Freeze-drying of human platelets: influence of saccharide, freezing rate and cell concentration

Abstract

Freeze-drying of human platelets is one potentially ideal approach for long-term preservation of platelets. In this study, effects of concentration and type of saccharides, freezing rate and initial cell concentration on the recovery of freeze-dried platelets were investigated. Annexin V binding platelet activation assays, scanning electron microscopy and platelet aggregation upon thrombin (1 U/ml) addition were used to evaluate the effectiveness of platelet freeze-drying. The numerical recovery of freeze-dried platelets was reached as high as 93.0+/-5.2 percent and the recovery of nonactive platelets was reached up to 85.7 +/- 3.4 percent in the presence of 1% BSA and 20% trehalose. Frozen by shelf pre-cooling was the best way to freeze the sample in this study and the numerical recovery of freeze-dried platelets was reached 93.0 +/- 5.2 percent at about 10 degree C/min. When the platelet concentration was increased from 0.2 to 4x10(9) platelets/ml, recovery remained higher than 81.4 percent. The morphology of freeze-dried and rehydrated platelets was intact but a little rounder compared with fresh platelets. The maximum aggregation rate to thrombin (1 U/ml) of freeze-dried platelets was 83.9 percent of the fresh ones, but aggregation speed was 43.0 percent of the fresh ones. Further research on rehydration process and scale up are required.