Comparison of assays measuring extracellular vesicle-tissue factor in plasma samples: Communication from the ISTH SSC Subcommittee on Vascular Biology.
(2024) Journal of thrombosis and haemostasis — Vol. 22, n° 10, p. 2910-2921 (2024)
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- (en) BACKGROUND: Scientific and clinical interest in extracellular vesicles (EVs) is growing. EVs that expose tissue factor (TF) bind factor VII/VIIa and can trigger coagulation. Highly procoagulant TF-exposing EVs are detectable in the circulation in various diseases, such as sepsis, COVID-19, or cancer. Many in-house and commercially available assays have been developed to measure EV-TF activity and antigen, but only a few studies have compared some of these assays. OBJECTIVES: The International Society on Thrombosis and Haemostasis Scientific and Standardization Committee Subcommittee on Vascular Biology initiated a multicenter study to compare the sensitivity, specificity, and reproducibility of these assays. METHODS: Platelet-depleted plasma samples were prepared from blood of healthy donors. The plasma samples were spiked either with EVs from human milk or EVs from TF-positive and TF-negative cell lines. Plasma was also prepared from whole human blood with or without lipopolysaccharide stimulation. Twenty-one laboratories measured EV-TF activity and antigen in the prepared samples using their own assays representing 18 functional and 9 antigenic assays. RESULTS: There was a large variability in the absolute values for the different EV-TF activity and antigen assays. Activity assays had higher specificity and sensitivity compared with antigen assays. In addition, there was a large intra-assay and interassay variability. Functional assays that used a blocking anti-TF antibody or immunocapture were the most specific and sensitive. Activity assays that used immunocapture had a lower coefficient of variation compared with assays that isolated EVs by high-speed centrifugation. CONCLUSION: Based on this multicenter study, we recommend measuring EV-TF using a functional assay in the presence of an anti-TF antibody.
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Bonifay, A., Mackman, N., Hisada, Y., Sachetto, A. T. A., Hau, C., Gray, E., Hogwood, J., Aharon, A., Badimon, L., Barile, L., Baudar, J., Beckmann, L., Benedikter, B., Bolis, S., Bouriche, T., Brambilla, M., Burrello, J., Camera, M., Campello, E., et al. (2024). Comparison of assays measuring extracellular vesicle-tissue factor in plasma samples: Communication from the ISTH SSC Subcommittee on Vascular Biology. Journal of thrombosis and haemostasis, 22(10), 2910-2921. https://doi.org/10.1016/j.jtha.2024.05.037 (Original work published 2024)