(107 days)
The Immunetics® QuickELISA™ Anthrax-PA Kit is intended for use in the qualitative detection of antibodies to the Protective Antigen (PA) protein of B. anthracis in human serum. The assay should be used only on serum samples from individuals with a clinical history, signs or symptoms consistent with anthrax infection as an aid in the diagnosis of anthrax, or from recipients of anthrax vaccine.
- The diagnosis of anthrax infection must be made based on history, signs, symptoms, Exposure likelihood, and when possible other laborative rooults in addition to the presence of antiation from other evidence to exclude anthrax.
- The assay has not been evaluated in individuals without clinical signs on symptoms who were probamou oxposures on information available to interpret cutaneous of innulation antinentive test result for such individuals
- the meaning of a positive of negativens from patients infected by the The assay fluo not boon orded results with such infections are unknown.
- The minimum level of anti-PA antibodies that confers protection following the vaccination is not known. This galue and protective immunity has not been established.
- The affinity and/or avidity of anti-PA IgG and IgM for the rPA antigen have not been determined with this assay.
The Immunetics® QuickELISA Anthrax-PA Kit allows qualitative detection in human serum of total antibodies to the Protective Antigen (PA) protein of B. anthracis, in a non-subtype specific, non-species specific microwell ELISA format. The PA toxin component of B. anthracis has been shown to elicit a detectable IgG response in naturally infected individuals and vaccine recipients. The assay is based on a single binding step, in which serum antibodies are incubated with a mixture of two rPA conjugates: Streptavidin-rPA and rPA-horseradish peroxidase (rPA-HRP). PA-specific, multivalent antibodies are able to form ternary complexes in which streptavidin-TPA and rPA-HRP are bound to different antigen combining sites on a single antibody molecule. The complexes are bound to the biotin-coated microplate via the streptavidin conjugate and detected via the HRP conjugate.
In the assay procedure Sample Diluent, serum samples and Conjugate mixture are added to wells and incubated. Unbound antibodies and Conjugate are removed by wash steps, and bound anti-PA antibodies and HRP Conjugate are detected by adding a chromogenic peroxidase substrate containing tetramethylbenzidine (TMB). A blue product is produced in wells where antibody has been bound to the solid phase. The color development reaction is quenched by addition of an acidic Stop Solution, which causes a change in color from blue to yellow, after which optical Absorbance at 450nm corrected by 620-650mm background subtraction is measured in each well using an ELISA microplate reader. Turn-around time for the test is approximately 35 minutes if agitation is used, or 70 minutes without agitation. While the assay provides a qualitative result when undiluted samples are tested, it may also be used to obtain semi-quantitative anti-PA IgG titer results when serial dilution data are collected for the scrum sample, in similar fashion to that described by U.S. Centers for Disease Control (CDC) Quinn, C.P. et al. Emerging Infectious Diseases 2002; 8(10):1103-1110.
The provided 510(k) summary for the QuickELISA Anthrax-PA Kit describes performance studies, but it does not explicitly define formal acceptance criteria in terms of specific thresholds for sensitivity, specificity, or reproducibility that the device needed to meet. Instead, it presents various performance metrics as "Summary of Performance."
Therefore, the table below will report the observed performance from the studies rather than predefined acceptance criteria.
1. Table of Acceptance Criteria and Reported Device Performance
Note: Formal acceptance criteria (pre-defined thresholds for success) are not explicitly stated in the provided document. The table below presents the reported performance from the studies.
| Performance Metric | Acceptance Criteria (Not Explicitly Stated) | Reported Device Performance |
|---|---|---|
| Specificity (Normal Sera) | Not explicitly stated | 99.14% (578/583) with 95% CI: 98.01% - 99.72% (Table 1.1) |
| 100% (100/100) for Stationary incubation | ||
| 100% (104/104) for Agitated incubation (Table 1.2) | ||
| Cross-Reactivity (Interference Study) | Not explicitly stated | Overall: 99.56% (224/225) with 95% CI: 97.55% - 99.99% (Table 2.1) |
| Specific conditions: HIV (100%), HBV (100%), H. pylori (100%), Flu vaccines (100%), etc. | ||
| RPR (syphilis): 95.0% (19/20) (Table 2.2) | ||
| Sensitivity | Not explicitly stated | AVA Vaccinees: 100% (49/49) with 95% CI: 92.75% - 100% |
| All Patients: 100% (34/34) with 95% CI: 89.72% - 100% (Note: Appears to be based on 19 positive cases and 15 negative cases based on the "19" in the table's "No. Positive" column for "All Patients". The presented number of total cases is inconsistent.) | ||
| Cutaneous Anthrax: 100% (18/18) with 95% CI: 81.47% - 100% (Note: Appears to be based on 13 positive cases and 5 negative cases.) | ||
| Inhalational Anthrax: 100% (16/16) with 95% CI: 79.41% - 100% (Note: Appears to be based on 6 positive cases and 10 negative cases.) (Table 3.1) | ||
| Analytical Sensitivity | Not explicitly stated | ≤31.3 ng/mL (based on LL-95%-C-INT (REF dilution) vs. UL-95%-C-INT (NC)) |
| 229 ng/mL (Sensitivity @ Cutoff based on Best-fit 4-Parameter Curve Equation) (Table 4.1) | ||
| Reproducibility | Not explicitly stated | Inter-Assay %CVs ranging from 2.2% to 14.8% for various samples (Table 6.1) |
| Inter-Lot %CVs ranging from 0.9% to 26.3% for various samples (Table 6.2) |
2. Sample Size Used for the Test Set and Data Provenance
- Specificity (Normal Human Sera):
- Sample Size:
- Table 1.1: 583 samples (578 Negative, 5 Positive)
- Table 1.2: 100 samples (Stationary incubation), 104 samples (Agitated incubation) - These appear to be subsets or different studies for normal human sera.
- Data Provenance: "non-endemic normal population" (implied healthy individuals without anthrax). No specific country of origin is mentioned, but "Normal Human Sera" suggests common laboratory sample sources. Retrospective.
- Sample Size:
- Cross-Reactive Conditions (Interference Study):
- Sample Size: 225 samples (224 Negative, 1 Positive) - This appears to be the total count for the interference study, with breakdown provided for specific conditions. E.g., ANA (21), EBV (20), HIV (20), RPR (20), etc.
- Data Provenance: From individuals with specific disease conditions (e.g., ANA, EBV, HIV, H. pylori, Influenza, Legionella, M. pneumoniae, SLE, Rheumatoid Factor, RPR (syphilis), HAV, HBV). Not specified if retrospective or prospective, or country of origin, but generally such panels are commercially sourced or from research cohorts. The note "CDC published results (JEID 2002); archived data on unrelated samples" suggests some samples were previously characterized by the CDC.
- Sensitivity (Positive Serum Panels):
- Sample Size:
- AVA Vaccinees: 49 samples (all tested positive)
- All Patients: 34 samples (all tested positive, but the table shows '19' in the positive column, which suggests an inconsistency or a subset of 19 positive cases among 34, and then "100%" reported for "All Patients" overall for the device. Given the specificity and sensitivity are percentage of correct detections, this would imply 34 actual positive cases were tested and all were detected. This requires clarification beyond the text).
- Cutaneous Anthrax: 18 samples (all tested positive)
- Inhalational Anthrax: 16 samples (all tested positive)
- Data Provenance:
- AVA Vaccinees: From recipients of the Anthrax Vaccine Adsorbed (AVA).
- Patients: From individuals with confirmed anthrax infection (cutaneous and inhalational).
- The note "Sensitivity in CDC PA IgG-specific ELISA = 100 % in all above groups" suggests these samples were characterized by the CDC.
- Not specified if retrospective or prospective, or country of origin, but likely from US clinical/public health sources given CDC involvement.
- Sample Size:
- Analytical Sensitivity (PA IgG Reference Dilutions):
- Sample Size: Not a patient sample size, but a set of reference material dilutions (e.g., 1000 ng/mL down to 0 ng/mL).
- Data Provenance: The 1.0 ug/mL PA IgG Reference Sample (REF-1) was "prepared from CDC standard serum pool designated AVR414 (described in JEID, 2002)".
3. Number of Experts Used to Establish the Ground Truth for the Test Set and Qualifications of Those Experts
The document does not provide information on the number or qualifications of experts used to establish the ground truth for the test set.
However, the "Sensitivity in CDC PA IgG-specific ELISA = 100 % in all above groups" for positive samples and the reference to "CDC published results (JEID 2002); archived data on unrelated samples" for cross-reactivity and normal sera analysis strongly suggest that the ground truth for patient samples and reference materials was established by the U.S. Centers for Disease Control (CDC). The CDC, as a leading public health agency, would be considered the authoritative source for anthrax diagnosis and characterization of antibody responses. While specific expert qualifications are not listed, the involvement of the CDC implies a high level of expertise in infectious diseases and serological testing.
4. Adjudication Method for the Test Set
The document does not explicitly describe any adjudication method (e.g., 2+1, 3+1, none) for establishing the ground truth for the test set.
As mentioned above, the ground truth appears to be established by the CDC, likely through their existing laboratory protocols and reference assays (e.g., CDC PA IgG-specific ELISA). This implies a high standard of reference, but the specific process of resolving discordant results among multiple readers or tests is not detailed for the ground truth establishment.
5. If a Multi-Reader Multi-Case (MRMC) Comparative Effectiveness Study Was Done
No, a multi-reader, multi-case (MRMC) comparative effectiveness study was not done.
This device is an in-vitro diagnostic (IVD) kit designed for laboratory use to detect antibodies, not for interpretation by human readers in the same way as an imaging device might be. Therefore, the concept of human readers improving with AI vs. without AI assistance does not apply to this type of device. The performance is based on the assay's ability to detect antibodies in serum samples.
6. If a Standalone (i.e., algorithm only without human-in-the-loop performance) Was Done
Yes, the provided studies describe the standalone performance of the QuickELISA Anthrax-PA Kit.
The performance metrics (specificity, sensitivity, cross-reactivity, analytical sensitivity, reproducibility) are all measured directly from the kit's output (optical absorbance, then translated into a qualitative positive/negative result) when testing serum samples. This is inherently a standalone performance evaluation of the device itself, without human interpretation as a variable in the core diagnostic result generation. The human "in-the-loop" for this device would be the laboratory technician performing the assay and reading the microplate reader, but this is standard operation for IVDs and not considered "human-in-the-loop performance" in the context of an AI-assisted diagnostic.
7. The Type of Ground Truth Used
The type of ground truth used appears to be a combination of:
- Expert Consensus/Reference Standard Serology: For positive samples (AVA vaccinees, cutaneous anthrax, inhalational anthrax patients), the ground truth was established by the CDC PA IgG-specific ELISA, which is referred to as having 100% sensitivity for these groups. This indicates a high-level reference standard.
- Clinical Diagnosis/Outcomes Data: For anthrax patients, their classification (cutaneous, inhalational) implies a clinical diagnosis confirmed by other means (symptoms, history, exposure, and possibly other laboratory data), consistent with the intended use statement.
- Known Negative Status: For specificity and cross-reactivity studies, the ground truth was "Normal Human Sera" (presumably healthy individuals without anthrax exposure/vaccination) and "Interference Sera" from individuals with documented conditions known not to be anthrax (e.g., HIV, EBV, Influenza).
8. The Sample Size for the Training Set
The document does not provide information on a dedicated training set or its sample size.
Given that this is a 510(k) submission for a non-AI/ML in-vitro diagnostic kit (an ELISA), there isn't typically a "training set" in the machine learning sense. The device's components (antigens, antibodies, reagents) and assay protocol are developed through R&D and optimization phases, which would involve internal testing and validation. The data presented in the summary represents the analytic and clinical validation of the final device, effectively acting as the "test set" to demonstrate its performance against established ground truth.
9. How the Ground Truth for the Training Set Was Established
As there is no explicit "training set" mentioned in the context of the 510(k) submission for this ELISA kit, the method for establishing its ground truth is not applicable. The data presented is from the validation studies of the final product.
§ 866.3045 In vitro diagnostic device for
Bacillus spp. detection.(a)
Identification. An in vitro diagnostic device forBacillus species (spp.) detection is a prescription device used to detect and differentiate amongBacillus spp. and presumptively identifyB. anthracis and otherBacillus spp. from cultured isolates or clinical specimens as an aid in the diagnosis of anthrax and other diseases caused byBacillus spp. This device may consist ofBacillus spp. antisera conjugated with a fluorescent dye (immunofluorescent reagents) used to presumptively identify bacillus-like organisms in clinical specimens; bacteriophage used for differentiatingB. anthracis from otherBacillus spp. based on susceptibility to lysis by the phage; or antigens used to identify antibodies toB. anthracis (anti-toxin and anti-capsular) in serum. Bacillus infections include anthrax (cutaneous, inhalational, or gastrointestinal) caused byB. anthracis, and gastrointestinal disease and non-gastrointestinal infections caused byB. cereus. (b)
Classification. Class II (special controls). The special controls are set forth in FDA's special controls guideline document entitled “In Vitro Diagnostic Devices forBacillus spp. Detection; Class II Special Controls Guideline for Industry and Food and Drug Administration Staff.” For availability of the guideline document, see § 866.1(e).(c)
Restriction on Distribution. The distribution of these devices is limited to laboratories that follow public health guidelines that address appropriate biosafety conditions, interpretation of test results, and coordination of findings with public health authorities.(d)
Restriction on Use. The use of this device is restricted to prescription use and must comply with the following:(1) The device must be in the possession of:
(i)(A) A person, or his agents or employees, regularly and lawfully engaged in the manufacture, transportation, storage, or wholesale or retail distribution of such device; or
(B) A practitioner, such as a physician, licensed by law to use or order the use of such device; and
(ii) The device must be sold only to or on the prescription or other order of such practitioner for use in the course of his professional practice.
(2) The label of the device shall bear the statement “Caution: Federal law restricts this device to sale by or on the order of a ____”, the blank to be filled with the word “physician” or with the descriptive designation of any other practitioner licensed by the law of the State in which he practices to use or order the use of the device.
(3) Any labeling, as defined in section 201(m) of the Federal Food, Drug, and Cosmetic Act, whether or not it is on or within a package from which the device is to be dispensed, distributed by, or on behalf of the manufacturer, packer, or distributor of the device, that furnishes or purports to furnish information for use of the device contains adequate information for such use, including indications, effects, routes, methods, and frequency and duration of administration and any relevant hazards, contraindications, side effects, and precautions, under which practitioners licensed by law to employ the device can use the device safely and for the purposes for which it is intended, including all purposes for which it is advertised or represented. This information will not be required on so-called reminder-piece labeling which calls attention to the name of the device but does not include indications or other use information.
(4) All labeling, except labels and cartons, bearing information for use of the device also bears the date of the issuance or the date of the latest revision of such labeling.