Rapid reactions of platelets studied by a quenched-flow approach: aggregation kinetics

Understanding the nature and regulation of very early events in the platelet activation sequence is an important goal. A general approach based on quenched-flow principles has therefore been developed. Platelet reactions are initiated by mixing inducing agent with whole blood, PRP, or washed platelets and pumping the mixture through narrow-bore tubing with quenching (stopping the reaction) at the outlet. Reaction times less than 1 sec are feasible. The quenched-flow system has been combined with a continuous-flow modification of resistive-particle counters to follow aggregation kinetics of "single" particles. Aggregation at 37 degrees is extremely rapid: after a lag period of about 1 sec, between 20% and 50% of platelets then aggregate per second (10 micro M ADP). Although the kinetics are second order, data can be expressed as the percent of "single" platelets aggregating per second and treated as pseudo-first order. This enables derivation of apparent maximal velocities of aggregation and inducer activation constants, for characterizing platelet reactivity.