Abstract: Acanthopagrus latus is one of the important breeding species in China, whose sperm cryopreservation can provide technical supports for its breeding, prevent the decline of its germplasm resources and maintain its sustainable development of the breeding industry effectively. In this experiment, we screened the diluent, glucose concentration, antifreeze type and concentration as well as dilution ratio and cooling procedure of the spermatozoa of A. latus. The results show that when the sperm were diluted with MPRS solution containing 10 g·L⁻¹ glucose and 5% ethylene glycol at the ratio of 1∶2, balanced at 4 ℃ for 30 min, fumigated 5 cm on the liquid nitrogen surface for 5 min, and finally stored in liquid nitrogen for 2 h, the frozen sperm had the best motility after thawing in 37 ℃ water bath, and the sperm motility, motility time and life span could reach (85.170±3.66)%, (9.16±70) s and (94.297±9.55) s, respectively.