(117 days)
The AmpliVue® Trichomonas Assay is an in vitro diagnostic test, uses isothermal amplification technology (helicase-dependent amplification, HDA) for the qualitative detection of Trichomonas vaginalis nucleic acids isolated from clinician-collected vaginal swab specimens obtained from symptomatic or asymptomatic females to aid in the diagnosis of trichomoniasis.
The AmpliVue® Trichomonas Assay combines simple processing, an isothermal amplification technology named Helicase-Dependent Amplification (HDA), and a selfcontained disposable amplicon detection device for the detection of T. vaginalis in clinician-collected vaginal swabs from symptomatic and asymptomatic women. The assay targets a conserved multi-copy sequence of the T. vaginalis DNA. The vaginal swab is eluted in a lysis tube, and the cells are lysed by simple heat treatment. After heat treatment, an aliquot of the lysed specimen is transferred into a dilution tube. An aliquot of the diluted sample is added to a reaction tube containing a lyophilized mix of HDA reagents including primers specific for the amplification of a conserved DNA sequence only found in T. vaginalis. The assay also includes an internal control to confirm the integrity of the assay reagents and cassette detection as well as to control for (or determine whether) HDA-inhibitors that may be present within the clinical specimens. The HDA reaction is asymmetric so that an excess of single stranded DNA (amplicon) is formed. The sequence specific capture probes as well as a biotinylated detection probe shared by both target and internal control bind to the corresponding single-stranded amplicons, forming dual labeled probe-amplicon hybrid. After completion of the HDA reaction, the reaction tube is transferred to a cassette for rapid detection with the test result displayed as test and/or control lines in the window of the cassette. The dual-labeled probe-amplicon hybrid is then detected by the lateral flow strip within the cassette. The bottom line captures the test amplicon and the top line captures the control amplicon. The biotin label binds the streptavidin-conjugated color particles for visualization and the test result is shown as colored lines visible to the naked eye. The cassette is comprised of two individual components: an amplicon cartridge that holds the running buffer and a single 0.2 mL thin wall reaction tube containing the amplified product; and the detection chamber which houses the amplicon cartridge and a verticalflow DNA detection strip embedded into the cassette. The DNA detection strip is coated with different anti-hapten antibodies that serve as the T. vaginalis test (T) line and the control (C) line in the assay. A razor blade and a plastic pin located at the bottom of the detection chamber opens the HDA reaction tube and the running buffer bulb when the handle of the cassette is closed. The mixture flows through a fiberglass paper connected to the DNA detection strip that contains a fiberglass pad pre-loaded with streptavidinconjugated color particles for color visualization. Detection of T. vaginalis DNA is reported whenever the T2 (Test line 2) is visible through the detection window of the cassette. The presence of the C line is not required for positive results. No detection of T. vaginalis DNA is reported when only the C line is displayed. The assay is regarded as invalid when neither line is displayed.
Here's an analysis of the acceptance criteria and the study that proves the device meets them, based on the provided text:
Description of Acceptance Criteria and Device Performance Study for AmpliVue® Trichomonas Assay
1. Table of Acceptance Criteria and Reported Device Performance:
The document does not explicitly state formal "acceptance criteria" in terms of specific performance targets set prior to the study. However, the study aims to demonstrate substantial equivalence to a predicate device, and the performance characteristics reported serve as the de facto demonstration of meeting the required clinical performance for this type of in vitro diagnostic device. The performance characteristics of the predicate device (APTIMA Trichomonas vaginalis Assay) are also provided for comparison.
| Performance Metric | Acceptance Criteria (from predicate performance) | Reported Device Performance (AmpliVue® Trichomonas Assay) |
|---|---|---|
| CLINICAL PERFORMANCE | ||
| Asymptomatic | ||
| Sensitivity | 100% (95% CI 75.8 – 100) | 100% (95% CI 94.1 – 100) |
| Specificity | 97.3% (95% CI 94.6 – 98.7) | 98.3% (95% CI 96.9 – 99.1) |
| Symptomatic | ||
| Sensitivity | 100% (95% CI 93.7 – 100) | 100% (95% CI 93.9 – 100) |
| Specificity | 98.8% (95% CI 97.0 – 99.5) | 97.9% (95% CI 95.5 – 99.0) |
| ANALYTICAL PERFORMANCE | ||
| Limit of Detection (LoD) | Not explicitly stated for predicate in this context, but typically a low concentration for high sensitivity. | G3 strain: 307 trophozoites/mL; CDC888 strain: 921 trophozoites/mL (defined as lowest concentration at which 95% of replicates tested positive) |
| Reproducibility | Consistent results across sites and operators. | Achieved 100% agreement for low and moderate positive and negative samples across 3 sites (95.9% to 100% CI). High negative showed 40% agreement (30.5% to 50.3% CI) within its expected range (20-80%). |
| Cross-Reactivity | No cross-reactivity with common microorganisms. | No cross-reactivity with 45 tested microorganisms. |
| Interference | No interference from common substances or microorganisms. | No interference from 45 tested microorganisms or 13 tested substances. |
| Analytical Reactivity (Inclusivity) | Detection of a broad range of T. vaginalis strains. | All 20 additional T. vaginalis strains tested were detected. |
2. Sample Size and Data Provenance for the Test Set:
- Sample Size for Test Set: 992 clinician-collected vaginal swab specimens.
- Data Provenance: The study was a multi-center study performed at four locations in the United States and one location in Canada. It was a prospective study, with specimens obtained from subjects after informed consent between April and November 2014.
3. Number of Experts and Qualifications for Ground Truth in the Test Set:
- The document states that the reference method for establishing ground truth was a composite of Wet Mount and InPouch TV culture. It does not specify the number or qualifications of experts involved in interpreting these reference methods. However, these methods are standard clinical laboratory procedures, implying trained laboratory personnel (e.g., medical technologists, clinical microbiologists) would have performed and interpreted them.
4. Adjudication Method for the Test Set:
- The ground truth was established using a composite reference method: Wet Mount and InPouch TV culture. A specimen was considered positive if either test was positive. This acts as a form of "OR" adjudication or a composite gold standard, rather than a direct agreement between multiple independent experts on the same test.
- For discordant results where the composite reference method was negative but the AmpliVue assay was positive, the FDA-cleared molecular device collection swab was used for discordant testing. Specifically, "Eight (8) of sixteen (16) Composite negative/AmpliVue positive specimens were positive by a FDA-cleared Trichomonas vaginalis molecular device." This implies a form of tie-breaking or re-evaluation using a third, highly sensitive method a posteriori.
5. Multi-Reader Multi-Case (MRMC) Comparative Effectiveness Study:
- No, an MRMC comparative effectiveness study was not done. This study focuses on the standalone performance of the AmpliVue® Trichomonas Assay compared to a composite reference method, and its substantial equivalence to a predicate device. It does not evaluate human reader performance with or without AI assistance.
6. Standalone Performance Study:
- Yes, a standalone performance study was done. The entire clinical study described evaluates the performance of the AmpliVue® Trichomonas Assay (algorithm only, as it's an in vitro diagnostic test) in detecting Trichomonas vaginalis nucleic acids from vaginal swab specimens. The reported sensitivity, specificity, PPV, and NPV are all measures of this standalone performance.
7. Type of Ground Truth Used:
- The primary ground truth used was a composite reference method consisting of:
- Wet Mount
- InPouch TV Culture
- For discordant cases (AmpliVue positive, composite negative), an FDA-cleared Trichomonas vaginalis molecular device was used for further investigation, suggesting a hierarchical or confirmatory approach for specific discrepancies.
8. Sample Size for the Training Set:
- The document does not specify a sample size for a training set. The AmpliVue® Trichomonas Assay is an in vitro diagnostic test based on Helicase-Dependent Amplification (HDA) technology. This type of assay does not typically involve traditional "training sets" in the same way machine learning algorithms do. Its development involves analytical validation (LoD, cross-reactivity, interference, inclusivity) and then clinical validation with a distinct set of patient samples, rather than a machine learning training/test split.
9. How the Ground Truth for the Training Set Was Established:
- As the device is an in-vitro diagnostic assay rather than an AI/ML algorithm requiring a training set, the concept of "ground truth for the training set" does not directly apply here. The assay's design and analytical parameters (e.g., specific primers, probes, Lysis Buffer, Dilution Buffer, Reaction Tubes quantities) were established through laboratory development and analytical studies (LoD, cross-reactivity, inclusivity, interference) which confirm the assay's ability to detect the target DNA sequence under various conditions.
§ 866.3860
Trichomonas vaginalis nucleic acid assay.(a)
Identification. ATrichomonas vaginalis nucleic acid assay is a device that consists of primers, probes, enzymes, and controls for the amplification and detection of trichomonas nucleic acids in endocervical swabs, vaginal swabs, and female urine specimens, from women symptomatic for vaginitis, cervicitis, or urethritis and/or to aid in the diagnosis of trichomoniasis in asymptomatic women. The detection of trichomonas nucleic acids, in conjunction with other laboratory tests, aids in the clinical laboratory diagnosis of trichomoniasis caused byTrichomonas vaginalis .(b)
Classification. Class II (special controls). The special controls are set forth in FDA's guideline document entitled: “Class II Special Controls Guideline: Nucleic Acid Amplification Assays for the Detection ofTrichomonas vaginalis; Guideline for Industry and Food and Drug Administration Staff.” See § 866.1(e) for information on obtaining this document.