(222 days)
The Biotest EBNA IgG ELISA is an enzyme immunoassay using a recombinant antigen for the qualitative detection of IgG antibodies to the Epstein-Barr Virus (EBV) EBNA-1 (Nuclear Antigen 1) in human serum or plasma. Results obtained with this test, in conjunction with other clinical and patient data obtained in assays for other Epstein-Barr antigens such as Early Antigen IgG and IgM, assist in serological diagnosis of EBV infection in pediatric and adult populations.
Using recombinant DNA technology, Biotest has developed three highly purified EBV antigens for use in their ELISA test system.
- EBNA-1 p72: Major antigen of the EBNA complex. The recombinant protein does not contain the glycine-alanine copolymer, a structural feature of EBNA-1, which shows cross-reactivities with certain autoantibodies and CMV (IgM response).
- EA-D p64: Early antigen. Dominant immunogen of the EA-D complex.
- EA p138: Early antigen and major DNA binding protein. This highly reactive antigen is not detectable in EA immunofluorescence assays based on chemically induced Raji cells (deletion within the Raji genome).
The EBV immune status will be determined by the detection of specific antibodies directed against EBV proteins according to the principle of the indirect ELISA. The antigens are purified to apparent homogeneity and immobilized on the solid phase (microtest plate, 96 wells). If the patient's serum contains specific antibodies they will bind during the first incubation. Non-specific antibodies are removed by washing steps. During a second incubation the captured IgG antibodies are labeled. This is performed by addition of murine monoclonal anti-human IgG antibody-enzyme-conjugates. The final reaction converts a colorless substrate to a colored product. The concentration of color after a definite time is related to the concentration of antibody in the serum sample.
Here's a detailed breakdown of the acceptance criteria and the study proving the device meets them, based on the provided text:
Acceptance Criteria and Device Performance
The acceptance criteria for the Biotest Anti-EBV Recombinant EBNA IgG ELISA device primarily revolve around its sensitivity and specificity in detecting IgG antibodies to EBV EBNA-1. The reported device performance is based on direct comparison with a predicate method (ACIF) and clinical interpretation of patient samples.
1. Table of Acceptance Criteria and Reported Device Performance
| Criteria Category | Acceptance Criteria (Implied/Derived) | Reported Device Performance (Biotest EBNA IgG ELISA) |
|---|---|---|
| Overall Relative Sensitivity | Comparable to the predicate method (EBNA ACIF). (No specific numerical threshold given, but strong agreement is implied). | 86.8% (95% C.I. = 82.1 to 91.5%) across both sites |
| Overall Relative Specificity | Comparable to the predicate method (EBNA ACIF). (No specific numerical threshold given, but strong agreement is implied). | 97.1% (95% C.I. = 93.9 to 98.9%) across both sites |
| Overall Relative Agreement | High agreement with the predicate ACIF. (No specific numerical threshold given). | 92.1% across both sites |
| Clinical Sensitivity (Total) | High sensitivity across various EBV infection stages. (No specific numerical threshold given). | 98.5% (95% C.I. = 96.6 to 99.5%) |
| Clinical Specificity (Total) | High specificity for negative cases. (No specific numerical threshold given). | 98.6% (95% C.I. = 92.3 to 100%) |
| Clinical Agreement (Total) | High agreement with clinical interpretation. (No specific numerical threshold given). | 98.5% |
| Cross-Reactivity | No cross-reactivity with common interfering viruses. | Not observed with HSV I/II, VZV, CMV. |
Note: The document does not explicitly state numerical acceptance criteria thresholds, but rather presents the study results as justification for the device's performance compared to established methods and clinical understanding. The implied acceptance is that the device performs comparably to or better than existing methods for the intended use.
2. Sample Size for the Test Set and Data Provenance
- Sample Size for Test Set: 408 patient samples.
- Data Provenance:
- Country of Origin: Not explicitly stated, but the study was conducted at "two geographically distinct locations" within the context of a US FDA submission, implying US-based data.
- Retrospective or Prospective: Not explicitly stated. The description "samples were obtained from both pediatric and adult patients (ages 1 to 74) representing acute (139), late acute (34), recent past (12), past (120), reactivation (22), past/probable reactivation (11), and negative (70) disease stages of EBV infection" suggests these were pre-existing samples categorized by their disease stage, which points towards a retrospective collection and analysis.
3. Number of Experts Used to Establish Ground Truth and Qualifications
The document doesn't explicitly state the "number of experts" or their specific "qualifications" for establishing the ground truth. Instead, it refers to:
- For "Direct Comparison": "commercially available or published EBNA anti-complement immunofluorescence (ACIF) tests." This implies that the ground truth for this comparison was established by the ACIF method, which is a recognized laboratory technique, not directly by human experts for each case in the same way a pathologist would review slides.
- For "Clinical Interpretation": "stage of infection based on antibody patterns and clinical diagnosis at the time the specimen was drawn." This suggests that the ground truth here was derived from established clinical diagnostics and antibody profiling done by medical professionals, but not necessarily a panel of "experts" specifically adjudicating each case for the study.
4. Adjudication Method for the Test Set
- None specified. The study utilized two primary comparisons:
- Direct comparison with ACIF: This is a direct laboratory test comparison, not a human adjudication process.
- Comparison with clinical interpretation: This relies on pre-existing clinical diagnoses and antibody patterns, also not implying an adjudication panel for the study itself.
5. Multi-Reader Multi-Case (MRMC) Comparative Effectiveness Study
- No. An MRMC study was not done. This device is an in-vitro diagnostic (IVD) ELISA kit, which is typically evaluated for its direct analytical performance and agreement with established reference methods or clinical gold standards, rather than assessing human reader improvement with AI assistance. The concept of "human readers" is not applicable to the performance evaluation of such a diagnostic kit.
6. Standalone (Algorithm Only) Performance
- The study is a standalone performance evaluation of the Biotest Anti-EBV Recombinant EBNA IgG ELISA kit. As an IVD, its performance is assessed independently (without human-in-the-loop assistance for the test itself), by comparing its results to a ground truth (ACIF or clinical interpretation).
7. Type of Ground Truth Used
Two types of ground truth were used:
- Reference Method (ACIF): For the "direct comparison," the ground truth was "commercially available or published EBNA anti-complement immunofluorescence (ACIF) tests."
- Clinical Interpretation: For the second evaluation, the ground truth was "clinical interpretation (stage of infection based on antibody patterns and clinical diagnosis at the time the specimen was drawn)." This incorporates a combination of established diagnostic criteria and patient medical history.
8. Sample Size for the Training Set
- Not applicable / Not explicitly mentioned. ELISA kits are laboratory assays, not machine learning algorithms that require a "training set" in the conventional sense. The "development" of the assay (e.g., antigen selection, antibody optimization) would involve extensive R&D, but this is distinct from the concept of a training set for an AI model. The provided document details the validation of the finalized device.
9. How the Ground Truth for the Training Set Was Established
- Not applicable. As explained above, this device does not utilize a training set in the machine learning context. The "ground truth" for the analytical performance of the antigens and antibodies used in the kit would have been established during the development phase through biochemical methods, reference panels, and expert characterization of EBV components, but this is an inherent part of IVD development, not a "training set" process for an algorithm.
§ 866.3235 Epstein-Barr virus serological reagents.
(a)
Identification. Epstein-Barr virus serological reagents are devices that consist of antigens and antisera used in serological tests to identify antibodies to Epstein-Barr virus in serum. The identification aids in the diagnosis of Epstein-Barr virus infections and provides epidemiological information on diseases caused by these viruses. Epstein-Barr viruses are thought to cause infectious mononucleosis and have been associated with Burkitt's lymphoma (a tumor of the jaw in African children and young adults) and postnasal carcinoma (cancer).(b)
Classification. Class I (general controls).