Automated Imaging and Analysis of Platelet, Coagulation and Fibrinolysis Activities Using a Novel Flow Chip-Based System at Physiological Temperature.

Conventional whole-blood flow assays for quantifying thrombus formation are typically performed at room temperature and are technically demanding, which limits their translational applicability. We engineered a novel, disposable, mountable, and single-channel microfluidic chip (MC-2S), which is based on the Maastricht chamber (MC) and designed for automated evaluation of platelet function, coagulation and fibrinolysis under physiological conditions. The MC-2S chip allows customizable choices of thrombogenic surfaces, such as collagen and tissue factor. The chip was used in combination with an adapted, 1.3 kg brightfield/fluorescence microscope, operating at physiological temperature (37 °C), and with scripts for automated multicolor analysis of image features. The integrated system enables a robust, rapid, and high-content quantification of the kinetics of thrombus formation and dissolution. In platelet-sensitive mode, MC-2S demonstrated high sensitivity to antiplatelet therapy with aspirin or cangrelor. In coagulation-sensitive mode, it detected the anticoagulant effect of rivaroxaban plus its reversal by andexanet-α. In fibrinolysis-sensitive mode, it monitored tissue-type plasminogen activator-induced thrombus dissolution, inhibited by tranexamic acid. Collectively, the MC-2S platform was found to provide a versatile, physiologically relevant tool for functional hemostasis testing, with high potential for the acute and subacute evaluation of patient blood samples in the context of bleeding disorders, thrombosis risk, and drug monitoring.