Development and applications of a new technique to study blood clot contraction (retraction)

Abstract

Despite the importance for hemostasis and thrombosis, platelet-governed clot shrinkage has not been systematically studied, partially due to the lack of methodology to follow and quantify clot contraction dynamics. We have developed a new technique based on the continuous tracking of clot size. An optical platform for this method is the commercially available Thrombodynamics Analyser System (HemaCore, Russia). A standard procedure includes recalcification of whole citrated blood and addition of thrombin to initiate blood clotting and platelet activation. The clot contraction is monitored by taking images every 15 seconds over 20 minutes or more followed by off-line computational analysis that provides a kinetic curve characterized by 6 numerical parameters. Clot contraction has at least two phases characterized by distinct rates. Exogenous Ca2+ is not indispensable for clot contraction to occur; however, the clots formed without addition of Ca2+ were less stable. Recalcification of blood with 2-5 mM [Ca2+] prevented the red blood cell fallout without an effect on the contraction kinetics. 10 mM [Ca2+] partially inhibited clot contraction. Iodoacetamide, an inhibitor of factor XIIIa, did not affect clot formation but abolished clot shrinkage, confirming that factor XIIIa is essential for clot contraction. Thrombin enhanced the rate and degree of clot contraction in a dose-dependent manner. Blood clot contraction was substantially delayed in patients on warfarin compared to healthy donors. We developed an accurate and simple assay for blood clot contraction which can be used for research and may be potentially useful for in vitro diagnostics.

Keywords

blood clotting / fibrin / platelet function

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