PLATELET-AGGREGATION MONITORED IN A 96-WELL MICROPLATE READER IS USEFUL FOR EVALUATION OF PLATELET AGONISTS AND ANTAGONISTS

Optimal conditions for a method to simultaneously measure aggregation in 96 samples using a microplate reader were developed. The temperatur e of the assay was set at 25 degrees C, the optimal platelet concentra tion range was determined to be from 1-3 x 10(8) per mt, the assay vol ume was determined to be best at 100 mu L and an agitation rate of set ting #5 on the vortex was found to yield the most reliable aggregation response. After these initial assay parameters were established, EC(5 0) values for standard platelet agonists including ADP, thrombin, coll agen and thrombin receptor activating peptides were determined using t he plate assay and compared to those obtained by measuring light trans mittance in an aggregometer. The results were quantitatively similar, and qualitatively the shapes of the aggregations as monitored by both methods were characteristic of those expected for each agonist. The us e of this assay was then extended to quantitate the inhibition of aggr egation by antagonists of the fibrinogen receptor as well as by an ina ctive thrombin receptor peptide and by antibodies against the thrombin receptor. This method provided useful data for characterization of bo th platelet agonists and antagonists and should be useful for future p latelet aggregation studies.