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American Journal of Pathology, Vol 79, 81-94, Copyright © 1975 by American Society for Investigative Pathology

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Characterization of human platelets separated from blood by ADP-induced aggregation

SF Mohammad, RL Reddick and RG Mason

Separation of platelets from plasma is achieved by adding ADP (final concentration 10-5 M) to platelet-rich plasma and allowing aggregates to form. Aggregates are removed quickly by brief, gentle centrifugation, washed two to three times with 0.9% NaCl (saline), and then incubated for 10 minutes in the presence of apyrase, albumin and calcium. Platelet aggregates deaggregate completely during this incubation period. The platelet suspension is then subjected to 1100g for 12 minutes, gently resuspended in a small volume of saline, and finally diluted with an appropriate medium to the desired concentration. The entire separation procedure requires approximately 30 minutes. Platelets obtained by this procedure are a) comparable in aggregability to the platelet preparations obtained by gel filtration, b) have normal intracellular amounts of ATP and ADP, and c) except for slight dilatation of the surface-connected canalicular system, have normal ultrastructural appearance. When suspended in an appropriate medium, these separated platelets take up serotonin 14-C and subsequently release it in nearly normal quantities when exposed to thrombin, collagen or ADP.