This study was aimed to investigate the effect of prefreezing parameters such as freezing rate, annealing, rate, annealing temperature, holding time in annealing before lyophilization on lyophilized platelets by orthogonal tests. The recovery rate, the morphological and ultrastructural changes, the activation and aggregation of lyophilized platelet after rehydratation were detected by cytometer, scan electron microscopy, flow cytometry and aggregation reaction on thrombin, respectively, and complex evaluation was carried out. The results showed that the recovery rate of lyophilized platelets was 91% - 53.5% at different conditions of lyophilization, the size and shape of ice crystals in lyophilized platelets of different test groups were not similar, the expression and distribution of platelet activation markers (PAC-1 and CD62p) after rehydration were relatively similar to fresh platelets, expression rate of PAC-1 was lower (0.03% - 0.22%), meanwhile there were differences of CD62p expression levels between different groups. The optimal theoretic composition of prefreezing conditions obtained on basis of platelet recovery rate was A(2)B(1)C(1)D(3), i.e, first, suspensions of platelets on the program control cooling apparatus were lyophilized with a rate of 20 degrees C/min and maintained at -40 degrees C for 2 hours, then the annealing was conducted at -30 degrees C for 0.5 hours with a heating rate of 1.5 degrees C/min; finally, the freeze-drying program was carried out to the end. It is concluded that the freezing rate, annealing rate, annealing temperature and holding time of annealing all impact on the recovery of lyophilized platelets. The preservation qualities of lyophilized platelets were affected by the various combinations of prefreezing parameters.