(328 days)
The Technozvm ADAMTS13 Activity assay is an enzyme-linked immunosorbent assay (ELISA) intended for the qualitative determination of ADAMTS13 activity in platelet poor human citrated plasma. The assay is intended to be used in conjunction with other clinical and laboratory findings as an aid in the diagnosis of thrombotic thrombocytopenic purpura (TTP) in adult and pediatric patients being evaluated for thrombotic microangiopathy (TMA).
The Technozym ADAMTS13 Activity assay is an enzyme linked immunosorbent assay (ELISA) used for detection of ADAMTS13 activity in citrated human plasma. The assay contains: ADAMTS13 Activity anti-GST coated test plate microplate coated with anti-GST antibody, ADAMTS13 Activity GST-VWF73 reagent that contains GST tagged peptide of 73 amino acids from the A2 domain of VWF with specific cleavage site for ADAMTS13 and serves as the in vitro substrate for ADAMTS13, ADAMTS13 Activity Calibrators-consists of six vials containing lyophilized plasma, each with a different level of ADAMTS13 activity, ADAMTS13 Activity Controls consists of two vials of lyophilized plasma, each with high or low levels of ADAMTS13 activity, ADAMTS13 Activity Conjugate - reagent that contains horseradish peroxidase (HRP) conjugated monoclonal antibody directed against the neoepitope exposed due to cleavage of GST-VWF73 by ADAMTS13 present in plasma, ADAMTS13 TMB substrate reagent contains tetramethylbenzidine (TMB) substrate for HRP, ADAMTS13 Activity Stop Solution reagent contains 2.5% sulfuric acid for stopping the conversion of TMB substrate.
The Technozym ADAMTS13 Activity assay is a manual enzyme-linked immunosorbent assay (ELISA) intended for the qualitative determination of ADAMTS13 activity in platelet-poor human citrated plasma. It is used in conjunction with other clinical and laboratory findings as an aid in the diagnosis of thrombotic thrombocytopenic purpura (TTP) in adult and pediatric patients being evaluated for thrombotic microangiopathy (TMA). The assay's clinical cutoff for TTP diagnosis is 0.1 IU/mL ADAMTS13 activity.
Acceptance Criteria and Device Performance
The primary acceptance criteria for this device appear to be related to its analytical performance (precision, specificity), and its clinical performance (sensitivity and specificity for TTP diagnosis).
Here's a table summarizing the reported device performance, which implicitly serves as the acceptance criteria based on the successful De Novo grant:
| Performance Metric | Acceptance Criteria (Effectively Met) | Reported Device Performance (Mean %CV / % Correct Call / Sensitivity/Specificity) | Study Type |
|---|---|---|---|
| Within-laboratory Precision | Quantitative: Low %CV | S1-S9: 4.52% - 9.26% Within-laboratory %CV | Within-laboratory Precision Study |
| Qualitative: High % Correct Call | S1-S9: 100% correct call (at cutoff) | Within-laboratory Precision Study | |
| Operator-to-operator Precision | Quantitative: Low %CV | S1-S9: 5.59% - 10.08% Within-laboratory %CV | Operator-to-operator Precision Study |
| Qualitative: High % Correct Call | S1-S9: 100% correct call (at cutoff) | Operator-to-operator Precision Study | |
| Site-to-site Reproducibility | Quantitative: Low %CV | S1-S9: 8.01% - 10.45% Reproducibility %CV | Site-to-site Reproducibility Study |
| Qualitative: High % Correct Call | S1-S9: 100% correct call (at cutoff) | Site-to-site Reproducibility Study | |
| Analytical Specificity/Interference | No clinically significant interference | None of 25 tested substances led to clinically significant interference | Interference Study |
| Clinical Sensitivity (TTP) | High Sensitivity | 84.8% (95% CI: 69.1% to 93.3%) | Clinical Performance Study |
| Clinical Specificity (TTP) | High Specificity | 97.1% (95% CI: 91.9% to 99%) | Clinical Performance Study |
| Positive Predictive Value (TTP) | High PPV | 90.2% (95% CI: 75.2% to 96.6%) | Clinical Performance Study |
| Negative Predictive Value (TTP) | High NPV | 95.3% (95% CI: 90.0% to 97.8%) | Clinical Performance Study |
| Prozone Effect (Hook Effect) | No significant hook effect | No significant hook effect up to 8 IU/mL | Prozone Effect Study |
| Cross-contamination | No cross-contamination | No cross-contamination observed | Cross-contamination Study |
| Reagent Shelf-life | Adequate Shelf-life | 24 months at 2-8°C | Real-time Shelf-life Stability Study |
| Frozen Sample Stability | Adequate Stability | 12 months at < -20°C | Frozen Sample Stability Study |
| Fresh Sample Stability | Adequate Stability | 8 hours at 18-25°C, 24 hours at 2-8°C | Fresh Sample Stability Study |
Study Details:
-
Sample sizes used for the test set and the data provenance:
- Precision/Reproducibility Studies: For each of the nine samples (S1-S9), 30 replicate measurements were performed. This involved human plasma sample pools, prepared by mixing human plasma from normal donors with clinical samples from TTP patients or heat-inactivated plasma. The origin of the normal donors and clinical samples is not explicitly specified beyond being "human plasma." These appear to be laboratory-prepared samples, not directly from patient cohorts.
- Analytical Specificity/Interference Study: Three base pools (mimicking high, medium, and low ADAMTS13 activity) were used. Each interfering substance was tested with five replicates per sample. The samples were human citrated plasma.
- Clinical Performance Study: A total of 137 samples were included. These were residual samples selected from a local repository of frozen human citrated plasma from patients diagnosed with thrombotic microangiopathies (TMA), with clinical suspicion of TTP. The study was conducted at two external sites, one in the U.S. and one outside the U.S. This indicates a multi-site validation. The study design ("residual samples from a local repository of frozen human citrated plasma") strongly suggests a retrospective approach.
- Prozone Effect, Cross-contamination, Reagent Stability, Sample Stability: Sample sizes vary for these specific studies but are generally sufficient for the analytical evaluations (e.g., 8 samples for reagent stability, 6-8 samples for sample stability, various low/high samples for cross-contamination).
-
Number of experts used to establish the ground truth for the test set and the qualifications of those experts:
- For the Clinical Performance Study, the ground truth for TTP diagnosis was established by a "board-certified clinician" according to the "local testing algorithm for TMAs." The exact number of clinicians involved is not specified, but it implies at least one per site or a consensus from a group. Their qualification as "board-certified clinician" indicates expertise in diagnosing TMAs.
- For the Precision/Reproducibility and Analytical Specificity/Interference studies, the ground truth for sample concentrations (e.g., "normal donors," "TTP patient plasma," "heat inactivated plasma") was established by how the samples were prepared or selected based on known clinical status. These are analytical ground truths, not clinical diagnoses requiring expert adjudication of individual cases.
-
Adjudication method (e.g., 2+1, 3+1, none) for the test set:
- For the Clinical Performance Study, the ground truth was established by "board-certified clinician" according to "local testing algorithm for TMAs." There is no explicit mention of an adjudication method like 2+1 or 3+1 for resolving discrepancies in the clinical diagnosis or establishing the final ground truth from multiple experts. The clinical diagnosis was the established baseline for comparison.
-
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:
- This is an in-vitro diagnostic (IVD) assay, not an AI-assisted diagnostic device that involves human readers interpreting images or complex data aided by AI. Therefore, an MRMC comparative effectiveness study, as typically understood in medical imaging AI, was not performed, nor is it applicable. The study evaluates the performance of the assay itself in providing a quantitative measurement (ADAMTS13 activity) leading to a qualitative TTP diagnosis (positive/negative). Human involvement is in performing the ELISA steps and interpreting the quantitative result against a defined cutoff, not in interpreting complex visual data.
-
If a standalone (i.e. algorithm only without human-in-the-loop performance) was done:
- The Technozym ADAMTS13 Activity assay is a manual ELISA kit, not an algorithm or AI system. Its performance is evaluated based on its accuracy in producing a quantitative result that is then interpreted qualitatively based on a predefined cutoff. Therefore, the concept of "standalone algorithm performance" (without human-in-the-loop) is not applicable in the context of this device. The device's performance metrics (precision, sensitivity, specificity) reflect its inherent analytical capabilities when properly executed.
-
The type of ground truth used (expert consensus, pathology, outcomes data, etc.):
- For the Clinical Performance Study, the ground truth for TTP diagnosis was based on the "clinical diagnosis of TTP" by "board-certified clinician(s)" according to "local testing algorithm for TMAs." This implies a combination of clinical findings, laboratory results, and potentially outcomes, but the primary reference is the established clinical diagnosis. It's akin to "expert clinical diagnosis" as the ground truth.
- For the Analytical Performance studies (Precision, Interference), the ground truth was based on known concentrations or characteristics of the prepared samples (e.g., specific concentrations, the presence/absence of interfering substances, known ADAMTS13 deficiency from TTP patient plasma or heat inactivation).
-
The sample size for the training set:
- This document describes the performance evaluation of a diagnostic assay kit, not a machine learning or AI model. Therefore, there is no "training set" in the context of developing a data-driven model. The assay's "training" or development would involve biochemical and reagent optimization, calibration curve establishment, and analytical validation steps, not a distinct data-driven training set as understood in AI/ML.
-
How the ground truth for the training set was established:
- As there is no "training set" for a machine learning model, this question is not applicable. The "ground truth" during the development of the assay would correspond to reference materials, known concentrations, and established clinical samples used for initial assay development and calibration. The calibrators themselves are traceable to the first International Standard for ADAMTS13 Activity and Antigen in Plasma (NIBSC WHO 1st international Standard ADAMTS13 Plasma 12/252), establishing a form of objective analytical ground truth for calibration.
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EVALUATION OF AUTOMATIC CLASS III DESIGNATION FOR Technozym ADAMTS13 Activity DECISION SUMMARY
Background Information: I
A De Novo Number
B Applicant
Technoclone Herstellung von Diagnostika und Arzneimitteln GmbH
C Proprietary and Established Names
Technozym ADAMTS13 Activity
D Regulatory Information
| ProductCode(s) | Classification | RegulationSection | Panel |
|---|---|---|---|
| SAC | Class II withspecial controls | 21 CFR 864.7297 | Hematology |
Submission/Device Overview: II
A Purpose for Submission:
De Novo request for evaluation of automatic class III designation for Technozym ADAMTS13 Activity
B Measurand:
ADAMTS13 Activity
C Type of Test:
Manual enzyme linked immunosorbent assay (ELISA)
Food and Drug Administration 10903 New Hampshire Avenue Silver Spring, MD 20993-0002 www.fda.gov
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III Indications for Use:
A Intended Use(s):
See Indications for Use below
B Indication(s) for Use:
The Technozvm ADAMTS13 Activity assay is an enzyme-linked immunosorbent assay (ELISA) intended for the qualitative determination of ADAMTS13 activity in platelet poor human citrated plasma. The assay is intended to be used in conjunction with other clinical and laboratory findings as an aid in the diagnosis of thrombotic thrombocytopenic purpura (TTP) in adult and pediatric patients being evaluated for thrombotic microangiopathy (TMA).
C Special Conditions for Use Statement(s):
For Prescription Use Only For In Vitro Diagnostic Use Only
D Special Instrument Requirements:
Microplate reader
IV Device/System Characteristics:
A Device Description:
The Technozym ADAMTS13 Activity assay is an enzyme linked immunosorbent assay (ELISA) used for detection of ADAMTS13 activity in citrated human plasma.
The assay contains:
- ADAMTS13 Activity anti-GST coated test plate microplate coated with anti-GST antibody .
- ADAMTS13 Activity GST-VWF73 reagent that contains GST tagged peptide of 73 amino . acids from the A2 domain of VWF with specific cleavage site for ADAMTS13 and serves as the in vitro substrate for ADAMTS13
- ADAMTS13 Activity Calibrators-consists of six vials containing lyophilized plasma, each . with a different level of ADAMTS13 activity
- ADAMTS13 Activity Controls consists of two vials of lyophilized plasma, each with high . or low levels of ADAMTS13 activity
- . ADAMTS13 Activity Conjugate - reagent that contains horseradish peroxidase (HRP) conjugated monoclonal antibody directed against the neoepitope exposed due to cleavage of GST-VWF73 by ADAMTS13 present in plasma
- ADAMTS13 TMB substrate reagent contains tetramethylbenzidine (TMB) substrate for . HRP
- ADAMTS13 Activity Stop Solution reagent contains 2.5% sulfuric acid for stopping the . conversion of TMB substrate
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B Test Principle
ADAMTS13, a disintegrin and metalloprotease with thrombospondin type 1 motif 13, is an enzyme (VWF-cleaving protease) that specifically cleaves yon Willebrand factor (VWF) under high shear stress conditions. The Technozym ADAMTS13 Activity assay is an enzyme linked immunosorbent assay for the detection of ADAMTS13 activity in human citrated plasma. GST-VWF73, a substrate that can be specifically cleaved by ADAMTS13 in vitro, is immobilized on to wells of a microplate that is pre-coated with an antibody specific to glutathione S-transferase (GST). After washing away unbound GST-VWF73, samples (i.e., clinical specimens, controls, and calibrators) are pipetted into wells and incubated with immobilized GST-VWF73. ADAMTS13 present in the samples cleaves the VWF73 peptide of immobilized GST-VWF73 at specific sites, exposing the neoepitope on VWF73. After washing away the excess sample, a second mouse monoclonal antibody specific to the neoepitope on GST-VWF73 that has been conjugated to the enzyme horseradish peroxidase (HRP) is added to the well. After washing away unbound HRP-conjugated antibody, the chromogenic substrate is added to the well. The HRP enzyme catalyzes a specific reaction with the chromogenic substrate, which produces a colored product that is detected as absorbance measurement (optical density, OD) at 450 nm with a microplate reader. The amount of absorbance (OD) generated is proportional to ADAMTS13 activity in the well. The results for the wells containing calibrators are used to create a reference curve to quantify the ADAMTS13 activity in the sample.
In line with the recommendation of the International Society of Thrombosis and Haemostasis (ISTH) in the Journal of Thrombosis and Haemostasis (2020), the assay results should be interpreted at the ADAMTS13 Activity assay cut-off of 0.1 IU/mL for thrombotic thrombocvtopenic purpura (TTP). Technozym ADAMTS13 Activity assay results > 0.1 IU/mL will be TTP negative and results ≤ 0.1 IU/mL will be TTP positive. The ADAMTS13 Activity assay results should be interpreted in conjunction with other clinical and laboratory findings.
V Standards/Guidance Documents Referenced:
CLSI EP05-A3: Evaluation of Precision of Quantitative Measurement Procedures: Approved Guideline - Third Edition
CLSI EP06-A2: Evaluation of the Linearity of Quantitative Measurement Procedures: A Statistical Approach; Approved Guideline - Second Edition
CLSI EP07-A2: Interference Testing in Clinical Chemistry; Approved Guideline - Second Edition
CLSI EP17-A2: Evaluation of Detection Capability for Clinical Laboratory Measurement Procedures: Approved Guideline - Second Edition
CLSI EP25-A: Evaluation of Stability of In Vitro Diagnostic Reagents; Approved Guideline
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Performance Characteristics: VI
Analytical Performance:
1. Precision/Reproducibility:
Precision studies were conducted according to recommendations in CLSI EP05-A3 using quality controls and nine human plasma sample pools, which were prepared by mixing human plasma from normal donors with clinical samples from patients diagnosed with thrombotic thrombocytopenic purpura (TTP) and deficient in ADAMTS13 activity or heat inactivated plasma.
Within-laboratory precision
To evaluate the within-laboratory precision, each sample was tested for five days with two runs per day and two replicates per run at a single site, using three reagent lots for a total of 30 replicate measurements per sample. The samples tested included levels below, around and above the assay cut-off of 0.1 IU/mL. The quantitative results are summarized in the tables below.
| Sample | N | Mean(IU/mL) | Repeatability | Between-run | Between-day | Between-lot | Within-laboratory | |||||
|---|---|---|---|---|---|---|---|---|---|---|---|---|
| SD | %CV | SD | %CV | SD | %CV | SD | %CV | SD | %CV | |||
| S1 | 30 | 0.65 | 0.03 | 4.46 | 0.03 | 5.10 | 0.02 | 3.60 | 0.00 | 0.00 | 0.05 | 7.66 |
| S2 | 30 | 0.45 | 0.01 | 2.26 | 0.04 | 8.50 | 0.00 | 0.00 | 0.01 | 3.00 | 0.04 | 9.26 |
| S3 | 29 | 0.24 | 0.01 | 3.25 | 0.01 | 2.50 | 0.00 | 1.00 | 0.00 | 1.70 | 0.01 | 4.52 |
| S4 | 29 | 0.19 | 0.00 | 1.84 | 0.01 | 4.20 | 0.00 | 0.00 | 0.00 | 2.10 | 0.01 | 4.97 |
| S5 | 30 | 0.14 | 0.00 | 2.19 | 0.01 | 4.90 | 0.00 | 0.00 | 0.00 | 0.00 | 0.01 | 5.17 |
| S6 | 30 | 0.08 | 0.00 | 2.05 | 0.01 | 8.10 | 0.00 | 0.00 | 0.00 | 0.00 | 0.01 | 7.87 |
| S7 | 30 | 0.65 | 0.03 | 4.88 | 0.03 | 3.80 | 0.02 | 2.90 | 0.01 | 0.90 | 0.05 | 6.90 |
| S8 | 30 | 0.23 | 0.01 | 3.04 | 0.01 | 5.40 | 0.00 | 0.00 | 0.00 | 0.80 | 0.01 | 6.24 |
| S9 | 30 | 0.12 | 0.00 | 2.62 | 0.01 | 4.10 | 0.00 | 0.00 | 0.00 | 0.00 | 0.01 | 4.91 |
| Sample | Mean(IU/mL) | Totalresults | Qualitative agreement | |
|---|---|---|---|---|
| Number of correctresults | % Correct call | |||
| S1negative | 0.65 | 30 | 30/30 | 100 |
| S2negative | 0.45 | 30 | 30/30 | 100 |
| S3negative | 0.24 | 29 | 29/29 | 100 |
| S4negative | 0.19 | 29 | 29/29 | 100 |
| S5negative | 0.14 | 30 | 30/30 | 100 |
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| Sample | Mean(IU/mL) | Totalresults | Qualitative agreement | |
|---|---|---|---|---|
| Number of correctresults | % Correct call | |||
| S6positive | 0.08 | 30 | 30/30 | 100 |
| S7negative | 0.65 | 30 | 30/30 | 100 |
| S8negative | 0.23 | 30 | 30/30 | 100 |
| S9negative | 0.12 | 30 | 30/30 | 100 |
Operator-to-operator
The study was conducted over five days using one reagent lot with two runs per day and two replicates per run by three operators for a total of 30 mean results per sample level. The study design included six samples prepared by mixing plasma from normal human donors with native deficient plasma (TTP patient plasma) in different ratios. In addition, three sample levels were prepared by mixing plasma from normal human donors with heat inactivated plasma. The samples tested included levels below, around and above the assay cut-off of 0.1 IU/mL.
| Sample | N | Mean (IU/mL) | Repeatability | Between-run | Between-day | Between-operator | Within-laboratory | |||||
|---|---|---|---|---|---|---|---|---|---|---|---|---|
| SD | %CV | SD | %CV | SD | %CV | SD | %CV | SD | %CV | |||
| S1 | 30 | 0.67 | 0.03 | 4.17 | 0.03 | 5.10 | 0.03 | 4.80 | 0.01 | 1.90 | 0.06 | 8.27 |
| S2 | 30 | 0.45 | 0.01 | 2.26 | 0.04 | 9.70 | 0.00 | 0.00 | 0.01 | 1.30 | 0.05 | 10.08 |
| S3 | 29 | 0.24 | 0.01 | 2.90 | 0.01 | 4.30 | 0.00 | 0.00 | 0.01 | 2.30 | 0.01 | 5.59 |
| S4 | 29 | 0.19 | 0.00 | 2.19 | 0.01 | 5.40 | 0.00 | 0.00 | 0.00 | 2.20 | 0.01 | 6.10 |
| S5 | 30 | 0.13 | 0.00 | 1.77 | 0.01 | 3.60 | 0.01 | 3.40 | 0.00 | 2.50 | 0.01 | 5.99 |
| S6 | 30 | 0.07 | 0.00 | 2.69 | 0.00 | 6.00 | 0.00 | 0.00 | 0.00 | 4.10 | 0.01 | 8.12 |
| S7 | 30 | 0.65 | 0.03 | 4.81 | 0.03 | 4.80 | 0.00 | 0.00 | 0.01 | 1.80 | 0.05 | 7.11 |
| S8 | 30 | 0.23 | 0.01 | 2.26 | 0.01 | 4.90 | 0.00 | 1.70 | 0.00 | 1.60 | 0.01 | 5.84 |
| S9 | 30 | 0.12 | 0.00 | 2.05 | 0.01 | 5.20 | 0.00 | 1.20 | 0.00 | 2.40 | 0.01 | 6.18 |
| Sample | Mean(IU/mL) | Totalresults | Qualitative agreement | |
|---|---|---|---|---|
| Number of correctresults | % Correct call | |||
| S1negative | 0.67 | 30 | 30/30 | 100 |
| S2negative | 0.45 | 30 | 30/30 | 100 |
| S3negative | 0.24 | 29 | 29/29 | 100 |
| S4negative | 0.19 | 29 | 29/29 | 100 |
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| Sample | Mean(IU/mL) | Totalresults | Qualitative agreement | |
|---|---|---|---|---|
| Number of correctresults | % Correct call | |||
| S5negative | 0.13 | 30 | 30/30 | 100 |
| S6positive | 0.07 | 30 | 30/30 | 100 |
| S7negative | 0.65 | 30 | 30/30 | 100 |
| S8negative | 0.23 | 30 | 30/30 | 100 |
| S9negative | 0.12 | 30 | 30/30 | 100 |
Site-to-site reproducibility
The study was performed at three study sites. At each site, the samples were assayed on each of five days, with two runs per day and two replicates per run, using one lot of reagents, resulting in a total of 30 mean results per sample level. The study design included six sample levels prepared by mixing plasma from normal human donors with native deficient plasma (TTP patient plasma) in different ratios. In addition, three sample levels were prepared by mixing plasma from normal human donors with heat inactivated plasma. To prepare heat inactivated plasma with no residual ADAMTS13 activity, citrated plasma samples from normal donors were heat-inactivated for 1 hour at 56℃. The samples tested included levels below, around and above the cut-off of 0.1 IU/mL.
| Sample | N | Mean (IU/mL) | Repeatability | Between-run | Between-day | Between-site | Reproducibility | |||||
|---|---|---|---|---|---|---|---|---|---|---|---|---|
| SD | %CV | SD | %CV | SD | %CV | SD | %CV | SD | %CV | |||
| S1 | 30 | 0.67 | 0.03 | 4.81 | 0.04 | 6.20 | 0.05 | 6.60 | 0.00 | 0.00 | 0.07 | 10.24 |
| S2 | 30 | 0.46 | 0.02 | 4.67 | 0.04 | 8.40 | 0.00 | 0.00 | 0.01 | 2.20 | 0.05 | 9.86 |
| S3 | 29 | 0.25 | 0.01 | 3.75 | 0.02 | 6.60 | 0.00 | 0.00 | 0.01 | 4.80 | 0.02 | 8.99 |
| S4 | 29 | 0.19 | 0.01 | 4.46 | 0.02 | 7.40 | 0.00 | 0.00 | 0.00 | 0.00 | 0.02 | 8.64 |
| S5 | 30 | 0.14 | 0.01 | 4.17 | 0.01 | 9.00 | 0.00 | 0.00 | 0.00 | 0.00 | 0.01 | 9.88 |
| S6 | 30 | 0.07 | 0.00 | 4.53 | 0.01 | 6.30 | 0.00 | 0.00 | 0.01 | 7.00 | 0.01 | 10.45 |
| S7 | 30 | 0.66 | 0.03 | 4.46 | 0.04 | 5.70 | 0.01 | 2.00 | 0.03 | 3.90 | 0.06 | 8.49 |
| S8 | 30 | 0.23 | 0.01 | 4.31 | 0.01 | 5.30 | 0.00 | 0.00 | 0.00 | 0.00 | 0.02 | 6.85 |
| S9 | 30 | 0.12 | 0.00 | 2.97 | 0.01 | 6.40 | 0.00 | 0.00 | 0.00 | 3.60 | 0.01 | 8.01 |
| Sample | Mean (IU/mL) | Total results | Qualitative agreement | |
|---|---|---|---|---|
| Number of correct results | % Correct call | |||
| S1 negative | 0.67 | 30 | 30/30 | 100 |
| S2 negative | 0.46 | 30 | 30/30 | 100 |
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| Mean | Total | Qualitative agreement | ||
|---|---|---|---|---|
| Sample | (IU/mL) | results | Number of correctresults | % Correct call |
| S3negative | 0.25 | 29 | 29/29 | 100 |
| S4negative | 0.19 | 29 | 29/29 | 100 |
| S5negative | 0.14 | 30 | 30/30 | 100 |
| S6positive | 0.07 | 30 | 30/30 | 100 |
| S7negative | 0.66 | 30 | 30/30 | 100 |
| S8negative | 0.23 | 30 | 30/30 | 100 |
| S9negative | 0.12 | 30 | 30/30 | 100 |
2. Analytical Specificity/Interference:
Interference studies were conducted based on the CLSI EP07 3rd Edition guideline. Three base pools mimicking high (1.0 IU/mL), medium (0.5 IU/mL) and low (0.1 IU/mL) levels of ADAMTS13 activity were prepared by mixing human citrated plasma (non-icteric, nonturbid and non-hemolyzed) with plasma rendered ADAMTS13 deficient by heat inactivation. Interference testing was conducted by paired-difference testing using one lot of reagents for both common endogenous and extrinsic interferents. Each sample was tested in five replicates. Samples with and without the interferent were measured, and the measurand concentration difference was determined.
None of the substances in the following table were found to lead to clinically significant interference.
| Potential interferingsubstance | No interference up to thefollowing evaluated clinicallysignificant concentration: |
|---|---|
| Exogenous | |
| Acetaminophen | 15.6 mg/dL |
| Acetylcysteine | 15.0 mg/dL |
| Ampicillin Na | 7.5 mg/dL |
| ASA | 3.0 mg/dL |
| Biotin | 0.351 mg/dL |
| Caplacizumab | 0.15 mg/dL |
| Cefoxitin Na | 660.0 mg/dL |
| Cyclosporine | 0.18 mg/dL |
| Doxycycline | 1.8 mg/dL |
| Heparin | 330 units/dL |
| Ibuprofen | 21.9 mg/dL |
| Levodopa | 0.75 mg/dL |
| Methyldopa | 2.25 mg/dL |
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| Potential interferingsubstance | No interference up to thefollowing evaluated clinicallysignificant concentration: |
|---|---|
| Exogenous | |
| Metronidazole | 12.3 mg/dL |
| Phenylbutazone | 32.0 mg/dL |
| Prednisolone | 0.12 mg/dL |
| Rifampicin | 4.8 mg/dL |
| Rituximab | 50.0 mg/dL |
| Theophylline | 6.0 mg/dL |
| Endogenous | |
| Intralipid | 500 mg/dL |
| Hemoglobin | 220 mg/dL |
| Unconjugated Bilirubin | 66.0 mg/dL |
| Conjugated Bilirubin | 66.0 mg/dL |
| GST | 0.02 mg/dL |
| VWF | 2.0 IU/mL |
| Human anti mouse antibody | titer >12 |
| Rheumatoid factor | 156 IU/mL |
3. Assay Reportable Range:
Not applicable
-
- Traceability, Stability, Expected Values (Controls, Calibrators, or Methods):
Traceability
Target values for calibrators and controls are traceable to the first International Standard for ADAMTS13 Activity and Antigen in Plasma (NIBSC WHO 1st international Standard ADAMTS13 Plasma 12/252).
Stability of calibrators and controls
Stability of calibrators and controls were evaluated in accordance with CLSI EP25A. Three lots of calibrators and controls were used in the study and stored in their final packaging at 2-8°C. At time points 0, 12, 24 and 30 months, sets of calibrators and controls were placed into stable storage (-70°C). At the end of the study (t=30 months), all calibrators and controls were tested in triplicate in one single run on one instrument using one lot of reagents. The data supported a real-time stability of 24 months.
-
- Assay Cut-Off:
Not applicable
- Assay Cut-Off:
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B Comparison Studies:
1. Clinical Performance Study:
The clinical performance study was conducted at two external sites, one located in U.S. and the other located outside of the U.S. Testing was performed double blinded. The clinician making the diagnosis decisions and selecting the samples was blinded to the Technozym ADAMTS 13 activity results and the laboratory technician conducting the Technozym assay was blinded to the diagnosis. Samples were tested in duplicate using the Technozym ADAMTS13 Activity assay. One kit lot was used per study site. At each study site, tests were performed by one laboratory professional. The study samples used in testing were residual samples selected from a local repository of frozen human citrated plasma from patients diagnosed with thrombotic microangiopathies (TMA) (i.e., clinical suspicion of thrombotic thrombocytopenic purpura (TTP)) by board-certified clinician according to the local testing algorithm for TMAs. All patient samples were from donors > 6 months of age and patient population is representative of intended use population.
Combined agreement analysis for both sites with a total of 137 samples included in the clinical performance study.
| Clinical diagnosis of TTP | ||||
|---|---|---|---|---|
| Positive | Negative | Total | ||
| TechnozymADAMTS13Activity | Positive | 28 | 3 | 31 |
| Negative | 5 | 101 | 106 | |
| Total | 33 | 104 | 137 | |
| Sensitivity = 84.8% (28/33); 95% CI: (69.1% to 93.3%)Specificity = 97.1% (101/104); 95% CI: (91.9% to 99%) | Positive Predictive Value (PPV) = 90.2% (28/31); 95% CI: (75.2% to 96.6%)Negative Predictive Value (NPV) = 95.3% (101/106); 95% CI: (90.0% to 97.8%) |
C Clinical Studies:
-
- Clinical Sensitivity:
Refer to Clinical Performance Study
- Clinical Sensitivity:
2. Clinical Specificity:
Refer to Clinical Performance Study
-
- Other Clinical Supportive Data (When 1. and 2. Are Not Applicable):
Not applicable
- Other Clinical Supportive Data (When 1. and 2. Are Not Applicable):
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D Clinical Cut-Off:
The clinical cut-off for TTP diagnosis is 10% or 0.1 IU/mL ADAMTS13 activity.
E Other Supportive Performance Characteristics Data:
1. Prozone Effect (Hook Effect)
Information was provided to support that no significant hook effect was observed up to activity levels of 8 IU/mL.
2. Cross-contamination Studies
A study was performed to evaluate if cross-contamination and/or carryover occurs between samples in the plate wells during the assay procedure. Low samples with a target concentration of 0.1 IU/mL and high samples with a target concentration of 1.0 IU/mL were used to perform the studies. In the first stage, the signal of only low samples was evaluated throughout the microplate. In the second stage, two test plates were run with an alternating pattern over all available patient sample locations. The pattern consisted of two wells containing only the low samples followed by two wells containing the high sample. The study was performed by three operators performing testing with three microplate readers and plate washer combinations with one lot of reagents. No cross-contamination was observed.
3. Reagent Stability Studies:
Real-time Shelf-life Stability Studies
The real-time stability study was conducted in accordance with CLSI guideline EP25-A. The study was conducted with three lots of Technozym ADAMTS13 Activity assay kits. Eight samples were prepared by mixing citrated human plasma in human ADAMTS13 activity deficient plasma (HIP) in different ratios. These samples were aliquoted and frozen at -20℃ and a fresh aliquot was used for every test time point. Reagent kits were stored in their final packaging at 2-8°C. Time points used in the real time stability study included: 0, 6. 12, 24 and 30 months. Reagent kits were retrieved and tested with different ADAMTS13 activity sample levels at the end of each designated time point in the study. The testing was done in duplicates for each ADAMTS13 activity level. Based on the real-time stability results, the data supports a shelf-life of the Technozym ADAMTS13 Activity assay kit for up to 24 months at 2-8°C.
4. Sample Stability Studies
Frozen sample stability
Eight samples were prepared by mixing citrated human plasma with heat treated citrated human plasma in different ratios, and aliquots was stored frozen at < - 20°C. At each test time point, a randomly selected set of aliquots was thawed at 37°C using a water bath and testing was performed. Samples were tested in duplicates in the Technozym ADAMTS13 Activity assay within one run. Testing was performed at the time points 0 (stored at -20°C for minimum of 5 days before testing), 6, 12, 18 and 24 months. One reagent lot was used
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throughout the study and testing was performed on one instrument. The study supports frozen sample stability of 12 months.
Fresh Sample stability
Six samples were prepared by mixing freshly drawn citrated human plasma with native TTP plasma (no ADAMTS13 activity) and aliquoted for testing at room temperature (18-25°C) and under refrigerated conditions (2-8°C). For samples stored at room temperature, testing was performed at time points 0, 4, 8, 9, 24 and 25 hours. For samples stored under refrigerated conditions, testing was performed at time points 0, 24, 25, 48 and 49 hours. All samples were tested in duplicates using three different reagent lots. The study supports a sample stability for up to 8 hours at room temperature (18-25°C) and up to 24 hours under refrigerated conditions.
Proposed Labeling: VII
The labeling supports the decision to grant the De Novo request for this device.
VIII Identified Risks and Mitigations:
| Risks to Health | Mitigation Measures |
|---|---|
| Clinical action based on false positive resultsmay lead to inappropriate patient management,or unnecessary treatments. | Certain design verification and validationactivities and documentation, includingcertain studies.Certain labeling information, includingcertain limiting statements and performancecharacteristics. |
| Clinical action based on false negative resultsmay lead to delayed diagnosis, misdiagnosis, ordiscontinuation of treatment. | Certain design verification and validationactivities and documentation, includingcertain studies.Certain labeling information, includingcertain limiting statements and performancecharacteristics. |
IX Benefit/Risk Assessment:
A Summary of the Assessment of Benefit:
There is currently no FDA market-authorized device for determining ADAMTS13 activity. Patients with thrombotic thrombocytopenic purpura (TTP) typically present with thrombocytopenia, microangiopathic hemolytic anemia (e.g., low hemoglobin, low hematocrit, low haptoglobin, elevated LDH, presence of schistocytes in peripheral blood smear) and various degrees of organ damage. These changes, however, are non-specific for TTP and can also occur in many of the differential diagnoses. TTP is caused by ADAMTS13 deficiency. The availability of the test may aid in the differential diagnosis of thrombocytopenic purpura (TTP) from other thrombotic microangiopathies (TMA) as described in the Journal of Thrombosis and
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Haemostasis (2020) "ISTH guidelines for the diagnosis of thrombotic thrombocytopenic purpura" (e.g., "although diagnosis of TTP relies on a high index of suspicion, based on clinical presentation and laboratory results, the panel recognized that the importance of having an ADAMTS13 activity test in the diagnosis and initial management process").
B Summary of the Assessment of Risk:
When used as intended, the risks of the device are mainly related to false positive or false negative test results. For a false positive test result, the risk could include unnecessary further testing or inappropriate patient management, including cessation of investigation for other diseases, resulting in missed opportunities to properly treat the patient. Additionally, a false positive test may lead to unnecessary treatments with side effects such as bleeding, fatigue, pyrexia, headache, paresthesia, urticaria, fatal infusion reactions, tumor lysis syndrome, severe mucocutaneous reaction and progressive multifocal leukoencephalopathy. Risks of a false negative test include a missed or delayed diagnosis, improper patient management including continuation of investigating the etiology of a patient's symptoms, which usually consists of further history, physical examination, and testing. The additional risk associated with a false negative test is related to the inappropriate discontinuation of treatment which can lead to missed opportunities for the timely treatment of TTP positive patients. Such treatment has been associated with faster normalization of platelet count, lower incidence of TTP-related death, lower rate of recurrence of TTP and lower incidence of thromboembolic event than placebo in clinical trials.
C Patient Perspectives:
This submission did not include specific information on patient perspectives for this device.
D Summary of the Assessment of Benefit-Risk:
Device design verification and validation, including precision, method comparison, and interference studies will help ensure that the device functions as intended and mitigate the risk of false positive or false negative test results. A limitation statement conveying that results from the assay alone should not be used in making treatment decisions will be included in the labeling, as an additional mitigation against the risk of false positive and false negative results. Overall, while general controls are insufficient to mitigate the risks of the device, in light of the special controls, the probable benefits outweigh the probable risks of incorrect test results for the proposed indications for use.
Conclusion: X
The De Novo request is granted, and the device is classified under the following and subject to the special controls identified in the letter granting the De Novo request:
Product Code(s): SAC Device Type: ADAMTS13 activity test system Class: II Regulation: 21 CFR 864.7297
N/A