(90 days)
Focus Technologies' HerpeSelect® 2 ELISA IgG test is intended for qualitatively detecting the presence or absence of human IgG class antibodies to HSV-2 in human sera. In conjunction with the Focus HerpeSelect® I ELISA IgG, the test is indicated for testing sexually active adults or expectant mothers for aiding in the presumptive digenosis of HSV infection. The assay can be used manually or in conjunction with an automated system as outlined in the package insert. The user is responsible for assay performance characteristics when an automated system is used. The performance of this assay has not been established for use in a pediatric population, for neonatal screening, or for testing of immunocompromised patients.
In the HerpeSelect 2 ELISA IgG assay, the polystyrene microwells are coated with recombinant gG-2 antigen. Diluted serum samples and controls are incubated in the wells to allow specific antibody present in the samples to react with the antigen. Nonspecific reactants are removed by washing, and peroxidase-conjugated anti-human IgG is added and reacts with specific IgG. Excess conjugate is removed by washing. Enzyme substrate and chromogen are added, and the color is allowed to develop. After adding the Stop Reagent, the resultant color change is quantified by a spectrophotometric reading of optical density (OD). Sample optical density readings are compared with reference cut-off OD readings to determine results.
The provided document describes the performance characteristics of the Focus Technologies HerpeSelect® 2 ELISA IgG assay for detecting HSV-2 IgG antibodies. The acceptance criteria are not explicitly stated in a single section but are implied through the reported performance metrics against a Western blot (WB) reference method across various populations.
1. Table of Acceptance Criteria and Reported Device Performance
Since explicit "acceptance criteria" are not given in a dedicated table, the reported performance metrics against the reference Western blot (considered the gold standard for serostatus) are used to infer them. Performance is presented for different populations, implying that satisfactory results in these different contexts are within the "acceptance criteria" for the device's intended use.
| Characteristic | Acceptance Criteria (Implied by Reported Performance) | Reported Device Performance (HerpeSelect 2 ELISA IgG) |
|---|---|---|
| For Expectant Mothers (n=241) | ||
| Relative Sensitivity (vs. WB) | High (e.g., above 90%) | 100% (58/58) (95% CI: 93.8-100%) |
| Relative Specificity (vs. WB) | High (e.g., above 90%) | 96.1% (172/179) (95% CI: 92.1-98.4%) |
| For Sexually Active Adults (n=246) | ||
| Relative Sensitivity (vs. WB) | High (e.g., above 90%) | 96.1% (73/76) (95% CI: 88.9-99.2%) |
| Relative Specificity (vs. WB) | High (e.g., above 90%) | 97.0% (159/164) (95% CI: 93.0-99.0%) |
| For Culture Positives (n=63) | ||
| Relative Sensitivity (vs. culture) | High (e.g., above 90%) | 96.8% (61/63) (95% CI: 89.0-99.6%) |
| Relative Sensitivity (vs. WB) | High (e.g., above 90%) | 98.4% (61/62) (95% CI: 91.3-100%) |
| For Low Prevalence Population (n=81) | ||
| Relative Specificity (vs. WB) | High (e.g., above 90%) | 98.7% (77/78) (95% CI: 93.1-100%) |
| Relative Sensitivity (vs. WB) | High (e.g., above 90%) | 100% (2/2) (95% CI: 15.8-100%) |
| Type Specificity with HSV-1 WB Positives (n=287) | High (e.g., above 90%) | 96.5% (276/286) (95% CI: 93.7-98.3%) |
| Cross-reactivity with Taxonomically Related Viruses (n=27) | High agreement (e.g., above 90% negative agreement) | Total 90.9% (70/77) (95% CI: 82.2-96.3%) |
| Agreement between Manual and Automated Methods (n=257) | High (e.g., above 95% overall agreement) | 98.1% (252/257) (95% CI: 95.5-99.4%) |
| Inter-assay Reproducibility (automated) | Low CV (acceptable variability) | Samples had inter-assay CVs ranging from 0.9% to 25.2% |
2. Sample Sizes and Data Provenance for Test Set
- Sexually Active Adults: n = 246 (retrospective, archived, masked sera)
- Expectant Mothers: n = 241 (retrospective, archived, masked sera)
- Culture Positive Patients: n = 63 (sera from culture positive patients)
- Low Prevalence Population: n = 81 (college students claiming to lack sexual experience)
- Type Specificity (HSV-1 WB positives): n = 287 (from expectant mothers, sexually active adults, low prevalence persons, and HSV-1 culture positives)
- Cross-reactivity with Taxonomically Related Viruses: n = 27 (HSV sero-negative by other FDA cleared HSV ELISAs, and IFA IgG positive for CMV, EBV VCA, HHV6, VZV)
- CDC Panel: Undisclosed specific number of samples (panel consists of 37% positive and 63% negative samples).
- Manual vs. Automated Method Agreement: n = 257 (adult samples submitted for HSV testing, 255 from US, 2 from outside US, retrospectively selected and manually tested by an external investigator as part of CLIA validation).
Data Provenance: The data primarily originated from a "Pacific Northwest university" for Western blot reference testing and a "major university located in the Northwestern United States" for discrepant analysis of cross-reactivity. One study on manual vs. automated method agreement involved a "major clinical laboratory located in Southern California" with samples predominantly from the US (255/257). The CDC panel information suggests samples from the US. The studies are described as using "masked, archived, and unselected sera" or "sequentially submitted" sera, indicating a retrospective data collection approach for most studies.
3. Number of Experts and Qualifications for Ground Truth
The document does not explicitly state the number of experts used to establish the ground truth for the test set or their specific qualifications (e.g., "radiologist with 10 years of experience"). However, it clearly states that the reference method for determining HSV-2 serostatus was an HSV-2 Western blot (WB) performed at a "Pacific Northwest university." For culture-positive patients, the reference methods included both culture (infection) and the HSV-2 Western blot. For cross-reactivity studies, a "type specific Western blot from a major university located in the Northwestern United States" was used for discrepant analysis.
For the "Agreement with CDC Panel," the ground truth was established by the CDC, implying a highly qualified and standardized reference.
The interpretation of Western blot results for HSV-2 would typically be performed by trained specialists in a reference laboratory, although their specific credentials are not detailed in this summary.
4. Adjudication Method for the Test Set
The document does not describe a formal adjudication method (like 2+1 or 3+1 consensus) for the initial Western blot interpretation or other reference methods. The Western blot results are presented as the established ground truth. Discrepant analysis between the HerpeSelect ELISA and other methods (e.g., other FDA cleared HSV ELISAs for cross-reactivity) was performed using a "type specific Western blot," suggesting this was the arbiter for discrepancies in those specific cases.
5. Multi-Reader Multi-Case (MRMC) Comparative Effectiveness Study
No multi-reader multi-case (MRMC) comparative effectiveness study comparing human readers with and without AI assistance (or in this case, the automated ELISA) is described. The studies focus on the performance of the HerpeSelect 2 ELISA IgG assay (both manual and automated options) against a reference standard (Western blot or culture), not on the improvement of human readers with AI assistance.
6. Standalone Performance Study
Yes, standalone performance studies were done. The entire document describes the performance of the HerpeSelect® 2 ELISA IgG device (both the manual and the automated option) as a standalone diagnostic tool. Its sensitivity, specificity, agreement, and reproducibility were evaluated independently against established reference methods (Western blot, culture) or in comparison with its own manual method.
7. Type of Ground Truth Used
The primary type of ground truth used was expert reference method, specifically:
- HSV-2 Western blot (WB): Considered the gold standard for HSV-2 antibody detection. This was used across most population studies (expectant mothers, sexually active adults, low prevalence population, type specificity).
- Culture (infection): Used as a reference alongside Western blot for the "culture positives" study.
- CDC Panel: A characterized serum panel from the CDC, implying a highly validated ground truth.
- Previously FDA cleared HSV ELISAs and IFA IgG for taxonomically similar viruses: Used as initial screening/reference for the cross-reactivity study, with Western blot for discrepant analysis.
8. Sample Size for the Training Set
The document does not provide information about a "training set" for the HerpeSelect® 2 ELISA IgG assay. This assay is a diagnostic laboratory test, not an AI or machine learning algorithm that typically requires distinct training sets. The studies described are performance evaluations of the final, already developed assay.
9. How Ground Truth for the Training Set Was Established
As there is no mention of a "training set" for algorithm development in this context (the device is an ELISA assay rather than an AI/ML algorithm), the method for establishing ground truth for a training set is not applicable or described. The performance data presented refers to the test set or validation set of samples on which the device's accuracy was demonstrated.
§ 866.3305 Herpes simplex virus serological assays.
(a)
Identification. Herpes simplex virus serological assays are devices that consist of antigens and antisera used in various serological tests to identify antibodies to herpes simplex virus in serum. Additionally, some of the assays consist of herpes simplex virus antisera conjugated with a fluorescent dye (immunofluorescent assays) used to identify herpes simplex virus directly from clinical specimens or tissue culture isolates derived from clinical specimens. The identification aids in the diagnosis of diseases caused by herpes simplex viruses and provides epidemiological information on these diseases. Herpes simplex viral infections range from common and mild lesions of the skin and mucous membranes to a severe form of encephalitis (inflammation of the brain). Neonatal herpes virus infections range from a mild infection to a severe generalized disease with a fatal outcome.(b)
Classification. Class II (special controls). The device is classified as class II (special controls). The special control for the device is FDA's revised guidance document entitled “Class II Special Controls Guidance Document: Herpes Simplex Virus Types 1 and 2 Serological Assays.” For availability of the guidance revised document, see § 866.1(e).