Analytical characterization and reference interval of an enzyme-linked immunosorbent assay for active von Willebrand factor

Lisa N. van der Vorm*, Li Li, Dana Huskens, Walid Chayoua, Hilde Kelchtermans, Philip G. de Groot, Mark Roest, Jasper A. Remijn, Bas de Laat

*Corresponding author for this work

Research output: Contribution to journalArticleAcademicpeer-review

9 Citations (Web of Science)

Abstract

Background

Interaction of von Willebrand factor (VWF) with platelets requires a conformational change that exposes an epitope within the VWF Al domain, enabling platelet glycoprotein Iba binding. Quantification of this "active" conformation of VWF has been shown to provide pathophysiological insight into conditions characterized by excessive VWF-platelet interaction.

Methods

We developed an immunosorbent assay based on a variable heavy chain antibody fragment against the VWF Al domain as a capture antibody. Assay performance in terms of specificity (binding to active R1306W- and sheared VWF), precision, accuracy, linearity, limits of detection and stability were determined. Active VWF, VWF antigen, VWF ristocetin cofactor activity, VWF:GP1bM and VWF propeptide were measured in citrated plasma and platelet-VWF binding in whole blood from 120 healthy individuals to establish a reference interval for active VWF and to assess associations with other VWF parameters.

Results

Intra- and inter-assay CVs were between 2.4-7.2% and 4.1-9.4%, depending on the level. Mean recovery of spiked recombinant R1306W VWF was 103 +/- 3%. The assay was linear in the range of 90.1-424.5% and had a limit of quantification of 101%. The reference interval for active VWF was 91.6-154.8% of NPP. Significant, positive correlations between active VWF and all other VWF parameters were found, with the strongest correlation with VWF: GP1bM binding.

Conclusions

We developed and validated an immunosorbent assay for the accurate detection of active VWF levels in plasma. The assay fulfilled all analytical criteria in this study and a reference interval was established, allowing its use to quantify active VWF in pathological conditions for future research.

Original languageEnglish
Article number0211961
Number of pages16
JournalPLOS ONE
Volume14
Issue number2
DOIs
Publication statusPublished - 13 Feb 2019

Keywords

  • PLATELET-ADHESION
  • FACTOR-VIII
  • VONWILLEBRAND-FACTOR
  • FACTOR LEVEL
  • BLOOD-GROUP
  • DISEASE
  • DIAGNOSIS
  • ADAMTS13
  • AGE
  • ASSOCIATION

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