Platelet retraction force measurements using flexible post force sensors

Platelets play an important role in hemostasis by forming a thrombotic plug that seals the vessel wall and promotes vascular healing. After platelets adhere and aggregate at the wound site, their next step is to generate contractile forces through the coordination of physicochemical interactions between actin, myosin, and alpha(IIb)beta(3) integrin receptors that retract the thrombus' size and strengthen its adhesion to the exposed matrix. Although platelet contractile forces (PCF) are a definitive feature of hemostasis and thrombosis, there are few approaches that can directly measure them. In this study, we describe the development of an approach to measure PCF in microthrombi using a microscopic flexible post force sensor array. Quasi-static measurements and live microscopic imaging of thrombin-activated platelets on the posts were conducted to assay the development of PCF to various hemostatic conditions. Microthrombi were observed to produce forces that monotonically increased with thrombin concentration and activation time, but forces subsided when thrombin was removed. PCF results were statistically similar on arrays of posts printed with fibronectin or fibrinogen. PCF measurements were combined with clot volume measurements to determine that the average force per platelet was 2.1 +/- 0.1 nN after 60 min, which is significantly higher than what has been measured with previous approaches. Overall, the flexible post arrays for PCF measurements are a promising approach for evaluating platelet functionality, platelet physiology and pathology, the impacts of different matrices or agonists on hemostatic responses, and in providing critical information regarding platelet activity that can guide new hemostatic or thrombotic strategies.