An Ex Vzvo Method for the Evaluation of Biomaterials in Contact with Blood

The use of extracoporeal (ex vivo) shunts for studying thrombus formation was given impetus by a number of studies from J. F. Mustard’s 1aboratory.lS In these studies the essential problems of this method were recognized and the requirements for a reproducible system were delineated. The basic features are: 1) uniform blood flow, 2 ) a chamber that represents preferential region for thrombus formation, and 3 ) access to the thrombus for analysis. Murphy et a1.l tried a number of flow chamber designs and finally settled on the use of a model bifurcation for their studies,2 and other investigators s , have evaluated alternative flow chamber configurations. This type of ex vivo test with recirculation of blood should be distinguished from other ex vivo procedures in which animals are bled through a test chamber.5, In the latter cases chronic shunts are not used, which simplifies the procedure, but the loss of blood usually limits the duration of the experiment to less than one hour. To develop an ex vivo method for the evaluation of the thrombogenicity of materials in contact with blood, a system was devised that exposes a test chamber to a dog’s systemic circulation by means of a chronic arteriovenous shunt. The general aspects of this approach are shown in the flow diagram in FIGURE 1 and the schematic FIGURE 2. Male dogs were selected after screening to ensure that they were normal with respect to weight and blood coagulation values. An arteriovenous shunt was implanted in accepted animals, and after surgery, coagulation factors and other physiological parameters were evaluated periodically to monitor the approach of the dog’s coagulation system toward steady state. After about one week, labeled fibrinogen and platelets were introduced systemically, and after 24-48 hours the chamber containing the test material was introduced into the bypass system. The rate of thrombus formation was determined by monitoring the buildup of radioactive fibrinogen and platelets in the test chamber. Blood flow through the chamber was measured with an electromagnetic or Doppler flow meter and controlled at a constant value. After a predetermined time the chamber was disconnected from the shunt, and any thrombus formed was removed and quantitated in terms of clot weight, moisture, fibrinogen, red cell, and platelet content. These values are used as indices of thrombus formation on a given biomaterial under defined flow conditions.