The influence of diluents, equilibration time and prefreezing storage time on the viability of cryopreserved Oreochromis niloticus (L.) spermatozoa

. This paper reports a series of trials on the cryopreservation of Oreochromis niloticus (L.) spermatozoa, using methanol as the cryoprotectant. Immotile milt samples pooled from four males were diluted with two diluents, each being subjected to equilibration periods of 15, 30, 45, 60 and 90 min. In addition, samples of fresh pooled milt were kept for 0, 2, 4, 6 and 8 days at 4°C prior to cryopreservation. Diluted samples were stored in 250-μl plastic straws and cooled to -50°C at 5°C/min and held under liquid nitrogen for between 1 and 3 weeks. Viability of post-thawed spermatozoa was estimated from video recordings of samples activated under a microscope and from fertilization rates of eggs. The type of diluent and its interaction with equilibration time significantly (P< 0.05) influenced the post-thaw motility of spermatozoa. Spermatozoa stored for up to 6 days prior to cryopreservation were as fertile as freshly frozen spermatozoa and yielded 60.6 (±SE, 8.4) developing embryos.