(90 days)
The INFINITI 2C9 & VKORC1 Multiplex Assay for Warfarin is an in vitro diagnostic test for the detection and genotyping of the *2 and *3 CYP4502C9 genetic variants and the VKORC1 3673 (-1639) intronic variant in genomic deoxyribonucleic acid (DNA) obtained from EDTA-anticoagulated whole blood samples. The INFINITI 2C9 & VKORC1 Multiplex Assay for Warfarin is a qualitative assay for use in clinical laboratories upon prescription by the attending physician.
The INFINITI 2C9 & VKORC1 Multiplex Assay for Warfarin is indicated for use to identify individuals at risk for sensitivity to warfarin.
The INFINITI 2C9 & VKORC1 Multiplex Assay for Warfarin is an in vitro diagnostic device which utilizes proprietary film-based microarray technology combined with process automation, reagent management, and software technology for the detection and genotyping of the 2C92, 2C93, and VKORC1 3673 (-1639) mutations from EDTA-anticoagulated whole blood samples.
The INFINITI 2C9 & VKORC1 Multiplex Assay for Warfarin is comprised of the BioFilmChip™ Microarray, the Intellipac Reagent Module and the PCR Amplification Mix. The INFINITI 2C9 & VKORC1 Multiplex Assay for Warfarin should be run using the AutoGenomics INFINITI Analyzer.
The BioFilmChip Microarray consists of a polyester film coated with proprietary multi-layer components designed for DNA analysis. The layers have been designed to provide a versatile surface to enhance test performance. There can be up to 240 spots per microarray with each spot representing a different allele. The microarrays are designed to be assay specific.
The Intellipac Reagent Module contains up to eight reservoirs that house the test reagents and has an integrated memory chip. Information on the reagent such as lot number, expiration date and volume usage, are archived in the memory.
The PCR Amplification Mix consists of the reagents needed for the PCR amplification step of the assay.
The INFINITI 2C9 & VKORC1 Multiplex Assay for Warfarin is based on the following processes:
(a) DNA extraction
(b) PCR amplification of purified DNA from human genomic DNA
(c) Labeling of the amplified product (allele specific primer extension)
(d) Hybridization of the labeled amplified product to a microarray by signature Tag/Capture probe hybridization under isothermal conditions.
(e) Scanning of the microarray
(f) Signal detection and analysis [determination of the 2C92, 2C93 and VKORC1 3673 (-1639) genotypes]
Steps (c) through (f) are automated by the INFINITI Analyzer.
The INFINITI Analyzer automates the 2C9 and VKORC1 assays and integrates all the discrete processes of sample (PCR amplicon) handling, reagent management, hybridization, detection, and results analysis. The assays are processed automatically and read by the built-in confocal microscope. Results are analyzed and presented as genotype calls.
Here's an analysis of the acceptance criteria and study details for the INFINITI 2C9 & VKORC1 Multiplex Assay for Warfarin, based on the provided text:
Acceptance Criteria and Device Performance
The document describes the performance characteristics without explicitly stating pre-defined "acceptance criteria" as pass/fail thresholds. Instead, it presents the "reported device performance" and implies that these results demonstrate the device's suitability. For the purpose of this analysis, I will synthesize the reported performance values as the de facto "acceptance criteria" that the device met.
1. Table of Acceptance Criteria and Reported Device Performance
| Performance Metric | Acceptance Criteria (Implied) | Reported Device Performance |
|---|---|---|
| Analytical Specificity | PCR primer, ASP primer, and capture probe specificity confirmed. | PCR primer specificity by amplicon size & sequencing. ASP primer specificity by correct calls with known genomic samples. Capture probe specificity by hybridization of correct oligo to known spot. |
| Limit of Detection (Analytical Sensitivity) | Minimum DNA concentration for correct calls. Recommended DNA concentration for reliable results. | Minimum DNA concentration for correct calls: 1 ng DNA. Recommended DNA concentration: 25 ng/µl (50 ng per test). Correct calls at 10 ng DNA. |
| Agreement with Bi-directional Sequencing (1st run) | High percentage agreement for *2, *3, and VKORC1 variants. | 2C9*2: 98.0% agreement (95% CI Lower Limit: 95.09%) |
| 2C9*3: 97.3% agreement (95% CI Lower Limit: 94.09%) | ||
| VKORC1 3673 (-1639): 98.0% agreement (95% CI Lower Limit: 95.09%) | ||
| Overall assay agreement. | Total for Assay: 97.8% agreement (95% CI Lower Limit: 96.86%) | |
| Agreement with Bi-directional Sequencing (Final Result - after re-run of no calls) | High percentage agreement, minimizing no calls and incorrect calls. | Total for Assay: 99.3% correct call rate (149/150 samples) for Table 2b (by Sample Type) |
| Assay Inter-Laboratory Reproducibility (Genotype Call Rate) | High percentage of correct genotype calls across sites, within each genotype, and overall. | 2C9*2 (Total): 99.07% (98.06% 95% CI Lower Limit) |
| 2C9*3 (Total): 97.22% (90.47% 95% CI Lower Limit) | ||
| VKORC1 3673 (-1639) GA (Total): 99.44% (98.08% 95% CI Lower Limit) | ||
| VKORC1 3673 (-1639) AA (Total): 97.22% (92.73% 95% CI Lower Limit) | ||
| VKORC1 3673 (-1639) GG (Total): 100.0% (98.08% 95% CI Lower Limit) | ||
| Total for Assay (Reproducibility): 98.99% correct calls (98.42% 95% CI Lower Limit) after 1st run; 99.96% after repeat. | ||
| Drug Interference | No interference from common interfering substances. | No interference from bilirubin (8mg/dl), cholesterol (70mg/dl), and heparin (133v/dl). |
| Sample Carry-Over | No detection of sample carry-over. | 100% correct genotype calls when positive sample followed by another positive or "No Template Control". |
| Assay Interference | No interference between this assay and other assays on the same instrument. | No interference when run simultaneously with INFINITI Assay for Factor II & Factor V. |
| Reagent Stability | Demonstrated shelf-life at specified storage conditions. | BioFilmChip Microarray: 12 months at RT (15-30°C). Intellipac Reagent: 12 months Refrigerated (2-8°C). Amplification Mix: 12 months Frozen (-10°C). |
2. Sample Sizes Used for the Test Set and Data Provenance
Sample Size for Test Set:
- Agreement with Bi-directional Sequencing:
- 150 patient samples tested in the initial comparison (first run data for 2C9*2, 2C9*3, VKORC1).
- Table 2b "by Sample Type" also shows 150 samples, implying these are the same samples analyzed by genotype.
- Inter-Laboratory Reproducibility:
- 7 genomic DNA samples and 5 whole blood samples (total 12 unique samples).
- Each unique sample was run in duplicate per day/operator for six days at each of 3 sites.
- Total tests: 12 samples * 2 replicates/day * 6 days * 3 sites = 432 tests per site, for a total of 1296 tests across all sites for the "first time run" and "final result".
Data Provenance:
- Agreement with Bi-directional Sequencing:
- Origin: Not explicitly stated, but "patient samples" were used. Given the FDA submission context, it's highly likely these were de-identified samples from the United States.
- Nature: The description "Each site tested its own patient samples" and that they were "from patients using or have used warfarin" suggests these were retrospective samples, collected from a patient population relevant to the intended use.
- Inter-Laboratory Reproducibility:
- Origin: Not explicitly stated, but likely from the United States given the submission.
- Nature: Controlled study using "seven genomic DNA samples and five whole blood samples." These would be prospective in the sense that they were specifically prepared and distributed for this reproducibility study, though the original source of the genetic material might have been retrospective.
3. Number of Experts Used to Establish the Ground Truth for the Test Set and Qualifications
The document does not mention the use of experts to establish ground truth.
For the Agreement with Bi-directional Sequencing study, the ground truth was established by:
- Bi-directional DNA sequencing. This is a recognized laboratory method for determining genetic sequences and is considered the gold standard for many genetic variants. Thus, the ground truth was established by laboratory method, not expert consensus.
- The document implies that the sequencing results were accepted as the definitive truth without the need for expert adjudication or review.
For the Inter-Laboratory Reproducibility study, the ground truth for the 7 genomic DNA samples and 5 whole blood samples was their known genotypes, which would have been previously determined, presumably also by a method like bi-directional sequencing.
4. Adjudication Method for the Test Set
The document does not describe an adjudication method involving multiple human reviewers for the test set.
- In the "Agreement with Bi-directional Sequencing" study, "bi-directional sequencing" served as the comparator/ground truth. The device results were directly compared to these sequencing results.
- For the inter-laboratory reproducibility study, device results from different sites and operators were compared to the known genotype of the samples.
5. Multi-Reader Multi-Case (MRMC) Comparative Effectiveness Study
No MRMC comparative effectiveness study was done. This study assesses the improvement of human readers with AI assistance versus without AI assistance. The INFINITI 2C9 & VKORC1 Multiplex Assay for Warfarin is an in vitro diagnostic device for genotype detection, not an imaging or diagnostic support tool that assists human readers. Its output is a genotype call, not an interpretation that radiologists or other human experts would then refine or improve upon.
6. Standalone Performance Study (Algorithm Only)
Yes, a standalone performance study was done. The entire submission details the performance of the "INFINITI 2C9 & VKORC1 Multiplex Assay for Warfarin" as a standalone device. The device automates key steps and provides genotype calls:
- "Steps (c) through (f) are automated by the INFINITI Analyzer."
- "The INFINITI Analyzer automates the 2C9 and VKORC1 assays and integrates all the discrete processes of sample (PCR amplicon) handling, reagent management, hybridization, detection, and results analysis. The assays are processed automatically and read by the built-in confocal microscope. Results are analyzed and presented as genotype calls."
- The performance metrics (agreement with sequencing, reproducibility, LOD, specificity, etc.) are all measures of the algorithm's direct output.
There is no human-in-the-loop component described for its routine operation or performance evaluation.
7. Type of Ground Truth Used
The primary ground truth used for the performance studies was established laboratory method results, specifically bi-directional DNA sequencing. This method is considered highly accurate for determining DNA sequences and genetic variants.
8. Sample Size for the Training Set
The document does not specify a sample size for a training set. This is typical for in vitro diagnostic devices based on established molecular biology principles (PCR, hybridization, genotyping microarrays) where the design is more mechanistic and less dependent on machine learning models requiring extensive "training data" in the conventional sense. The "development" of the assay mentioned for analytical specificity would involve testing various known samples, but these are generally considered part of the assay development and validation rather than a distinct "training set" for an algorithm in the AI context.
9. How the Ground Truth for the Training Set Was Established
Since no explicit training set is detailed, the method for establishing its ground truth is also not described. If one were to consider the samples used during "assay development" as a form of training/optimization, their ground truth would also have been established by known genomic samples (e.g., cell lines with characterized genotypes, or samples sequenced by a gold-standard method).
§ 862.3360 Drug metabolizing enzyme genotyping system.
(a)
Identification. A drug metabolizing enzyme genotyping system is a device intended for use in testing deoxyribonucleic acid (DNA) extracted from clinical samples to identify the presence or absence of human genotypic markers encoding a drug metabolizing enzyme. This device is used as an aid in determining treatment choice and individualizing treatment dose for therapeutics that are metabolized primarily by the specific enzyme about which the system provides genotypic information.(b)
Classification. Class II (special controls). The special control is FDA's guidance document entitled “Class II Special Controls Guidance Document: Drug Metabolizing Enzyme Genotyping Test System.” See § 862.1(d) for the availability of this guidance document.