, it does not provide the specific performance metrics, sample sizes, ground truth establishment methods, or expert qualifications that would be expected in a detailed study report.

Here's what can be extracted and what is missing:

Information Present in the Document:

• Device Name: Verigene® Respiratory Virus Nucleic Acid Test
• Target Pathogens: Influenza A Virus, Influenza B Virus, and Respiratory Syncytial Virus (RSV)
• Specimen Type: Nasopharyngeal swab
• Intended Use: Qualitative multiplex in vitro diagnostic test as an aid in the differential diagnosis of Influenza A, Influenza B, and RSV infections.
• Limitations:
  • Not intended to detect Influenza C virus.
  • Negative results do not preclude infection and should not be used as the sole basis for treatment or management.
  • Negative results recommended to be confirmed by culture.
  • Performance characteristics for Influenza A Virus were established when A/H3 and A/H1 were predominant; performance may vary with other emerging strains.
  • If a novel Influenza A virus is suspected, specimens should be sent to public health authorities for testing (with appropriate precautions).

Missing Information (Required for the question):

1. A table of acceptance criteria and the reported device performance: This document does not contain this information. It only states that the device is "substantially equivalent."
2. Sample size used for the test set and the data provenance: Not mentioned.
3. Number of experts used to establish the ground truth for the test set and the qualifications of those experts: Not mentioned.
4. Adjudication method (e.g. 2+1, 3+1, none) for the test set: Not mentioned.
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: Not applicable as this is an in-vitro diagnostic (IVD) test, not an AI-assisted imaging device with human readers.
6. If a standalone (i.e. algorithm only without human-in-the-loop performance) was done: As an IVD test, its performance is inherently standalone (the instrument provides the result). However, the specific metrics (sensitivity, specificity, etc.) are not provided.
7. The type of ground truth used (expert consensus, pathology, outcomes data, etc): While it mentions confirmation by culture for negative results, the primary ground truth for the study (if one was conducted for the 510(k)) is not explicitly detailed. Often for such tests, ground truth would be established by culture or a highly sensitive and specific PCR method.
8. The sample size for the training set: Not mentioned (and likely not applicable in the same way as an AI model, as this is a traditional IVD).
9. How the ground truth for the training set was established: Not mentioned.

Conclusion:

The provided document is an FDA 510(k) clearance letter, which confirms substantial equivalence to a predicate device but does not detail the specific performance study, acceptance criteria, and results in the format requested. To obtain this information, one would typically need to review the full 510(k) submission summary or a peer-reviewed publication of the clinical validation study for the device.

Ask a specific question about this device

The Verigene® Respiratory Virus Nucleic Acid Test on the Verigene SP System (RVNATsg) is a qualitative multiplex in vitro diagnostic test for the detection and identification of Influenza A Virus, Influenza B Virus, and Respiratory Syncytial Virus (RSV) nucleic acids purified from nasopharyngeal swab specimens obtained from patients symptomatic for viral upper respiratory infection. The test is intended to be used on the Verigene® SP System as an aid in the differential diagnosis of Influenza A, Influenza B, and RSV infections. The test is not intended to detect Influenza C virus.

Negative results do not preclude influenza virus or RSV infection and should not be used as the sole basis for treatment or other management decisions. It is recommended that negative test results be confirmed by culture.

Performance characteristics for Influenza A Virus were established when Influenza A/H3 and A/H1 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.

The entire RVNATSP is performed on the Verigene® SP System, which is a bench-top molecular diagnostics workstation that consists of two instruments, the Verigene SP Processor and the Verigene Reader. The Verigene SP Processor performs the assay steps on each sample by using a robotic pipettor to transfer and mix reagents within and between separate testing modules designed for nucleic acid extraction, target amplification, and the Verigene Hybridization test. The Verigene hybridization test module is the same as in the previous Verigene System with added modules for nucleic acid extraction and RT-PCR target amplification. Key functions of the Verigene SP Processor include:

1. Reading of the barcode identification label on inserted Test Consumables to maintain positive identification of patient samples throughout processing.
2. Facilitation of nucleic acid extraction, multiplex RT-PCR target amplification, and the Verigene Hybridization Test.
3. Real-time communication of test processing status to the Reader.

The Verigene Reader is the same instrument as in the FDA-cleared VRNAT. It is a free-standing instrument with a touch screen control panel and a wand-based barcode scanner. It utilizes a graphical user interface to guide the user through the process of ordering tests and reporting results. There are no serviceable parts and no user calibration is required. Interaction with the touch screen is minimized through barcode use. This instrument also serves as the reader of the Test Cartridges using advanced optics. The key functions of the Verigene Reader include:

1. Entry and tracking of specimen identification numbers via manual keyboard input or via barcode-reader wand.
2. Test selection for each specimen.
3. Automated transfer of specimen processing instructions on Test Cartridge-specific basis to linked Processor unit(s). A single Reader unit can control up to 32 Processor units.
4. Automated imaging and analysis of Test Cartridges.
5. Results display.
6. Results report generation.

RVNATSP consumables within each single-use disposable test kit include: (i) Tip Holder Assembly; (ii) Extraction Tray; (iii) Amplification Tray; and (iv) RV Test Cartridge. The kit components are inserted into the corresponding module of the Verigene SP Processor prior to each test, and the sample is added to the Extraction Tray. Patient information is entered into the Reader to initiate the test procedure.

1. Tip Holder Assembly – The robotic pipettor picks up pipettes from the Tip Holder Assembly. The pipettes are used for mixing and transferring reagents within the test procedure.
2. Extraction Tray – Nucleic acids are extracted from the sample by using magnetic bead-based methods within the Extraction Tray. Each Tray contains reagents for a single extraction procedure. A robotic pipette transfers reagents to designated wells within the Extraction Tray to affect the steps of lysis, capture of nucleic acids onto the magnetic beads, washing, and eluting the isolated nucleic acids from the magnetic beads.
3. Amplification Tray – The isolated nucleic acids are amplified by using multiplex RT-PCR within the Amplification Tray. Each Tray contains reagents for a single multiplex RT-PCR procedure. A robotic pipette transfers the reagents to a specific well within the Amplification Tray. A set thermal profile is then initiated to perform all of the amplification related steps including UDG-based decontamination, reverse transcription, and multiplex PCR in a single tube. Upon completion, an aliquot of the amplified sample is mixed with hybridization buffer containing the virus specific mediator probes. The sample is then transferred to the Test Cartridge.
4. RV Test Cartridge for Verigene Hybridization Test - The virus-specific amplicons are detected and identified within a Test Cartridge by using specific nucleic acid probes in conjunction with gold nanoparticle probe-based detection technology. Each Test Cartridge is a self-contained, laboratory consumable that consists of two parts. The upper housing of each cartridge is called the "reagent pack" and contains reservoirs filled with the detection reagents. When in place with the 'substrate holder', the reagent pack creates an air-tight hybridization chamber surrounding the region of the substrate containing a target-specific capture array. As each step of the test is completed, old reagents are moved out of the hybridization chamber and new reagents are added from the reagent pack via microfluidic channels and pumps. Once the test is complete, the Test Cartridge is removed from the Verigene SP Processor unit and the reagent pack is snapped off and discarded. The remaining slide is now ready for imaging and analysis in the Verigene Reader.
5. End-point detection on the Verigene Reader: The test slide is inserted into the Verigene Reader wherein it is illuminated along its side. The gold-silver aggregates at the test sites scatter the light, which is in turn captured by a photosensor. The relative intensity arising from each arrayed test site is tabulated. Net signals, defined as the absolute signal intensities with background signals subfracted, are compared with thresholds determined by negative controls within the slide in order to arrive at a decision regarding the presence or absence of target. These results are linked to the test and patient information entered at the beginning of each test session to provide a comprehensive results file.

The provided document describes the Verigene® Respiratory Virus Nucleic Acid Test on the Verigene® SP System (RVNATSP), a qualitative multiplex in vitro diagnostic test for the detection and identification of Influenza A Virus, Influenza B Virus, and Respiratory Syncytial Virus (RSV) nucleic acids. The study aims to demonstrate substantial equivalence to a previously cleared predicate device, the Verigene® Respiratory Virus Nucleic Acid Test (VRNAT, K083088).

1. Table of Acceptance Criteria and Reported Device Performance:

The primary acceptance criteria for the RVNATSP were established by demonstrating equivalence to the predicate VRNAT device (K083088) in terms of analytical sensitivity (Limit of Detection), lack of carryover/crossover contamination, precision/reproducibility, and method comparison (percent agreement). Based on the provided information, the acceptance criteria are implicitly that the performance of the RVNATSP should be comparable or identical to that of the cleared VRNAT.

2. Sample Size Used for the Test Set and Data Provenance:

• Analytical Sensitivity (LOD) Study: For each virus strain, 20 replicates were tested at the LOD concentration after an initial quadruplicate testing of serially diluted stocks. This was done for Influenza B, RSV A, RSV B, and Influenza A. The samples were prepared from strains with established titers serially diluted into Universal Transport Media (Copan). The country of origin of the data is not explicitly stated but implies internal laboratory studies. The data is prospective for the purpose of this study.
• Carryover and Crossover Contamination Studies: High positive samples of Influenza A, Influenza B, and RSV B were alternated with high negative samples. The exact number of samples or alternations is not specified but the "collective data" demonstrated no cross-contamination. This appears to be a prospective internal study.
• Precision/Reproducibility Studies: Eight unique sample panels (viral strains at high negative, low positive, and moderate positive concentrations) were tested.
  • Site 1 (Precision Study): The sample set was tested over 12 non-consecutive days with two operators performing the test in duplicate (total of 48 tests per panel member for Influenza A, Influenza B, RSV A, RSV B).
  • Sites 2 and 3 (Reproducibility Study): The sample set was tested in triplicate daily by 2 operators on each of five non-consecutive days (total of 15 tests per panel member per site for Influenza A, Influenza B, RSV A, RSV B).
  • The total number of individual results used for precision/reproducibility comparison across all sites for each panel member was 78 (48 from Site 1 + 15 from Site 2 + 15 from Site 3). This was a prospective, multi-site study. Data provenance is not specified beyond "three sites."
• Method Comparison Studies: A total of 62 unique samples (from culture positive and negative nasopharyngeal swab samples) were prepared, representing Influenza A (15 positive, 47 negative), Influenza B (16 positive, 46 negative), and RSV A/B (34 positive, 28 negative). These were diluted to yield viral load levels close to low positive. Each sample provided a decision for all three viruses, resulting in 186 decisions per test.
  • The sample set was tested at one internal site (Site 1) using the cleared VRNAT.
  • The same sample set was tested at all three sites (Site 1, Site 2, and Site 3) using the RVNATSP.
  • "Total of 62x4 = 248 unique tests" (implying 62 samples tested once on VRNAT at Site 1, and once on RVNATSP at each of the 3 sites). The comparison data in Tables 7-9 use the VRNAT (Old System) results (presumably from Site 1) as the reference for each RVNATSP site's results. The combined data in Table 10 then aggregates results from all three RVNATSP sites against the VRNAT reference.
  • Data provenance is not explicitly stated, but "culture positive nasopharyngeal swab samples" suggests patient samples. This was a prospective, multi-site study.

3. Number of Experts Used to Establish the Ground Truth for the Test Set and Their Qualifications:

This device is a molecular diagnostic test. The "ground truth" for the test set is established by the known concentration of viral strains (for analytical sensitivity/LOD and precision/reproducibility) or by the results from the predicate device (cleared VRNAT) for the method comparison study. There are no human "experts" required to establish a ground truth in the way a radiologist would for an imaging study. The reference method for comparison is a previously cleared, similar molecular diagnostic test.

4. Adjudication Method for the Test Set:

Not applicable in the context of this molecular diagnostic device comparison. The "ground truth" for method comparison is the result obtained from the predicate device (cleared VRNAT). For discordant results in the method comparison study (e.g., in Site 2 and Site 3 performance for Influenza A), "Repeat tests were positive and gave the expected result," suggesting internal re-testing or confirmation processes were employed for discrepancies, but not necessarily a formal adjudication by a panel of human experts.

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:

Not applicable. This is a molecular diagnostic device, not an imaging device requiring human readers or AI assistance in interpretation. The output is a qualitative "Detected" or "Not Detected" result for specific viral nucleic acids.

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

The device (RVNATSP) operates as a standalone system once the sample is loaded. The "algorithm" in this context refers to the molecular diagnostic assay steps and the instrument's internal decision algorithm for generating "Detected" or "Not Detected" results. The performance studies described (analytical sensitivity, contamination, precision, method comparison) inherently evaluate the standalone performance of the RVNATSP system without human intervention in the result interpretation after the run is initiated. The comparison against the predicate device also assesses standalone performance.

7. The Type of Ground Truth Used (expert consensus, pathology, outcomes data, etc.):

• For Analytical Sensitivity (LOD) and Precision/Reproducibility: The ground truth was based on known concentrations of specific viral strains (e.g., TCID50/mL values) that were serially diluted into a sample matrix.
• For Method Comparison: The ground truth was established by the results obtained from the FDA-cleared predicate device (VRNAT, K083088). This is a "device-to-device" comparison where the cleared device serves as the reference standard.

8. The Sample Size for the Training Set:

This document describes a 510(k) submission for a molecular diagnostic device, not a machine learning or AI-based system that typically uses a "training set." The RVNATSP is built upon established molecular biology principles and reagents. Therefore, the concept of a "training set" for an algorithm, as described in AI development, is not directly applicable to this device. The development of such assays involves extensive R&D and analytical optimization, which could be considered an iterative "training" process for the assay design itself, but not a distinct dataset for algorithm training in the computational sense.

9. How the Ground Truth for the Training Set Was Established:

As noted above, the concept of a "training set" with established ground truth as understood in AI/ML development does not directly apply to this molecular diagnostic device. The assay design and optimization would rely on established molecular biology techniques, purified viral samples, and potentially a range of clinical samples, but these are part of the assay development and validation rather than a formally defined "training set" for an algorithm.

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The Verigene® Respiratory Virus Nucleic Acid Test is a qualitative multiplex in vitro diagnostic test for the detection and identification of Influenza A Virus, Influenza B Virus, and Respiratory Syncytial Virus (RSV) nucleic acids purified from nasopharyngeal swab specimens obtained from patients symptomatic for viral upper respiratory infection. The test is intended to be used on the Verigene® System as an aid in the differential diagnosis of Influenza A, Influenza B, and RSV infections. The test is not intended to detect Influenza C virus.

Negative results do not preclude influenza virus or RSV infection and should not be used as the sole basis for treatment or other management decisions. It is recommended that negative test results be confirmed by culture.

Performance characteristics for Influenza A Virus were established when Influenza A/H3 and A/H1 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.

The Verigene Respiratory Virus Nucleic Acid Test (VRNAT) is based on uniquely identifying virus-specific nucleic acids for Influenza A virus, Influenza B virus, and Respiratory Syncytial Virus (RSV). The VRNAT involves the following steps: (i) Sample Preparation: Isolation of the viral RNA from nasopharyngeal swab specimens obtained from symptomatic patients. Sample preparation is conducted on an automated sample isolation system (NucliSens® easyMAG™ System, bioMérieux); (ii) Multiplex RT-PCR procedure for the generation of virus-specific amplicons: A specified volume of the eluted genomic RNA from the sample preparation step is subjected to an RT-PCR target amplification step; (iii) Verigene Test: Conducted on the Verigene® System for detection and identification of virus-specific amplicons.

The Verigene System consists of two instruments, the Verigene Processor and the Verigene Reader, and utilizes single-use, disposable Test Cartridges.

The Verigene Reader is a bench-top, free-standing instrument with a touch screen control panel and a wand-based barcode scanner. It utilizes a graphical user interface to guide the user through the process of ordering tests and reporting results. There are no serviceable parts and no user calibration is required. Interaction with the touch screen is minimized through barcode use. This instrument also serves as the reader of the hybridization substrate using optical detection.

The key functions of the Verigene Reader include:

• Entry and tracking of specimen identification numbers via manual keyboard input or via barcode-reader wand.
• Test selection for each specimen.
• Automated transfer of specimen processing instructions on Test Cartridge specific basis to linked Processor unit(s).
• Automated imaging and analysis of Test Cartridges.
• Results display.
• Results report generation.

The Verigene Processor performs the Verigene Test under the direction of the Verigene Reader. It has been designed to be simple and easy to use with minimal user interaction. It contains no fluids and has no user calibration requirements. There are four hybridization modules in each Verigene Processor and up to eight Verigene Processors may be connected to a single Verigene Reader. The modules within a Verigene Processor can simultaneously run different tests. When a Test Cartridge containing the sample mix is inserted, a barcode reader internal to the Verigene Processor modules reads the cartridge ID and sends it to the Verigene Reader. From this information the Verigene Reader establishes the hybridization parameters and automatically starts the Verigene Test.

The Test Cartridge consists of two parts: a Reagent Pack with reservoirs preloaded with test-specific reagents, and a Substrate Holder. The reagent pack creates an air-tight hybridization chamber surrounding the region of the substrate-containing target-specific capture array. As each step in the Verigene Test is completed, old reagents are moved out of the hybridization chamber and new reagents are added from the reagent pack via microfluidics.

To run the Verigene Test, the user mixes the sample with a test-specific Sample Buffer and loads this sample mix into the Test Cartridge. The Test Cartridge is subsequently inserted into the Processor. Both the identity of the test contained in the Test Cartridge and the associated patient specimen are linked with a common barcode that is unique to each Test Cartridge. Test information entered at the beginning of each test session is used to direct the associated module of the Processor unit on what steps to execute to process the Test Cartridge, and the patient specimen information is used for tracking and results reporting. Once the test is complete, the Test Cartridge is removed from the Processor unit and the reagent pack is snapped off and discarded. The remaining Test Substrate is now ready for imaging and analysis in the Reader unit.

The Test Substrate is inserted into the Reader wherein it is illuminated along its side. The gold-silver aggregates at the test sites scatter the light, which is in turn captured by a photosensor. The relative intensity arising from each arrayed test site is tabulated. Net signals, defined as the absolute signal intensities with background signals subtracted, are compared with thresholds determined by negative and positive controls within the slide in order to arrive at a decision regarding the presence or absence of target. These results are linked to the test and patient information entered at the beginning of each test session to provide a comprehensive results file.

The Nanosphere Verigene® Respiratory Virus Nucleic Acid Test (VRNAT) is a qualitative multiplex in vitro diagnostic test for the detection and identification of Influenza A Virus, Influenza B Virus, and Respiratory Syncytial Virus (RSV) nucleic acids.

Here’s a breakdown of the acceptance criteria and the study that supports it:

1. Table of Acceptance Criteria and Reported Device Performance

The acceptance criteria are not explicitly stated as numerical targets for sensitivity and specificity in the provided document. However, the reported device performance is presented. The "predicate device" section and the clinical performance data imply that the acceptance criteria are generally to achieve high sensitivity and specificity in comparison to a culture-based reference method, and to demonstrate acceptable reproducibility and analytical sensitivity. Based on the clinical performance, the implicit acceptance criteria would involve a high level of agreement with the reference method (DFA/Viral Culture), particularly for sensitivity, and acceptable specificity.

2. Sample Size Used for the Test Set and Data Provenance

• Sample Size for Clinical Performance (Test Set): A total of 720 nasopharyngeal swab specimens were used for the clinical performance study.
• Data Provenance: The specimens were prospectively collected during the 2007-2008 respiratory season at three clinical sites. The country of origin is not explicitly stated but can be inferred to be the USA given the FDA submission. The residual specimens were frozen and later tested.

3. Number of Experts Used to Establish the Ground Truth for the Test Set and Qualifications of those Experts

The ground truth for the clinical performance study was established using a culture-based reference method followed by direct fluorescent antibody (DFA) identification of all culture positive samples.

• Number of Experts: The document does not specify the exact number of experts involved in the DFA identification or culture interpretation. It refers to it as a "culture-based reference method" and that discordant results were followed up by using "bidirectional sequencing at an independent reference laboratory."
• Qualifications of Experts: The qualifications of the personnel performing the DFA and culture interpretation are not explicitly stated. For the discordant results, the follow-up involved an "independent reference laboratory," implying expert-level analysis.

4. Adjudication Method for the Test Set

• Adjudication Method: For the clinical performance study, discordant results between the VRNAT and the reference method (DFA/Viral Culture) were followed up by using bidirectional sequencing at an independent reference laboratory. This acts as a form of "tie-breaker" or confirmatory adjudication.
• Initial "No Call" Results: Samples with an initial 'No Call' result (2.9% of samples) were re-tested following the recommendations in the 'Results Interpretation' section. All these retested samples were resolved.

5. Multi-Reader Multi-Case (MRMC) Comparative Effectiveness Study

• Not Applicable: The provided document describes the performance of a diagnostic test for detecting viral nucleic acids, not an imaging device or an AI system intended to assist human readers. Therefore, an MRMC comparative effectiveness study, which typically assesses the impact of AI on human reader performance, was not performed or described.

6. Standalone (Algorithm Only Without Human-in-the-Loop Performance) Study

• Yes: The entire clinical performance study and analytical studies (reproducibility, analytical sensitivity, analytical reactivity, analytical specificity/cross-reactivity, competitive inhibition, fresh vs. frozen, interferences) assess the performance of the Verigene® Respiratory Virus Nucleic Acid Test (VRNAT) system itself, which includes the automated Verigene Processor and Verigene Reader. This represents the standalone performance of the algorithm and system in identifying the target viruses. Users perform sample preparation and load cartridges, but the detection and interpretation leading to a "Detected" or "Not Detected" call are automated by the system based on pre-defined clinical cut-off criteria.

7. Type of Ground Truth Used

• Clinical Performance (Test Set): The primary ground truth for the clinical performance study was culture-based reference method followed by direct fluorescent antibody (DFA) identification of all culture positive samples. For discordant results, bidirectional sequencing was used as a confirmatory method.
• Analytical Studies: For analytical sensitivity, reactivity, and specificity, the ground truth was established using known concentrations of well-characterized viral strains grown in culture or other established microorganisms.

8. Sample Size for the Training Set

• The document does not explicitly mention a separate "training set" or its sample size in the context of machine learning. The Verigene system uses a defined algorithm with empirically determined clinical cut-offs (Noise Threshold, Ratio-to-NC, Ratio-to-PC) rather than a continuously learning AI model that would typically require a distinct training set. The determination of these cut-offs (e.g., using ROC curves) might involve a dataset, but it is not specified as a "training set" in the conventional machine learning sense, nor is its size provided.

9. How the Ground Truth for the Training Set Was Established

• As a distinct "training set" for an AI model is not described, the method for establishing its ground truth is not applicable. The clinical cut-offs for the VRNAT call algorithm were determined empirically (e.g., Noise Threshold = Negative Control + 2 SD) and by using ROC curves for the normalized ratios. These methods rely on data with established true positive and true negative results, but the dataset used for this "training" of the algorithm's thresholds is not detailed as a formal training set with specific ground truth establishment methods.

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