The overall quality of platelets for transfusion is not considered satisfactory by most researchers in the field. The problem is due, in part, to the difficulty of collecting and storing these highly reactive cells, and also to the lack of adequate in vitro methods for assessing quality. We have sought to bring methods of modern cell biology to the investigation of the cellular and biochemical basis of the platelet storage lesion. We have available several very sensitive methods for identifying storage-induced changes in the platelet membrane, the membrane skeleton and cytoskeletal proteins. Characterization of protein patterns has been achieved by one- and two-dimensional SDS-PAGE and by immunoblotting using a variety of monoclonal antibodies to cytoskeletal and membrane proteins. Flow cytometry has proven to be a very sensitive technique for monitoring activation of platelets and the mobilization of glycoprotein receptors in response to agonists. A method for analysis of activation-dependent changes in platelets sampled directly from platelet concentrates has been developed and includes formalin-fixation of platelets diluted in plasma. Washing steps have been eliminated from this procedure, allowing platelets to be analyzed directly without centrifugation which may cause additional activation. Fluorescently-labeled antibodies to P-selectin (CD62) and p53 (CD63), a lysosomal granule protein, allow rapid, direct detection of activation. Regulation f glycoprotein expression under conditions of agonist-induced activation has been monitored with antibodies to GPIb, GPIb/IX complex, GPIIb, and GPIIb/IIIa complex. In addition, the amount of filamentous actin, another indicator of activation, is measured with the sensitive probe, rhodamine-labelled phalloidin. The platelet activation state will be correlated with morphological changes demonstrated by phase microscopy and with any alterations in aggregation response to agonists. Results of a platelet storage study were presented at a symposium - """"""""The Cellular and Molecular Basis of the Platelet Storage Lesion """""""", held in April, 1991 and were published in March, 1992.
