(197 days)
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.
| Performance Metric | Acceptance Criteria (Implied) | Reported Device Performance |
|---|---|---|
| Influenza A: | ||
| Sensitivity | High (e.g., >95% based on predicate device expectations) | 99.2% (95% CI = 95.5% - 99.9%) |
| Specificity | High (e.g., >90% based on predicate device expectations) | 90.1% (95% CI = 87.5% - 92.3%) |
| Influenza B: | ||
| Sensitivity | High (e.g., >95% based on predicate device expectations) | 96.8% (95% CI = 83.5% - 99.4%) |
| Specificity | High (e.g., >95% based on predicate device expectations) | 98.5% (95% CI = 97.3% - 99.2%) |
| RSV: | ||
| Sensitivity | High (e.g., >85% based on predicate device expectations) | 89.8% (95% CI = 78.2% - 95.6%) |
| Specificity | High (e.g., >90% based on predicate device expectations) | 91.5% (95% CI = 89.2% - 93.4%) |
| Reproducibility (High Negative): | High (e.g., >90% agreement) | Influenza A: 96.2%; Influenza B: 100%; RSV A: 97.4%; RSV B: 94.9% |
| Reproducibility (Low Positive): | High (e.g., >95% agreement) | Influenza A: 98.7%; Influenza B: 98.7%; RSV A: 98.7%; RSV B: 100% |
| Reproducibility (Moderate Positive): | Very High (e.g., 100% agreement) | Influenza A: 100%; Influenza B: 100%; RSV A: 100%; RSV B: 100% |
| Analytical Reactivity: | 100% concordance with expected results | 100% concordance with expected results |
| Analytical Specificity/Cross-Reactivity: | No cross-reactivity observed | No cross-reactivity observed with 38 common respiratory pathogens and microorganisms |
| Competitive Inhibition: | No evidence of competitive inhibition | No evidence of competitive inhibition |
| Fresh vs. Frozen Samples: | High positive and negative agreement | Influenza A: 100% PPA, 100% NPA; Influenza B: 100% PPA, 100% NPA; RSV: 100% PPA, 96.3% NPA |
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.
§ 866.3980 Respiratory viral panel multiplex nucleic acid assay.
(a)
Identification. A respiratory viral panel multiplex nucleic acid assay is a qualitative in vitro diagnostic device intended to simultaneously detect and identify multiple viral nucleic acids extracted from human respiratory specimens or viral culture. The detection and identification of a specific viral nucleic acid from individuals exhibiting signs and symptoms of respiratory infection aids in the diagnosis of respiratory viral infection when used in conjunction with other clinical and laboratory findings. The device is intended for detection and identification of a combination of the following viruses:(1) Influenza A and Influenza B;
(2) Influenza A subtype H1 and Influenza A subtype H3;
(3) Respiratory Syncytial Virus subtype A and Respiratory Syncytial Virus subtype B;
(4) Parainfluenza 1, Parainfluenza 2, and Parainfluenza 3 virus;
(5) Human Metapneumovirus;
(6) Rhinovirus; and
(7) Adenovirus.
(b)
Classification. Class II (special controls). The special controls are:(1) FDA's guidance document entitled “Class II Special Controls Guidance Document: Respiratory Viral Panel Multiplex Nucleic Acid Assay;”
(2) For a device that detects and identifies Human Metapneumovirus, FDA's guidance document entitled “Class II Special Controls Guidance Document: Testing for Human Metapneumovirus (hMPV) Using Nucleic Acid Assays;” and
(3) For a device that detects and differentiates Influenza A subtype H1 and subtype H3, FDA's guidance document entitled “Class II Special Controls Guidance Document: Testing for Detection and Differentiation of Influenza A Virus Subtypes Using Multiplex Nucleic Acid Assays.” See § 866.1(e) for the availability of these guidance documents.