Search Results
Found 4 results
510(k) Data Aggregation
(123 days)
ONLINE TDM Methotrexate is an in vitro test for the quantitative determination of methotrexate in human serum and plasma on cobas c systems. The determination of methotrexate is used for monitoring levels of methotrexate to ensure appropriate therapy.
The ONLINE TDM Methotrexate assay is an in vitro test for the quantitative determination of methotrexate in human serum and plasma on cobas c systems. The determination of methotrexate is used for monitoring levels of methotrexate to ensure appropriate therapy.
The ONLINE TDM MTX assay is a two-reagent system used for the detection of methotrexate in serum and plasma. In this technology drug hapten attached to the enzyme glucose 6 phosphate dehydrogenase (G6PDH) serves as the binding partner to anti-methotrexate antibody. A competitive reaction to a limited amount of specific anti-methotrexate antibody takes place between the enzyme bound hapten and free methotrexate in the sample. Enzyme activity is reduced with bound antibody. Only active enzymes reduce NAD+ to NADH. The rate of NADH formation during the reaction correlates to the methotrexate concentration and is measured photometrically.
The ONLINE TDM MTX assay is a homogeneous enzyme-immunoassay.
Reagents - working solutions
R1: Anti-methotrexate antibody (rabbit monoclonal), 3 µg/mL; NAD, G6P, bovine serum albumin in water, pH 6.3; preservative
R3: Methotrexate hapten conjugated to G6PDH, 0.3 µg/mL; bovine serum albumin in buffer, pH 7.8; preservative
The provided document describes the analytical performance evaluation of the "ONLINE TDM Methotrexate" assay. This is an in vitro diagnostic device used for the quantitative determination of methotrexate in human serum and plasma. The study aims to demonstrate that the device meets predefined acceptance criteria.
Here's an analysis of the acceptance criteria and the study proving the device meets them:
1. Table of Acceptance Criteria & Reported Device Performance:
The document doesn't explicitly present a single table of "acceptance criteria" alongside "reported device performance" in a comparative format for all tests. Instead, it states for each section: "All acceptance criteria were met," and then provides the results. We can infer the acceptance criteria from the context and the reported successful outcomes.
Here is a reconstructed table based on the provided text, indicating the acceptance criteria (inferred from the "All acceptance criteria were met" statements and industry standards implicitly followed) and the reported performance.
| Performance Characteristic | Acceptance Criteria (inferred/implied) | Reported Device Performance |
|---|---|---|
| Precision | CV (Coefficient of Variation) within acceptable limits for repeatability & intermediate precision (e.g., generally lower CVs for higher concentrations). | Repeatability:- Control 1 (0.0863 µmol/L): 4.4% CV- Control 2 (0.485 µmol/L): 0.9% CV- Control 3 (0.849 µmol/L): 0.7% CV- Human Serum 1 (0.0872 µmol/L): 4.0% CV- Human Serum 2 (0.526 µmol/L): 0.8% CV- Human Serum 3 (0.889 µmol/L): 0.7% CV- Human Serum 4 (4.85 µmol/L): 1.0% CV- Human Serum 5 (44.2 µmol/L): 4.0% CV- Human Serum 6 (449 µmol/L): 3.7% CV- Human Serum 7 (1334 µmol/L): 3.1% CVIntermediate Precision:- Control 1 (0.0737 µmol/L): 10.9% CV- Control 2 (0.487 µmol/L): 1.2% CV- Control 3 (0.841 µmol/L): 0.8% CV- Human Serum 1 (0.0752 µmol/L): 11.2% CV- Human Serum 2 (0.526 µmol/L): 1.3% CV- Human Serum 3 (0.889 µmol/L): 1.1% CV- Human Serum 4 (4.91 µmol/L): 2.0% CV- Human Serum 5 (44.2 µmol/L): 5.3% CV- Human Serum 6 (449 µmol/L): 6.3% CV- Human Serum 7 (1316 µmol/L): 5.1% CVAll stated to have met acceptance criteria. |
| Analytical Sensitivity | LoB, LoD, LoQ within predefined thresholds to ensure accurate measurement at low concentrations. | LoB: ≤ 0.0250 µmol/L (claim in labeling)LoD: < 0.0350 µmol/L (claim in labeling)LoQ: 0.0400 µmol/L (claim in labeling)All stated to have met acceptance criteria. |
| Linearity/Reportable Range | Assay results must be linear across the claimed measuring range. | Confirmed for the measuring range of 0.0400 – 1.20 µmol/L.All stated to have met acceptance criteria. |
| Dilution | Expected recovery of concentration from diluted samples. | Post-dilution checks performed, supporting instrument and/or manual dilution for samples above the measuring range. (Implied acceptance criteria met, as no issues reported). |
| Endogenous Interferences | No significant interference from common endogenous substances at specified concentrations. | No interference claims for: Icterus (up to I index of 60/~1026 µmol/L bilirubin), Hemolysis (up to H index of 1000/~621 µmol/L hemoglobin), Lipemia (up to L index of 1000), Albumin (60 g/L), Immunoglobulin G (60 g/L), Total protein (2-12 g/dL), Rheumatoid factors (1000 IU/mL).All predefined acceptance criteria were met. |
| Analytical Specificity/ Cross-Reactivity | Expected cross-reactivity with structurally similar compounds. | DAMPA cross-reactivity: 87.6% at 0.1 µmol/L MTX, 64.6% at 1.0 µmol/L MTX.DAMPA can cross-react between 50% and 150% (This range implies the acceptance criteria for DAMPA cross-reactivity). |
| Exogenous Interferences (Drugs) | No significant interference from commonly used pharmaceuticals. | A long list of common and special pharmaceuticals were tested with no significant interference (implicitly meaning acceptance criteria were met). |
| Sample Matrix Comparison | Acceptable correlation between different sample matrices (serum vs. various plasma types). | Correlation (Pearson r) with Serum vs.:- Li-Heparin plasma: 0.998 - K2-EDTA plasma: 0.999 - K3-EDTA plasma: 0.998 - Na-Heparin plasma: 0.998 Slopes are close to 1, intercepts close to 0.All predefined acceptance criteria were met, supporting the claims for acceptable sample types. |
| Method Comparison | Acceptable correlation/agreement with a validated comparator method (LC-MS/MS). | Deming Regression: y = 1.032x + 0.000831 µmol/LCorrelation (r): 0.997(Implied acceptance criteria met, as close agreement shown). |
| Stability | Device must maintain performance throughout its claimed shelf life and on-board stability. | Stability studies were conducted to support Roche Diagnostic's claims as reported in the package labeling. (Implied acceptance criteria met). |
2. Sample Sizes Used for the Test Set and Data Provenance:
- Precision (Repeatability & Intermediate Precision):
- Repeatability: n = 84 measurements (total across all samples/controls).
- Intermediate Precision: Not explicitly stated as a single 'n'. It involved 2 aliquots per run, 2 runs per day, over 21 days for controls and human serum samples.
- Analytical Sensitivity (LoB, LoD, LoQ):
- LoB: One analyte-free sample measured with three reagent lots, in 6 runs, each with 10-fold determination, distributed over 3 days.
- LoD: 5 serum samples measured on three lots with 2-fold determination per run, across 6 runs distributed over 3 days.
- LoQ: 5 serum samples measured with three reagent lots, across 6 runs distributed over 3 days.
- Linearity/Assay Reportable Range:
- A dilution series spanning ≥ 9 levels, assayed on one analyzer using 3 reagent lots and 4 replicates per sample.
- Endogenous Interferences: Not explicitly stated as a numerical sample size but implies a study design for each substance.
- Analytical Specificity/Cross-Reactivity (DAMPA):
- Two human serum sample pools.
- Exogenous Interferences (Drugs): Not explicitly stated as a numerical sample size but implies a study design for each drug.
- Sample Matrix Comparison:
- ≥ 50 samples compared across serum and various plasma types.
- Method Comparison:
- 105 native human plasma and serum samples.
Data Provenance: The studies were conducted on "human serum" and "human plasma" samples. No specific country of origin is mentioned for the samples. The studies are retrospective in nature, as they involve testing existing samples (or creating spiked samples from existing matrices) within a laboratory setting to characterize the device performance. This is a typical approach for IVD device validation.
3. Number of Experts Used to Establish Ground Truth for the Test Set and Qualifications:
This information is not provided in the document. The type of device (in vitro diagnostic for quantitative determination of a drug) means that "ground truth" is typically established via a reference method (like LC-MS/MS), not by expert human graders.
4. Adjudication Method for the Test Set:
This information is not applicable for this type of in vitro diagnostic device study. Adjudication methods (like 2+1, 3+1) are typically used in clinical studies where human readers evaluate medical images and their interpretations need to be reconciled. For a quantitative assay like this, the 'ground truth' is established by a reference analytical method.
5. If a Multi-Reader Multi-Case (MRMC) Comparative Effectiveness Study was done and Effect Size:
No, a MRMC comparative effectiveness study was not done. This type of study is relevant for AI-powered devices that assist human interpretation (e.g., radiologists reading images). This device is a fully automated in vitro diagnostic assay, so human interpretation assistance is not part of its function.
6. If a Standalone (algorithm only without human-in-the-loop performance) was done:
Yes, this entire study describes the standalone performance of the "ONLINE TDM Methotrexate" assay. It is an automated laboratory test that measures methotrexate levels, meaning it operates "without human-in-the-loop performance" during the measurement process. The results are quantitative values generated directly by the instrument using the assay's reagents.
7. The Type of Ground Truth Used:
The primary type of "ground truth" used for performance evaluation, particularly for method comparison and potentially for linearity/dilution studies, is a validated comparator method, specifically LC-MS/MS (Liquid Chromatography-Mass Spectrometry/Mass Spectrometry). This is a highly accurate and precise analytical technique often considered a reference method for drug quantification. The document explicitly states: "The target value was determined with the LC-MS/MS" for dilution checks and "A method comparison of the ONLINE TDM Methotrexate on the cobas c 503 analyzer versus a validated comparator method on LC-MS/MS was completed."
For other tests like precision, sensitivity, interference, matrix comparison, ground truth is established by the known concentrations of controls, spiked samples, or comparison against established acceptable ranges as per CLSI (Clinical and Laboratory Standards Institute) guidelines.
8. The Sample Size for the Training Set:
The document does not provide any details about a training set. This is expected because this device is an in vitro diagnostic reagent assay, not an AI/Machine Learning model that undergoes a "training" phase. The assay relies on established biochemical reactions (homogeneous enzyme-immunoassay) and instrumental calibration rather than iterative learning from a dataset.
9. How the Ground Truth for the Training Set Was Established:
As there is no training set for this type of IVD device (it's not an AI/ML product), this question is not applicable. The "ground truth" for the performance evaluation (test set) relied on validated analytical methods and known sample concentrations.
Ask a specific question about this device
(166 days)
The ARK Methotrexate II Assay is a homogeneous enzyme immunoassay intended for the quantitative determination of methotrexate in human serum or plasma on automated clinical chemistry analyzers. The measurements obtained are used in monitoring levels of methotrexate to help ensure appropriate therapy. Specimens from patients who have received glucarpidase (carboxypeptidase G2) as a high dose methotrexate rescue therapy should not be tested with the ARK Methotrexate II Assay.
The ARK Methotrexate II Assay is a homogeneous immunoassay based on competition between drug in the specimen and methotrexate labeled with the recombinant enzyme glucose-6phosphate dehydrogenase (rG6PDH) for binding to the antibody reagent. As the latter binds antibody, enzyme activity decreases. In the presence of drug from the specimen, enzyme activity increases and is directly related to the drug concentration. Active enzyme converts the coenzyme nicotinamide adenine dinucleotide (NAD) to NADH that is measured spectrophotometrically as a rate of change in absorbance. Endogenous serum G6PDH does not interfere with the results because the coenyzme NAD functions only with the bacterial enzyme (rG6PDH) used in the assay. The ARK Methotrexate II Assay consists of reagents R1 anti-methotrexate monoclonal antibody with substrate and R2 methotrexate labeled with recombinant G6PDH enzyme. The test system includes the ARK Methotrexate II Calibrator, ARK Methotrexate II Control, and ARK Methotrexate II Dilution Buffer.
The ARK Methotrexate II Assay is a homogeneous enzyme immunoassay for the quantitative determination of methotrexate in human serum or plasma.
Here's an analysis of the acceptance criteria and the study proving the device meets them:
1. Table of Acceptance Criteria and Reported Device Performance:
| Acceptance Criteria | Device Performance (Reported) |
|---|---|
| Limit of Quantitation (LoQ) | Established as 0.030 umol/L. Acceptable inter-assay precision (<0.010 SD) and recovery (±0.010 umol/L) observed at 0.030 umol/L. |
| Measurement Range | 0.030 - 1.300 umol/L. |
| Recovery | All concentrations tested showed ±10% recovery of the expected sample concentration. |
| Linearity | A linear relationship was demonstrated between 0.030 and 1.300 umol/L with a maximum deviation of 8.73% from linearity. |
| Clinical Accuracy (vs. LC-MS/MS) | Slope of 1.03, intercept of 0.00, and Pearson correlation (r²) of 0.98. Mean bias of 0.01 with 95% Limits of Agreement from -0.11 to 0.14. |
| Method Comparison (vs. Predicate Device) | Slope = 0.98, y-intercept = -0.02, and correlation coefficient (r²) = 0.97. |
| Precision (Total CV) | ≤10% for all tested levels of controls and human serum samples. |
| Interference by Endogenous Substances | Elevated concentrations of common endogenous substances (albumin, bilirubin, cholesterol, etc.) did not interfere significantly (<±7.48% interference). |
| Analytical Specificity (7-OH-MTX) | Not substantially affected by 7-Hydroxymethotrexate (8.72% interference at 0.050 µmol/L MTX, 0.58% at 0.500 µmol/L MTX). |
| Analytical Specificity (DAMPA) | Significant interference with DAMPA (up to 57.50% cross-reactivity at 0.040 µmol/L DAMPA). Device should not be used during glucarpidase rescue therapy. |
| Analytical Specificity (Other Compounds) | Methotrexate-selective antibody did not cross-react with various potentially co-administered drugs and folate derivatives (all within ±10% interference). |
| Sample Stability | Stable for at least 14 days refrigerated (2-8 °C), 14 days at room temperature (25 °C), 15 months frozen (-20 °C), and after 3 freeze/thaw cycles. |
| Product Stability | Shelf-life stability claim of up to 18 months when stored unopened at 2-8°C. |
| On-Board Stability | Reagents stable up to 100 days on-board the instrument. |
| Calibration Curve Stability | Effective up to at least 100 days. |
2. Sample Size Used for the Test Set and Data Provenance:
- Clinical Accuracy (vs. LC-MS/MS): 90 patient samples.
- Method Comparison (vs. Predicate Device): 123 patient samples.
- LoQ, Recovery, Linearity, Precision, Interference, Analytical Specificity: These studies involved spiked human serum samples, pooled human serum, and controls, rather than a specific number of unique patient samples for the test set. For LoQ, 40 replicates were tested. For Recovery, 6 replicates per sample were tested. For Linearity, 6 replicates per sample were tested. For Precision, 160 replicates were tested for each of the 6 control levels and 6 human serum levels. For Interference and Analytical Specificity, 6 replicates per sample were tested.
- Data Provenance: "Leftover specimens were obtained from persons receiving high-dose methotrexate therapy" for the method comparison studies. The document does not specify the country of origin of the data or whether the studies were retrospective or prospective, though "leftover specimens" suggests a retrospective collection.
3. Number of Experts Used to Establish the Ground Truth for the Test Set and Qualifications of Those Experts:
- The ground truth for the clinical accuracy study was established by an LC-MS/MS reference method. This is an analytical laboratory technique, not based on expert consensus. Therefore, no "experts" in the traditional sense (e.g., radiologists) were used for ground truth establishment. The performance of LC-MS/MS as a reference method typically implies its results are considered the gold standard.
4. Adjudication Method for the Test Set:
- Not applicable. The ground truth for the clinical accuracy and method comparison studies was established using objective analytical methods (LC-MS/MS and the predicate device), not through expert review requiring an adjudication method.
5. If a Multi-Reader Multi-Case (MRMC) Comparative Effectiveness Study was done, If so, what was the effect size of how much human readers improve with AI vs without AI assistance:
- Not applicable. This device is an in vitro diagnostic (IVD) assay for quantitative determination of a substance in human samples. It does not involve human readers interpreting medical images or data.
6. If a Standalone (i.e. algorithm only without human-in-the-loop performance) was done:
- Yes, the performance data presented (Limit of Quantitation, Measurement Range, Recovery, Linearity, Clinical Accuracy, Method Comparison, Precision, Interference, Analytical Specificity) reflects the standalone performance of the ARK Methotrexate II Assay (an automated immunoassay) without human intervention in the result generation. The device itself is designed to provide quantitative measurements.
7. The Type of Ground Truth Used:
- Clinical Accuracy: Liquid Chromatography with Tandem Mass Spectrometry (LC-MS/MS) was used as the reference method (gold standard) for ground truth.
- Method Comparison: The legally marketed predicate device, ARK™ Methotrexate Assay (K111904), was used to compare results.
- Other performance metrics (LoQ, Recovery, Linearity, Precision, Interference, Analytical Specificity): Ground truth was established by preparing samples with known concentrations of methotrexate or interfering substances, typically by spiking human serum with certified reference materials.
8. The Sample Size for the Training Set:
- Not applicable. This device is an immunoassay (laboratory test kit), not a machine learning or AI algorithm that requires a "training set" in the conventional sense of computational models. The development and optimization of the assay would involve various experiments and reagent formulations, but not a distinct "training set" like for AI models.
9. How the Ground Truth for the Training Set Was Established:
- Not applicable, as there is no "training set" in the context of an AI/ML algorithm for this type of medical device. The "ground truth" for the development of the assay components would involve standard analytical chemistry practices, such as using reference materials and established analytical methods to verify the concentration and purity of substances used in the reagents.
Ask a specific question about this device
(261 days)
The ARK™ Methotrexate Assay is a homogeneous enzyme immunoassay intended for the quantitative determination of methotrexate in human serum or plasma on automated clinical chemistry analyzers. The measurements obtained are used in monitoring levels of methotrexate to help ensure appropriate therapy.
Specimens from patients who have received glucarpidase (carboxypeptidase G2) as a high dose methotrexate rescue therapy should not be tested with the ARK Methotrexate Assay.
The ARK Methotrexate Assay is a homogeneous immunoassay based on competition between drug in the specimen and Methotrexate labeled with the enzyme glucose-6-phosphate dehydrogenase (G6PDH) for binding to the antibody reagent. As the latter binds antibody, enzyme activity decreases. In the presence of drug from the specimen, enzyme activity increases and is directly proportional to the drug concentration. Active enzyme converts the coenzyme nicotinamide adenine dinucleotide (NAD) to NADH that is measured spectrophotometrically as a rate of change in absorbance. Endogenous serum G6PDH does not interfere with the results because the coenzyme NAD functions only with the bacterial enzyme used in the assay.
The ARK Methotrexate Assay consists of reagents R1 anti-Methotrexate polyclonal antibody with substrate and R2 Methotrexate labeled with bacterial G6PDH enzyme. The ARK Methotrexate Calibrator consists of a six-level set to calibrate the assay, and the ARK Methotrexate Control consists of a six-level set used for quality control of the assay (tri-level calibration range set and tri-level high range set). The ARK Methotrexate Dilution Buffer is equivalent to zero calibrator (Calibrator A).
Here's an analysis of the provided text to extract the acceptance criteria and study information:
Acceptance Criteria and Device Performance for ARK™ Methotrexate Assay
The provided document describes the acceptance criteria and the study that proves the ARK™ Methotrexate Assay, when used on the Beckman Coulter AU680 analyzer, meets these criteria. The study aims to demonstrate substantial equivalence to the assay's performance on the predicate device, the Roche/Hitachi 917 analyzer.
1. Table of Acceptance Criteria and Reported Device Performance
The document states: "The pre-determined Pass/Fail acceptance criteria were met for all of the above performance studies." However, the specific numerical acceptance criteria or performance values for each characteristic are not provided in the given text. It only lists the types of performance characteristics evaluated.
| Performance Characteristic | Acceptance Criteria (Not Detailed) | Reported Device Performance |
|---|---|---|
| Limit of Blank | Undisclosed Pass/Fail Criteria | Met Acceptance Criteria |
| Limit of Detection | Undisclosed Pass/Fail Criteria | Met Acceptance Criteria |
| Limit of Quantitation | Undisclosed Pass/Fail Criteria | Met Acceptance Criteria |
| Recovery | Undisclosed Pass/Fail Criteria | Met Acceptance Criteria |
| Linearity | Undisclosed Pass/Fail Criteria | Met Acceptance Criteria |
| Accuracy (Method Comparison) | Undisclosed Pass/Fail Criteria | Met Acceptance Criteria |
| Precision | Undisclosed Pass/Fail Criteria | Met Acceptance Criteria |
| Specificity | Undisclosed Pass/Fail Criteria | Met Acceptance Criteria |
| Cross-reactivity | Undisclosed Pass/Fail Criteria | Met Acceptance Criteria |
| Carry-over | Undisclosed Pass/Fail Criteria | Met Acceptance Criteria |
| Dilution Recovery | Undisclosed Pass/Fail Criteria | Met Acceptance Criteria |
| On-board Stability | Undisclosed Pass/Fail Criteria | Met Acceptance Criteria |
2. Sample Size Used for the Test Set and Data Provenance
The document does not specify the sample sizes used for the test set for any of the performance studies.
The document refers to "human serum or plasma" as the sample type. The country of origin of the data is not explicitly stated. The study is a "validation and verification" activity, implying it was conducted specifically for this submission, likely making it a prospective study for the purpose of device validation.
3. Number of Experts and Qualifications for Ground Truth
The concept of "experts" and their qualifications for establishing ground truth, as typically seen in image analysis or diagnostic interpretation, is not applicable to this type of in vitro diagnostic device (immunoassay) study. The ground truth for this device's performance characteristics is established through analytical methods and reference standards, not expert interpretation.
4. Adjudication Method for the Test Set
The concept of an "adjudication method" (e.g., 2+1, 3+1) is not applicable to this type of in vitro diagnostic device (immunoassay) study. Performance is assessed through quantitative measurements against established analytical criteria, not through expert consensus on diagnostic outcomes.
5. Multi-Reader Multi-Case (MRMC) Comparative Effectiveness Study
A Multi-Reader Multi-Case (MRMC) comparative effectiveness study is not applicable to this type of in vitro diagnostic device (immunoassay). This study design is typically used for imaging or diagnostic interpretation tasks where human readers' performance is being evaluated, often with and without AI assistance. This device is an automated assay, not an AI for human interpretive tasks.
6. Standalone (Algorithm Only) Performance Study
The study described is a standalone performance study for the algorithm (assay system) on the Beckman Coulter AU680 analyzer. The validation and verification activities listed (Limit of Blank, Limit of Detection, Accuracy, Precision, etc.) directly evaluate the performance of the device itself, without human interpretation in the loop. The purpose is to demonstrate that the assay system alone performs comparably to its predicate.
7. Type of Ground Truth Used
For an immunoassay like the ARK™ Methotrexate Assay, the "ground truth" for evaluating its performance characteristics (analytical accuracy, precision, linearity, etc.) would be established through:
- Reference materials/standards: Calibrators and controls with precisely known concentrations of methotrexate.
- Reference methods: Comparison with established, validated analytical methods (e.g., LC-MS/MS, or the predicate device's performance which was previously validated) for samples with unknown concentrations.
- Spiking studies: Adding known amounts of analyte to a sample and measuring recovery to assess accuracy.
The document implicitly refers to these as part of the "analytical performance studies" and "validation and verification activities."
8. Sample Size for the Training Set
The document does not mention a training set or its sample size. This is expected as the ARK™ Methotrexate Assay is a homogeneous enzyme immunoassay kit, not a machine learning or AI model that requires a "training set" in the conventional sense. The "training" for such a system would involve validating its components and establishing its operational parameters on the target analyzer.
9. How Ground Truth for the Training Set Was Established
Since there is no "training set" in the context of machine learning, this question is not applicable. The assay is a chemical and biological system where parameters are set based on chemical principles and reagent characteristics, and then validated through performance studies.
Ask a specific question about this device
(105 days)
The ARK™ Methotrexate Assay is a homogeneous enzyme immunoassay intended for the quantitative determination of Methotrexate in human serum or automated clinical chemistry analyzers. The measurements obtained are used in monitoring levels of methotrexate to help ensure appropriate therapy.
The ARK™ Methotrexate Calibrator is intended for use in calibration of the ARK Methotrexate Assay.
The ARKTM Methotrexate Control is intended for use in quality control of the ARK Methotrexate Assay.
Specimens from patients who have received glucarpidase (carboxypeptidase G2) as a high dose methotrexate rescue therapy should not be tested with the ARK Methotrexate Assay.
The ARK Methotrexate Assay is a homogeneous immunoassay based on competition between drug in the specimen and Methotrexate labeled with the enzyme glucose-6-phosphate dehydrogenase (G6PDH) for binding to the antibody reagent. As the latter binds antibody, enzyme activity decreases. In the presence of drug from the specimen, enzyme activity increases and is directly proportional to the drug concentration. Active enzyme converts the coenzyme nicotinamide adenine dinucleotide (NAD) to NADH that is measured spectrophotometrically as a rate of change in absorbance. Endogenous serum G6PDH does not interfere with the results because the coenyzme NAD functions only with the bacterial enzyme used in the assay.
The ARK Methotrexate Assay consists of reagents R1 anti-Methotrexate polyclonal antibody with substrate and R2 Methotrexate labeled with bacterial G6PDH enzyme. The ARK Methotrexate Calibrator consists of a six-level set to calibrate the assay, and the ARK Methotrexate Control consists of a six-level set used for quality control of the assay (tri-level calibration range set and tri-level high range set). The ARK Methotrexate Dilution Buffer is equivalent to zero calibrator (Calibrator A).
Here's a breakdown of the acceptance criteria and the studies that prove the device meets them, based on the provided text:
Acceptance Criteria and Reported Device Performance
| Acceptance Criteria Category | Specific Acceptance Criteria | Reported Device Performance |
|---|---|---|
| Limit of Quantitation (LoQ) | LoB < Result < LoQ: report "analyte detected; concentration < LoQ" Result ≥ LoQ: report the result as measured | LoB: 0.01 µmol/L LoD: 0.02 µmol/L LoQ: 0.04 µmol/L (LoQ-2 SD > LoD met) |
| Accuracy (Analytical Recovery) | Not explicitly stated as a numerical criterion in the "Acceptance Criteria" section for each data point, but typically implied to be close to 100% recovery. | Mean percentage recovery: 102.1% (Individual recoveries ranged from 98.3% to 111.1%) |
| Linearity | Percent difference was ±10% between the predicted 1st and 2nd order regressed values for concentrations >0.10 µmol/L or ±0.01 µmol/L at concentrations ≤ 0.10 µmol/L. | All observed differences for concentrations >0.10 µmol/L were within ±10% (-2.3% to 4.8%).All observed differences for concentrations ≤ 0.10 µmol/L were within ±0.01 µmol/L (-0.010 µmol/L to -0.004 µmol/L). The highest theoretical concentration tested was 1.30 µmol/L. Regression of assayed methotrexate concentrations was linear throughout the range of 2 to 1200 µmol/L when proportionally diluted. |
| Assay Range | Not explicitly stated as an acceptance criterion, but the functional range. | Measurement range: 0.04 - 1.20 µmol/L |
| Method Comparison | Not explicitly stated as a numerical acceptance criterion (e.g., minimum correlation coefficient or confidence intervals for slope/intercept), but comparison to a predicate device is expected. | Range 0.04 to 1.19 µM (102 samples): Slope: 1.00 (95% CI: 1.00 to 1.02) y-intercept: 0.01 (95% CI: 0.00 to 0.01) Correlation Coefficient (r²): 0.978 (95% CI: 0.968 to 0.985)Range 0.04 to 1440 µM (147 samples, including diluted): Slope: 0.99 (95% CI: 0.96 to 1.00) y-intercept: 0.01 (95% CI: 0.01 to 0.01) Correlation Coefficient (r²): 0.998 (95% CI: 0.997 to 0.998) |
| Precision | <10% total CV at >0.10 µmol/L SD ≤0.01 at ≤0.10 µmol/L | All control and patient pool samples met the criteria: - ARK Methotrexate Control (MID, HIGH, 5, 50, 500 µmol/L): Total CVs ranged from 3.8% to 7.0%. - ARK Methotrexate Control (LOW, 0.06 µmol/L): Total SD was 0.007, which is ≤0.01. - Patient Pool (MID, HIGH, 5, 50, 500 µmol/L): Total CVs ranged from 5.3% to 7.2%. - Patient Pool (LOW, 0.07 µmol/L): Total SD was 0.008, which is ≤0.01. |
| Interfering Substances | Not substantially affected by tested endogenous substances. | Measurement of methotrexate was not substantially affected by clinically high concentrations of Albumin, Bilirubin (conjugated & unconjugated), Cholesterol, Gamma-Globulin, Hemoglobin, Intralipid®, Rheumatoid Factor, Triglycerides, and Uric Acid. |
| Specificity - Crossreactivity | < 0.07% crossreactivity with 7-Hydroxymethotrexate. | 7-Hydroxymethotrexate: ≤ 0.07% crossreactivity. DAMPA: 64.3% to 100% crossreactivity (significant). Triamterene: Crossreactivity ranged from 1.85% to 3.32%. Trimethoprim: Crossreactivity ranged from 0.12% to 0.54%. Folate analogs and other compounds (e.g., Adriamycin, Cyclophosphamide): ≤ 0.01% crossreactivity at ≥ 1000 µmol/L. |
| Anticoagulants | No significant difference in recovery between serum and plasma. | No significant difference found in methotrexate recovery between serum and plasma samples. |
| Sample Stability | Acceptable stability under various storage conditions. | Stable frozen (at least 15 months), 48 hours at room temperature, refrigerated (at least 21 days), and after three (3) successive freeze/thaw cycles. |
| On-Board Stability | Acceptable stability of calibration curve and reagents on the analyzer. | Calibration curve effective up to at least 19 days. Reagents effective for up to at least 25 days in analyzer-specific containers. In-use stability of calibrator and controls also demonstrated. |
Study Details
2. Sample sizes used for the test set and the data provenance (e.g. country of origin of the data, retrospective or prospective):
- Limit of Quantitation (LoQ):
- LoB, LoD: N = 60
- LoQ: N = 40
- Data provenance: Not specified, but generally analytical studies like this are conducted with prepared samples in a laboratory setting (prospective).
- Accuracy: Concentrated methotrexate drug added into human serum negative for methotrexate. Six replicates of each sample were assayed.
- Sample size: 5 target concentrations, each with 6 replicates (30 assays).
- Data provenance: Not specified, but likely prospective, prepared samples in a lab.
- Linearity: Dilutions of a 1.30 µmol/L serum sample and proportional dilutions of samples between 2 and 1200 µmol/L in pooled human serum.
- Sample size: 13 different concentrations were tested for the initial linearity study. The second linearity study for higher concentrations (2-1200 µmol/L) stated "samples containing methotrexate... were prepared proportionally," implying multiple samples across this range.
- Data provenance: Not specified, but likely prospective, prepared samples in a lab.
- Method Comparison:
- Sample size: 102 specimens within the measurement range (0.04 to 1.19 µM) and 147 specimens including those above the measurement range requiring dilution (total range 0.04 to 1440 µM).
- Data provenance: Not specified, but these are likely clinical samples (human serum/plasma), which could be retrospective or prospective. The document does not specify country of origin.
- Precision:
- Sample size: Each of the 6 control levels and 6 patient pool levels were assayed in quadruplicate, twice a day for 20 days. This means 4 * 2 * 20 = 160 measurements per level.
- Data provenance: Not specified, but likely prospective, prepared controls and pooled patient samples in a lab.
- Interfering Substances: Multiple substances tested, each with samples at ~0.05 and ~0.50 µmol/L methotrexate.
- Sample size: Each interfering substance was tested at two methotrexate levels, likely with duplicates or triplicates, plus control samples. The specific number of individual assays is not given but implies a significant number of tests.
- Data provenance: Not specified, but likely prospective, prepared samples in a lab setting.
- Specificity (Crossreactivity):
- Sample size: Two concentrations of methotrexate (0.05 and 0.50 µmol/L) were tested with triamterene and trimethoprim. Various other compounds and folate analogs were tested at high concentrations (e.g., ≥ 1000 µmol/L), presumably just once for cross-reactivity.
- Data provenance: Not specified, but likely prospective, prepared samples in a lab setting.
- Anticoagulants:
- Sample size: "Studies were conducted," implying multiple samples, but no specific number is given.
- Data provenance: Not specified, but likely prospective, prepared samples in a lab setting.
- Sample Stability:
- Sample size: Not specified, but involved human specimens.
- Data provenance: Not specified, but likely prospective.
- On-Board Stability:
- Sample size: Not specified, but utilized a calibration curve and reagents over time.
- Data provenance: Not specified, but likely prospective, lab-based testing.
3. Number of experts used to establish the ground truth for the test set and the qualifications of those experts (e.g. radiologist with 10 years of experience):
This device is an in vitro diagnostic assay for quantitative determination of methotrexate. The "ground truth" for such assays is typically established by:
- Reference materials: Certified reference standards for methotrexate concentration (used in Accuracy, LoQ, Linearity, Crossreactivity).
- Predicate device comparison: The "gold standard" or accepted method (Fluorescence Polarization Immunoassay - FPIA in this case, specifically the Abbott TDx®/TDxFLx® Methotrexate II assay) for method comparison.
Therefore, the concept of "experts" establishing ground truth as it applies to image interpretation (like radiologists) or pathological diagnosis is not directly relevant here. The ground truth is analytical and based on established methodology and certified materials.
4. Adjudication method (e.g., 2+1, 3+1, none) for the test set:
Not applicable. Adjudication methods like 2+1 or 3+1 are typically used in clinical studies where subjective human interpretation (e.g., reading medical images) is involved and discrepancies between readers need to be resolved to establish a consensus ground truth. For in vitro diagnostic device performance studies, the results are quantitative and compared against analytical or predicate methods, not subjective expert consensus.
5. If a multi-reader multi-case (MRMC) comparative effectiveness study was done, If so, what was the effect size of how much human readers improve with AI vs without AI assistance:
Not applicable. This device is an in vitro diagnostic assay, not an AI-powered image analysis tool or a device requiring human "readers" in the context of an MRMC study. Its performance is evaluated analytically, not by assessing human performance improvement with or without AI.
6. If a standalone (i.e. algorithm only without human-in-the-loop performance) was done:
Yes, all the performance studies described (analytical recovery, linearity, method comparison, precision, interference, specificity, stability) represent the standalone performance of the ARK Methotrexate Assay. There is no human-in-the-loop aspect described in these tests; the assay directly provides quantitative results.
7. The type of ground truth used (expert consensus, pathology, outcomes data, etc):
The ground truth used for this in vitro diagnostic assay consists of:
- Reference Standards: For accuracy, LoQ, linearity, and specificity, the ground truth was based on precisely known concentrations of methotrexate prepared using certified stock concentrates.
- Predicate Device (Monoclonal FPIA): For method comparison, the ground truth was the results obtained from the legally marketed predicate device (Abbott TDx®/TDxFLx® Methotrexate II), which is an established method for methotrexate measurement.
- Pre-defined Parameters/Conditions: For interference, specificity, and stability studies, the ground truth was the expectation of what the device should measure under specific well-defined conditions (e.g., no cross-reactivity with certain compounds, stability over a given period).
8. The sample size for the training set:
Not applicable. This document describes the validation of an immunoassay kit, not a machine learning or AI algorithm that requires a "training set." Immunoassays are based on biochemical reactions with antibodies, not pattern recognition learned from data.
9. How the ground truth for the training set was established:
Not applicable, as there is no "training set" in the context of this immunoassay device.
Ask a specific question about this device
Page 1 of 1