Summary
This protocol paper describes the RT-FluFF assay, a novel in vitro thrombolysis testing platform that addresses a significant gap in drug development by incorporating physiologically relevant shear flow during both clot formation and digestion phases. Unlike conventional static clot models, the assay allows real-time tracking of fibrinolysis under conditions that more accurately represent human circulatory physiology whilst maintaining the reproducibility and control of laboratory testing. The flexible system enables testing across diverse flow conditions and pharmaceutical interventions, potentially improving translational success rates for thrombolytic drug candidates.
UK applicability
The assay development has potential application to UK pharmaceutical research and drug development pipelines, particularly for companies and academic institutions developing thrombolytic therapeutics. Adoption could reduce reliance on animal models in early-stage drug testing, aligning with UK and EU regulatory trends favouring in vitro alternatives.
Key measures
Real-time fluorometric measurement of clot digestion; clot mass at discrete timepoints; flow rates during clot formation and digestion; shear conditions across arterial, pulmonary, and venous settings
Outcomes reported
The study describes development and characterisation of the Real-Time Fluorometric Flowing Fibrinolysis (RT-FluFF) assay, which tracks clot digestion in real-time under shear flow conditions mimicking arterial, pulmonary, and venous circulation. Measurements include continuous fluorometric tracking and endpoint clot mass determination.
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