ACTIVATION DURING PREPARATION OF THERAPEUTIC PLATELETS AFFECTS DETERIORATION DURING STORAGE - A COMPARATIVE FLOW CYTOMETRIC STUDY OF DIFFERENT PRODUCTION METHODS

Three different separation methods, all using centrifugation, are rout inely used to prepare therapeutic platelet concentrates from human don or blood. Platelet concentrates derived from platelet-rich plasma (PRP -PC), buffy coat (RC-PC) and apheresis (AP-PC) were investigated at th e end of production, and over an 8d storage period. Change in platelet surface markers were measured by flow cytometry, using fluorescein-co njugated antibodies to fibrinogen, P-selectin (CD62P), GPIIb-IIIa (CD4 1), GPIb alpha (CD42b) and GPV (CD42d), and fluorescein-conjugated Ann exin V was used to measure expression of anionic phospholipid. All con centrates showed some changes during preparation but PRP-PC underwent the greatest changes with significantly higher levels of P-selectin (P < 0.001) and bound Annexin V (P = 0.001) than AP-PC or BC-PC, and low er levels of GPIb alpha (P = 0.002) and GPV (P < 0.001). These changes were attributable to component separation rather than venesection. Th ese markers all continued to change on storage with a strong positive correlation between the changes seen during production and those after 5d storage. PRP-PC continued to show the greatest changes whereas BC- PC showed the least. Fibrinogen was bound to 40-50% of platelets in al l preparations and this did not alter significantly on storage whereas total expression of GPIIb-IIIa remained unchanged throughout. There w as no evidence that the platelet surface changes were thrombin-mediate d and leucocyte depletion of BP-PC by filtration had no effect on the changes. It is proposed that the deterioration of platelet concentrate s during storage may be related to activation occurring during prepara tion. 'Whole blood' flow cytometry using a panel of fluorescein-labell ed reagents provides an informative method for evaluating platelet con centrates.