Background and Objectives: Preliminary studies indicated that α1-antitrypsin (A1AT) is the most abundant protein that is non-covalently bound to fibrin clots prepared from plasma. The aim of this study was to identify and characterize fibrin(ogen)-bound A1AT. Methods and Results: Plasma clots were prepared and extensively washed with saline. Clot-bound A1AT could only be extracted using denaturing agents such as urea, thiourea or SDS, pointing to an apparently strong association. Purified fibrinogen, but still containing A1AT as a contaminant, was gel filtered, which showed that the A1AT was bound to fibrinogen. A specific ELISA detected the presence of A1AT-fibrinogen complexes in both purified fibrinogen and pooled normal plasma. Finally, fibrin(ogen)-Sepharose chromatography indicated that A1AT purified from plasma contained a small fraction of fibrin(ogen)-binding A1AT. To study the inhibitory activity of fibrin(ogen)-bound A1AT, both fibrinogen containing A1AT and washed plasma clots were incubated with increasing amounts of elastase. SDS-PAGE and Western blotting showed under both conditions the generation of the A1AT-elastase complex as well as cleaved A1AT. The inhibitory activity of fibrin(ogen)-bound A1AT was also demonstrated by measuring elastase-induced lysis of fibrin clots. Conclusion: Fibrin clots contain strongly bound A1AT, which is functionally active as a serine protease inhibitor (serpin). This A1AT might play a role in the local regulation of proteases involved in coagulation or fibrinolysis and represent a novel link between the inflammatory and hemostatic systems.

Western blotting, article, chromatography, clinical article, controlled study, enzyme linked immunosorbent assay, fibrin clot, human, human tissue, lysis, normal human, plasma, polyacrylamide gel electrophoresis, priority journal, protein analysis, protein binding,
Journal of Thrombosis and Haemostasis
Erasmus MC: University Medical Center Rotterdam

Talens, S, Malfliet, J.J.M.C, van Hal, P.Th.W, Leebeek, F.W.G, & Rijken, D.C. (2013). Identification and characterization of α1-antitrypsin in fibrin clots. Journal of Thrombosis and Haemostasis, 11(7), 1319–1328. doi:10.1111/jth.12288