(87 days)
The Panther Fusion Paraflu assay is a multiplex real-time PCR (RT-PCR) in vitro diagnostic test for the rapid and qualitative detection and differentiation of parainfluenza 1 virus, parainfluenza 2 virus, parainfluenza 3 virus and parainfluenza 4 virus (HPIV-1, HPIV-2, HPIV-3, and HPIV-4). Nucleic acids are isolated and purified from nasopharyngeal (NP) swab specimens obtained from individuals exhibiting signs and symptoms of a respiratory tract infection.
This assay is intended to aid in the differential diagnosis of HPIV-1, HPIV-2, HPIV-3, and HPIV-4 infections in humans. Negative results do not preclude HPIV-1, HPIV-2, HPIV-3, and HPIV-4 infections and should not be used as the sole basis for treatment or other management decisions. This assay is designed for use on the Panther Fusion system.
The Panther Fusion Paraflu assay is a multiplex real-time PCR (RT-PCR) in vitro diagnostic test developed for use on the fully automated Panther Fusion system to detect and differentiate parainfluenza 1 virus, parainfluenza 2 virus, parainfluenza 3 virus, and parainfluenza 4 virus directly from nasopharyngeal swab specimens.
The Panther Fusion Paraflu assay involves the following steps: Sample lysis, nucleic acid capture and elution, and multiplex RT-PCR where analytes (when present) are simultaneously amplified, detected and differentiated. Nucleic acid capture and elution takes place in a single tube on the Panther Fusion system. The eluate is transferred to the Panther Fusion system reaction tube containing the assay reagents. Multiplex RT-PCR is then performed for the eluted nucleic acid on the Panther Fusion system.
Acceptance Criteria and Study for Panther Fusion Paraflu Assay
1. Table of Acceptance Criteria and Reported Device Performance
The acceptance criteria for the Panther Fusion Paraflu assay are largely based on the performance characteristics of its predicate device, the Prodesse ProParaFlu+ Assay, as well as satisfactory analytical and clinical performance established in the studies. While explicit "acceptance criteria" for sensitivity and specificity are not directly stated as numerical thresholds in this document, the clinical performance study aims to demonstrate that the device performs comparably to the reference methods (viral culture/DFA and PCR/sequencing) and that the observed performance (sensitivity, specificity, PPA, NPA) is clinically acceptable. The analytical studies establish the device's technical capabilities.
Here's a summary of the reported device performance from the clinical study (Table 12), which implicitly meets the acceptance threshold for substantial equivalence:
| Analyte | Performance Metric | Reported Device Performance (95% CI) |
|---|---|---|
| HPIV-1 | Sensitivity | 97.1 (85.1-99.5) |
| HPIV-1 | Specificity | 99.6 (99.4-99.8) |
| HPIV-2 | Sensitivity | 91.7 (74.2-97.7) |
| HPIV-2 | Specificity | 99.5 (99.1-99.7) |
| HPIV-3 | Sensitivity | 96.3 (87.5-99.0) |
| HPIV-3 | Specificity | 99.0 (98.6-99.3) |
| HPIV-4 | Positive Percent Agreement (PPA) | 96.7 (83.3-99.4) |
| HPIV-4 | Negative Percent Agreement (NPA) | 99.8 (99.6->99.9) |
2. Sample Size and Data Provenance for the Test Set
- Sample Size Used for the Test Set: 2870 evaluable samples were used for the clinical performance analyses for each analyte. (Initially 2961 specimens were obtained, but 31 were withdrawn, and some had invalid reference results, leading to 2870 evaluable samples).
- Data Provenance: The samples were "leftover, remnant nasopharyngeal (NP) swab specimens" collected from "male and female individuals of all ages exhibiting signs and/or symptoms of a respiratory tract infection." The samples were obtained from "Four participating US pediatric/adolescent, private, and/or university hospitals." The study was prospective in terms of data collection for the clinical performance assessment.
3. Number of Experts and Qualifications for Ground Truth
The document does not explicitly state the number or specific qualifications of experts used to establish the ground truth. However, it indicates the methods used:
- For HPIV-1 and HPIV-3: Reference viral culture followed by direct fluorescent antibody (DFA) identification. This implies laboratory personnel experienced in virology and cell culture techniques.
- For HPIV-4: 2 unique and independently developed reverse transcriptase PCR assays followed by bi-directional sequencing. This would involve molecular biology experts and potentially bioinformaticians for sequence analysis.
4. Adjudication Method for the Test Set
The adjudication method differed by analyte:
- For HPIV-1 and HPIV-3: A validated PCR assay was used for discordant resolution testing.
- For HPIV-4: No discordant resolution testing was performed. The reference assay for HPIV-4 was based on a composite result of two different reverse transcriptase PCR assays followed by bi-directional sequencing. This implies that the consensus of these two independent PCR/sequencing results served as the definitive ground truth for HPIV-4.
5. Multi-Reader Multi-Case (MRMC) Comparative Effectiveness Study
No, a multi-reader multi-case (MRMC) comparative effectiveness study was not done. This device is an in vitro diagnostic test (a laboratory assay) and does not involve human readers in the same way an imaging or AI diagnostic algorithm would. The performance is assessed against established laboratory reference methods.
6. Standalone (Algorithm Only) Performance
Yes, a standalone performance study was done. The "Panther Fusion Paraflu assay" is a fully automated system that performs nucleic acid extraction, amplification, and detection. Its performance as an algorithm/assay without human intervention during the diagnostic process is precisely what the analytical and clinical performance studies describe. The results presented in the tables (e.g., Table 12 for clinical performance, Tables 8, 9, 13, 14, 15 for analytical precision and reproducibility) represent the standalone performance of the device.
7. Type of Ground Truth Used
The ground truth varied by analyte:
- For HPIV-1 and HPIV-3: Viral culture followed by direct fluorescent antibody (DFA) identification. In cases of discordance, a validated PCR assay was used for resolution. This represents a combination of traditional laboratory methods (culture/DFA) and molecular diagnostic (PCR) as expert consensus for confirmed cases.
- For HPIV-4: Two unique and independently developed reverse transcriptase PCR assays followed by bi-directional sequencing. This represents molecular diagnostic confirmation via sequencing.
8. Sample Size for the Training Set
The document does not explicitly state a "training set" size for the Panther Fusion Paraflu assay in the context of an algorithm that learns from data. This is likely because the Panther Fusion Paraflu assay is a molecular diagnostic test based on pre-designed primers and probes for specific viral targets, rather than a machine learning algorithm that is "trained" on a dataset in the conventional sense. The development of such assays involves extensive analytical validation to ensure specificity and sensitivity through iterative design and testing for the targeted genetic sequences.
The analytical studies (e.g., LoD, interference, specificity) involved specific panels and spiked samples to characterize the assay's performance, but these are not referred to as a "training set."
9. How the Ground Truth for the Training Set Was Established
As noted above, there isn't a "training set" in the machine learning sense. The ground truth for the analytical validation (e.g., LoD, interference, specificity) was established by using:
- Well-characterized viral strains (e.g., HPIV-1, HPIV-2, HPIV-3, HPIV-4) at known concentrations (e.g., TCID50/mL).
- Spiking these known viral strains into negative clinical specimens or simulated clinical matrix.
- Testing panels of known interfering organisms (viral, bacterial, yeast) and substances to confirm lack of cross-reactivity or interference.
The positivity or negativity in these analytical studies is determined by the known content of the prepared spiked samples.
§ 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.