(28 days)
The Aptima® CMV Quant Assay is an in vitro nucleic acid amplification test for the quantitation of human cytomegalovirus (CMV) DNA in human EDTA plasma on the fully automated Panther® system.
The Aptima CMV Quant Assay is intended for use to aid in the management of solid-organ transplant patients and hematopoietic stem cell transplant patients receiving anti-CMV therapy, serial DNA measurements can be used to assess viral response to treatment. The results from Aptima CMV Quant assay must be interpreted within the context of all relevant clinical and laboratory findings.
The Aptima CMV Quant Assay is not intended for use as a screening assay for the presence of CMV in blood or blood products.
The Aptima CMV Quant Assay is an in vitro nucleic acid amplification test that uses real-time transcription mediated amplification (TMA) technology on the Panther Fusion system to quantify CMV DNA, genotypes 1, 2, 3, and 4. The primer design targets the highly conserved UL56 gene to ensure accurate quantitation of the CMV DNA. The assay is standardized to the WHO International Standard for human cytomegalovirus. The Aptima CMV Quant Assay is intended for use to aid in the management of solid-organ transplant patients and hematopoietic stem cell transplant patients.
The addition of a protocol to treat specific plasma samples with proteinase K in the assay package insert will allow the retesting of those samples to obtain a valid result.
The provided text describes a 510(k) premarket notification for the "Aptima CMV Quant Assay" and specifically details an update to the assay's package insert to include a protocol for retesting plasma samples that initially yield an "ML2 flag" invalid result. This retesting involves a manual pretreatment with proteinase K. The information provided is for a diagnostic assay, not an AI/ML-based device. Therefore, many of the requested elements pertaining to AI/ML development, such as training sets, ground truth establishment for AI/ML, expert consensus for imaging, MRMC studies, and effect sizes of AI assistance, are not applicable.
However, I can extract information related to the acceptance criteria and the study that proves the device (specifically, the proteinase K pretreatment protocol) meets the acceptance criteria within the context of a diagnostic assay's performance and validation.
Acceptance Criteria and Study for the Proteinase K Pretreatment Protocol
The primary acceptance criterion for this change is to demonstrate that the proteinase K pretreatment protocol effectively resolves the "ML2 flag" invalid results and does not negatively impact the accuracy of the CMV quantification.
1. Table of Acceptance Criteria and the Reported Device Performance:
| Acceptance Criteria (Implicit) | Reported Device Performance |
|---|---|
| Reduction/Elimination of ML2 Flag Invalid Results: The protocol should effectively enable valid results for samples initially flagged with ML2. | The prevalence of specimens invalidated with ML2 flags at the NIH decreased from 0.87% (on testing 1039 plasma specimens without proteinase K treatment) to 0% (on testing 4098 specimens with proteinase K treatment) when the proteinase K protocol was used. |
| Maintenance of Accuracy: The proteinase K pretreatment should not negatively impact the accuracy of CMV quantification. | The NIH also demonstrated that using proteinase K for pretreatment of specimens did not impact the accuracy of CMV quantification in the Aptima Assay (as referenced by Youn et al. 2024). The Hologic risk assessment concluded that the benefits outweigh the risks and the "assay performance characteristics including accuracy are maintained with incorporation of this modification." |
| Feasibility and Safety of Protocol Implementation: The protocol should be practical for use in clinical laboratories and not introduce new safety concerns for the assay. | The protocol is described with clear steps (temperature, volume, timing). The risk assessment stated that "the benefits provided by the updates to the Hologic specimen handling workflow at customer sites outweigh the risks." |
2. Sample Size Used for the Test Set and the Data Provenance:
- Test Set Sample Sizes:
- Without Proteinase K: 1039 plasma specimens
- With Proteinase K: 4098 plasma specimens
- Data Provenance: The plasma specimens were collected from the transplant patient population by the National Institute of Health (NIH). This suggests a retrospective analysis of archived samples or a prospective study conducted at the NIH. The location is the USA.
3. Number of Experts Used to Establish the Ground Truth for the Test Set and Qualifications:
- Not applicable as this is a quantitative diagnostic assay for DNA detection, not an image-based AI/ML device requiring expert interpretation of images for ground truth. The "ground truth" here is the actual CMV DNA concentration, which the assay is designed to measure.
4. Adjudication Method for the Test Set:
- Not applicable for a quantitative diagnostic assay. The performance is assessed against the ability to produce a valid quantitative result for CMV DNA and maintain accuracy, not against human consensus.
5. If a Multi-Reader Multi-Case (MRMC) Comparative Effectiveness Study was done:
- No, this is not an MRMC study. MRMC studies are typically performed for visual diagnostic aids (e.g., medical imaging AI) to compare human reader performance with and without AI assistance. This device is a direct quantitative assay.
6. If a Standalone (i.e. algorithm only without human-in-the loop performance) was done:
- Yes, in a sense. The study validates the protocol (which includes a manual pretreatment step) and its effect on the automated assay system's ability to produce a valid result. The "performance" refers to the assay's output after the sample preparation, not an AI algorithm's independent decision. The core assay itself operates in a "standalone" fashion once the sample is loaded onto the Panther system.
7. The type of ground truth used:
- The implicit ground truth is the actual CMV DNA concentration in patient samples (or spiked controls used in accuracy assessments), as determined by a reference method or known concentrations. The study's focus, however, is on resolving an assay technical issue (ML2 flag) and demonstrating that the resolution method maintains the accuracy of CMV quantification. The reference to the NIH demonstrating that "using proteinase K for pretreatment of specimens did not impact the accuracy of CMV quantification" confirms that accurate quantitation (the ground truth outcome) was assessed.
8. The Sample Size for the Training Set:
- Not applicable. This is not an AI/ML device that requires a training set in the conventional sense. The "training" for such an assay would be its initial development and optimization, which isn't described in terms of a specific "training set" size for this modification. The data provided refers to validation/test sets for the new protocol.
9. How the ground truth for the training set was established:
- Not applicable, as it's not an AI/ML device with a training set for model development.
§ 866.3180 Quantitative cytomegalovirus nucleic acid tests for transplant patient management.
(a)
Identification. A quantitative cytomegalovirus (CMV) nucleic acid test for transplant patient management is identified as a device intended for prescription use in the detection of CMV and as an aid in the management of transplant patients to measure CMV deoxyribonucleic acid (DNA) levels in human plasma and/or whole blood using specified specimen processing, amplification, and detection instrumentation. The test is intended for use as an aid in the management of transplant patients with active CMV infection or at risk for developing CMV infection. The test results are intended to be interpreted by qualified healthcare professionals in conjunction with other relevant clinical and laboratory findings.(b)
Classification. Class II (special controls). The special controls for this device are:(1) The labeling required under § 809.10(b) of this chapter must include:
(i) A prominent statement that the device is not intended for use as a donor screening test for the presence of CMV DNA in blood or blood products.
(ii) Limitations, which must be updated to reflect current clinical practice. The limitations must include, but are not limited to, statements that indicate:
(A) Test results are to be interpreted by qualified licensed healthcare professionals in conjunction with clinical signs and symptoms and other relevant laboratory results;
(B) Negative test results do not preclude CMV infection or tissue invasive CMV disease, and that CMV test results must not be the sole basis for patient management decisions.
(iii) A detailed explanation of the interpretation of results and acceptance criteria must be provided and include specific warnings regarding the potential for variability in CMV viral load measurement when samples are measured by different devices. Warnings must include the following statement, where applicable: “Due to the potential for variability in CMV viral load measurements across different CMV assays, it is recommended that the same device be used for the quantitation of CMV viral load when managing CMV infection in individual patients.”
(iv) A detailed explanation of the principles of operation and procedures for assay performance.
(2) Design verification and validation must include the following:
(i) Detailed documentation of the device description, including all parts that make up the device, reagents required for use with the CMV assay but not provided, an explanation of the methodology, design of the primer/probe sequences, rationale for the selected gene target, and specifications for amplicon size, guanine-cytosine content, and degree of nucleic acid sequence conservation. The design and nature of all primary, secondary, and tertiary quantitation standards used for calibration must also be described.
(ii) A detailed description of the impact of any software, including software applications and hardware-based devices that incorporate software, on the device's function.
(iii) Documentation and characterization of all critical reagents (
e.g., determination of the identity, supplier, purity, and stability) and protocols for maintaining product integrity throughout its labeled shelf life.(iv) Stability data for reagents provided with the device and indicated specimen types, in addition to the basis for the stability acceptance criteria at all time points chosen across the spectrum of the device's indicated life cycle, which must include a time point at the end of shelf life.
(v) All stability protocols, including acceptance criteria.
(vi) Final lot release criteria, along with documentation of an appropriate justification that lots released at the extremes of the specifications will meet the claimed analytical and clinical performance characteristics as well as the stability claims.
(vii) Risk analysis and documentation demonstrating how risk control measures are implemented to address device system hazards, such as Failure Modes Effects Analysis and/or Hazard Analysis. This documentation must include a detailed description of a protocol (including all procedures and methods) for the continuous monitoring, identification, and handling of genetic mutations and/or novel CMV stains (
e.g., regular review of published literature and annual in silico analysis of target sequences to detect possible primer or probe mismatches). All results of this protocol, including any findings, must be documented.(viii) Analytical performance testing that includes:
(A) Detailed documentation of the following analytical performance studies: Limit of detection, upper and lower limits of quantitation, inclusivity, precision, reproducibility, interference, cross reactivity, carryover, quality control, specimen stability studies, and additional studies as applicable to specimen type and intended use for the device.
(B) Identification of the CMV strains selected for use in analytical studies, which must be representative of clinically relevant circulating strains.
(C) Inclusivity study results obtained with a variety of CMV genotypes as applicable to the specific assay target and supplemented by in silico analysis.
(D) Reproducibility studies that include the testing of three independent production lots.
(E) Documentation of calibration to a standardized reference material that FDA has determined is appropriate for the quantification of CMV DNA (
e.g., a recognized consensus standard).(F) Documentation of traceability performed each time a new lot of the standardized reference material to which the device is traceable is released, or when the field transitions to a new standardized reference material.
(ix) Clinical performance testing that includes:
(A) Detailed documentation of device performance data from either a method comparison study with a comparator that FDA has determined is appropriate, or results from a prospective clinical study demonstrating clinical validity of the device.
(B) Data from patient samples, with an acceptable number of the CMV positive samples containing an analyte concentration near the lower limit of quantitation and any clinically relevant decision points.
(C) The method comparison study must include predefined maximum acceptable differences between the test and comparator method across all primary outcome measures in the clinical study protocol.
(D) The final release test results for each lot used in the clinical study.