Evaluating Whole Blood Clotting in vitro on Biomaterial Surfaces

Biomaterial-associated thrombosis is still a major concern for blood-contacting implants. After the medical device is implanted and comes in contact with blood, several complex reactions occur, which may lead to thrombus formation and failure of the device. Therefore, it is essential to evaluate the biomaterial interaction with the whole blood. Several studies have been reported in the literature that evaluate different steps in the coagulation cascade, such as protein adsorption, plasma activation, and platelet adhesion in vitro, however, evaluation of whole blood clotting on biomaterial surfaces is not widely reported. Here, a protocol to evaluate whole blood clotting in vitro on 2D biomaterials surfaces via a simple and fast hemolysis assay is presented. Whole human blood is placed onto the biomaterial surfaces and is allowed to clot for different time periods. After the specific time intervals, the surfaces are transferred into deionized (DI) water to release the free hemoglobin and the absorbance of this solution is measured. The absorbance value is proportional to the free hemoglobin concentration in the DI water due to lysis of red blood cells and gives an indirect correlation to the extent of blood clotting on the biomaterial surfaces. This protocol provides a fast, facile and effective method to measure the anti-thrombogenic properties of biomaterials.