A highly-sensitive plasma von Willebrand factor ristocetin cofactor (VWF:RCo) activity assay by flow cytometry

Dong Chen, C. A. Daigh, J. I. Hendricksen, R. K. Pruthi, W. L. Nichols, J. A. Heit, W. G. Owen

Research output: Contribution to journalArticle

37 Scopus citations

Abstract

Background: Assays of plasma von Willebrand factor (VWF) ristocetin cofactor activity (VWF:RCo) are essential for the laboratory diagnosis of von Willebrand disease (VWD) and for monitoring therapy. However, current manual or automated VWF:RCo assay methods have relatively poor operating characteristics. Our goal was to develop and validate a simple, accurate, specific and sensitive platelet-based VWF:RCo assay. Methods: Using green or red fluorochrome-labeled, fixed normal platelets and normal or patient plasma, ristocetin-dependent and VWF-mediated platelet aggregation was detected by flow cytometry. VWF:RCo activity was assayed as the number of double-positive events (green and red) among all green or red events, relative to the calibrator plasma signal (6-150% or IUdL-1), and reported as percent or IUdL-1. We tested plasma samples from normal donors (n = 51) and known VWD patients (type 1, n = 16; type 2, n = 17) based on clinical history, levels of plasma VWF antigen (VWF:Ag), VWF:RCo activity (manual platelet aggregometry/agglutination assay), factor (F) VIII activity and VWF multimer analysis. Results: For normal donors and type 1 VWD patients, VWF:RCo activity by flow cytometry vs. manual platelet aggregation correlated closely (R2 = 0.74), and VWF:RCo/VWF:Ag ratios did not differ significantly. In contrast, VWF:RCo/VWF:Ag ratios for type 2 VWD subtypes were significantly lower using VWF:RCo by flow cytometry vs. manual platelet aggregation assay (P < 0.01), especially for type 2A VWD patients. Conclusions: This new flow cytometry-based VWF:RCo assay is simple, accurate, specific and sensitive, particularly for type 2 VWD.

Original languageEnglish (US)
Pages (from-to)323-330
Number of pages8
JournalJournal of Thrombosis and Haemostasis
Volume6
Issue number2
DOIs
StatePublished - Feb 2008

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Keywords

  • Flow cytometry
  • Platelet
  • Ristocetin cofactor activity
  • Von Willebrand disease
  • Von Willebrand factor activity

ASJC Scopus subject areas

  • Hematology

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