Abstract：For starry flounder (Platichthys stellatus), the male is less than the female and the volume of sperm is few, which limits the development for artificial reproduction and crossbreeding. In present study, the series of selecting experiments for sperm diluents, the osmotic pressure of diluents, the suitable temperature of seawater and the suitable dilution degree of frozen sperm were selected using the mature starry flounder. In order to check the application effect of frozen sperm, the experiments of fertilization and hatchability with starry flounder eggs were made. and made comparison with control group (fresh sperm). The results were analyzed by One-way ANOVA and Student-newman-keuls from SPSS 11.5. The results indicated that the frozen sperm motility was the highest in Ts-2 (37.65 g/L sucrose+10.01 g/L KHCO3+1.21 g/L Tris alkali) and SFs-4 (60 g/L glucose+10.01 g/L KHCO3+1.21 g/L Tris alkali) with (56.67±5.78) % and (66.67±5.77) %, respectively. It was also used computer aided sperm analyse(CASA) to analyze and test for motion parameters, (curvilinear velocity(VCL), straight line velocity(VSL), beat cross frequency(BCF), linearity(LIN), straightness(STR)) from frozen sperm sample in Ts-2 and SFs-4. The results indicated the motion parameters of frozen sperm with SFs-4 significantly higher than the ones in Ts-2. Using sucrose, KHCO3, and Tris alkali to prepare different osmotic pressure of sperm diluents, the frozen sperm motility was the highest (61.67±5.00)% in 319.94 Pa and it was no significant difference compared with fresh sperm (P＞0.05). In the experiment of selecting of the suitable temperature of seawater, the frozen sperm were activated with 3~18 ℃ seawater, theit showed the sperm motility was the highest at 12 ℃ and there was no significant difference compared with fresh sperm (P＞0.05), but there was significant difference between others and fresh sperm (P＜0.05). For the dilution degree of the post-thaw sperm was diluted for 10~40 time, and fertilized with eggs from starry flounder, it indicated that the frozen sperm rate of fertilization was the highest in the 20 times diluted, then it was stated the different dilution degree of frozen sperm had certain effect on fertilization ratio. In present study, the sperm from starry flounder could be effective cryopreserved with Ts-2 +20%DMSO and SFs-4+20% DMSO. This research result firstly provide important the theory and technology basis.
