A novel biosensor for measuring plasmin activity

Plasmin facilitates fibrinolysis by catalyzing the degradation of fibrin clots. Reliable methods to assess fibrinolysis by measuring plasmin generation are needed in the preclinical and clinical settings.

This study aimed to design, synthesize, and evaluate a fluorescence resonance energy quenching-based plasmin sensor (FPS) protein to monitor plasmin generation.

The hydrolysis of FPS was monitored by loss of fluorescence resonance energy quenching to evaluate efficiency and specificity. Plasma-based plasmin generation assays were used to evaluate fibrinolytic cofactors and inhibitors.

Compared with a peptide plasmin sensor, FPS demonstrated higher efficiency for plasmin and was not cleaved by plasmin inhibited by α₂-macroglobulin. FPS interacts with plasmin's kringle domain 5. FPS can measure plasmin generation in clotting plasma, which was increased by an inhibitory antibody against α₂-antiplasmin and attenuated by aprotinin, tranexamic acid, and fibrin absence. Maximal plasmin generation occurs after peak clot formation. Furthermore, when antiplasmin is inhibited, the presence of plasminogen activator inhibitor-1 can be measured. Unlike peptide sensors, FPS does not inhibit clot lysis. Lastly, plasmin generation measured using FPS is relatively consistent in plasma from healthy volunteers, with only a 2-fold variation in peak plasmin, peak time, and endogenous plasmin potential.

FPS may improve the evaluation of fibrinolysis in preclinical and clinical settings, thereby enhancing our understanding of fibrinolytic disorders.