Dataset Information

Multicenter performance evaluation and reference range determination of a new one-stage factor VIII assay.

ABSTRACT:

Introduction

We conducted a multicenter evaluation of a new one-stage factor VIII (FVIII) assay (Roche Diagnostics), intended for the quantitative assessment of FVIII activity. We evaluated the analytical performance of the FVIII assay on the cobas t 711 analyzer.

Methods

Experiments performed at three laboratories used 3.2% citrated residual or commercially purchased plasma samples. Five human plasma pools and two controls were used to determine assay within-run and within-laboratory precision, and total reproducibility; coefficients of variation (CVs) and/or standard deviations (SDs) were calculated. Lot-to-lot variability and method comparison (vs Coagulation FVIII Deficient Plasma/Dade Actin FS Activated PTT reagent/Standard Human Plasma Calibrator on the Sysmex CS-5100 analyzer; Siemens Healthineers) were evaluated by Passing-Bablok and Deming regression, respectively, and Pearson's r calculated. Assay-specific reference range was determined using 199 fresh plasma samples from healthy adults, not receiving anticoagulants.

Results

Across sites, SDs for repeatability were 0.016-0.046 for samples with ≤1.0 international units (IU)/dL FVIII activity; CVs were 0.9%-3.8% for samples with >1.0 IU/dl activity. Among samples with mean FVIII activity 0.344-133 IU/dl, good intermediate precision (SD 0.020 for samples with 0.344 IU/dl activity; CV 1.8%-4.7%) and good total reproducibility (CV 2.0%-13.3%) were observed. The FVIII assay showed excellent lot-to-lot variability (Pearson's r = .999) and good correlation with the comparator assay (Pearson's r = .993-.996). The reference range for FVIII activity was 82.2-218.0 IU/dl.

Conclusion

The one-stage FVIII assay demonstrated robust analytical performance on the cobas t 711 analyzer, supporting its use in routine laboratory practice.

SUBMITTER: Lowe AE

PROVIDER: S-EPMC8993620 | biostudies-literature | 2022 Apr

REPOSITORIES: biostudies-literature

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Publications

Multicenter performance evaluation and reference range determination of a new one-stage factor VIII assay.

Lowe Anna E AE Jones Robert R Kitchen Steve S Geisen Ulrich U Rozsnyai Gergely G Jilma Petra P Quehenberger Peter P

Journal of clinical laboratory analysis 20220311 4

<h4>Introduction</h4>We conducted a multicenter evaluation of a new one-stage factor VIII (FVIII) assay (Roche Diagnostics), intended for the quantitative assessment of FVIII activity. We evaluated the analytical performance of the FVIII assay on the cobas t 711 analyzer.<h4>Methods</h4>Experiments performed at three laboratories used 3.2% citrated residual or commercially purchased plasma samples. Five human plasma pools and two controls were used to determine assay within-run and within-labora ...[more]

PMID: 35274775

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Project description:IntroductionDiagnosis, treatment monitoring and assessment of desmopressin effect in haemophilia A patients are performed by measurement of factor VIII activity (FVIII). The two assays commonly applied are the one-stage assay and the chromogenic assay. Especially in non-severe haemophilia A, discrepancies between these assays are common. It is still unestablished which assay corresponds best with bleeding phenotype and desmopressin effect.AimTo correlate FVIII levels measured by the one-stage assay and by the chromogenic assay with bleeding phenotype and, additionally, to compare FVIII assay discrepancies before and after desmopressin administration.MethodFactor VIII was measured in 130 non-severe haemophilia A patients during routine visits to the outpatient clinic and/or during desmopressin testing. FVIII was measured by both the one-stage assay and the chromogenic assay. Discrepancies between assays were defined as at least a twofold difference of FVIII or an absolute FVIII difference between measurements of ≥0.10 IU/mL. Bleeding phenotype was defined as annual number of treated bleedings (adjusted ABR).ResultsHundred and thirty non-severe haemophilia A patients were included. In 31/130 patients, assay results were discrepant. However, FVIII measurements with both assays correlated adequately with adjusted ABR. In addition, in 27/130 patients FVIII measurements at baseline and after desmopressin administration were analysed. In 13/27 patients, all measurements were either equivalent or discrepant when results were compared. In 14/27 patients, this was not the case as both equivalent measurements and discrepant measurements at different time points within one patient were observed.ConclusionNeither the one-stage assay nor the chromogenic assay is superior in predicting bleeding phenotype. In addition, equivalent or discrepant FVIII results measured before desmopressin do not always predict FVIII assay results after desmopressin administration.

Project description:Essentials Performance of the one-stage clotting (OSC) assay varies with the clotting activator used. Recombinant FIX-albumin fusion protein (rIX-FP) was reliably monitored with most OSC reagents. rIX-FP shows comparable reagent-dependent variability to other rFIX products in the OSC assay. Actin® FS and kaolin-based reagents underestimated rIX-FP activity by around 50% in the OSC assay. SUMMARY: Background Measuring factor IX activity (FIX:C) with one-stage clotting (OSC) assays, based on the activated partial thromboplastin time (APTT), is the current mainstay of diagnostic techniques for hemophilia B. Assessing the performance of new recombinant FIX (rFIX) products in OSC assays is essential, as APTT reagents from different manufacturers yield different potency estimates for rFIX. Objectives To evaluate the extent to which choice of reagent composition influences rFIX potency measurements of recombinant FIX-albumin fusion protein (rIX-FP, IDELVION) activity in OSC assays. Methods rIX-FP was added to FIX-deficient plasma, and FIX:C was assessed centrally and locally in a multicenter international field study with a variety of commercial OSC APTT reagents. Paired sample analysis of clinical samples was performed to compare values of FIX:C from local and central laboratories. In-house bioanalytical investigations with spiked samples were conducted to compare the APTT-reagent dependent variability of rIX-FP with unmodified rFIX and rFIX Fc fusion protein (rFIXFc). Results Central and local assessments of FIX:C from 10 countries and 21 participating centers showed comparable results to those from the central laboratory across the majority of 18 different APTT reagents from both clinical and spiked samples. There was a consistent underestimation of rIX-FP activity of ≈ 50% with OSC assays using Actin FS or kaolin-based APTT reagents. In the bioanalytical study, rIX-FP showed comparable variability in OSC assays to unmodified rFIX and rFIXFc. Conclusions rIX-FP activity can be accurately measured by the use of OSC assays with the majority of commercial reagents. Actin FS or kaolin-based reagents will probably lead to a 50% underestimation of activity.

Project description:Development of neutralizing antibodies against biotherapeutic agents administered to prevent or treat various clinical conditions is a longstanding and growing problem faced by patients, medical providers and pharmaceutical companies. The hemophilia A community has deep experience with attempting to manage such deleterious immune responses, as the lifesaving protein drug factor VIII (FVIII) has been in use for decades. Hemophilia A is a bleeding disorder caused by genetic mutations that result in absent or dysfunctional FVIII. Prophylactic treatment consists of regular intravenous FVIII infusions. Unfortunately, 1/4 to 1/3 of patients develop neutralizing anti-FVIII antibodies, referred to clinically as "inhibitors," which result in a serious bleeding diathesis. Until recently, the only therapeutic option for these patients was "Immune Tolerance Induction," consisting of intensive FVIII administration, which is extraordinarily expensive and fails in ~30% of cases. There has been tremendous recent progress in developing novel potential clinical alternatives for the treatment of hemophilia A, ranging from encouraging results of gene therapy trials, to use of other hemostatic agents (either promoting coagulation or slowing down anti-coagulant or fibrinolytic pathways) to "bypass" the need for FVIII or supplement FVIII replacement therapy. Although these approaches are promising, there is widespread agreement that preventing or reversing inhibitors remains a high priority. Risk profiles of novel therapies are still unknown or incomplete, and FVIII will likely continue to be considered the optimal hemostatic agent to support surgery and manage trauma, or to combine with other therapies. We describe here recent exciting studies, most still pre-clinical, that address FVIII immunogenicity and suggest novel interventions to prevent or reverse inhibitor development. Studies of FVIII uptake, processing and presentation on antigen-presenting cells, epitope mapping, and the roles of complement, heme, von Willebrand factor, glycans, and the microbiome in FVIII immunogenicity are elucidating mechanisms of primary and secondary immune responses and suggesting additional novel targets. Promising tolerogenic therapies include development of FVIII-Fc fusion proteins, nanoparticle-based therapies, oral tolerance, and engineering of regulatory or cytotoxic T cells to render them FVIII-specific. Importantly, these studies are highly applicable to other scenarios where establishing immune tolerance to a defined antigen is a clinical priority.

Dataset Information

Multicenter performance evaluation and reference range determination of a new one-stage factor VIII assay.

Introduction

Methods

Results

Conclusion

Publications

Multicenter performance evaluation and reference range determination of a new one-stage factor VIII assay.

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