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

Autor: D, Chen, C A, Daigh, J I, Hendricksen, R K, Pruthi, W L, Nichols, J A, Heit, W G, Owen
Rok vydání: 2008
Předmět:
Zdroj: Journal of Thrombosis and Haemostasis. 6:323-330
ISSN: 1538-7836
1538-7933
DOI: 10.1111/j.1538-7836.2007.02845.x
Popis: 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.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 IU dL(-1)), and reported as percent or IU dL(-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.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 (P0.01), especially for type 2A VWD patients.This new flow cytometry-based VWF:RCo assay is simple, accurate, specific and sensitive, particularly for type 2 VWD.
Databáze: OpenAIRE
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