The choice of ideal permeable cryoprotectant for cock semen equilibration and freezing is critical. The aim of this study was to compare three different cryoprotectants [dimethyl sulfoxide (DMSO), ethylene glycol (EG), and propanediol (PND)] following cryopreservation. The abdominal massaging technique was used for semen collection from 5 Venda cocks. Individual ejaculates were diluted with modified Kobidil+ (mK+) extender (fraction A) at a ratio of 1:2 and equilibrated for 2 h at 5°C. Semen samples of 5 μL were taken at 0, 60, and 120 min and evaluated by CASA for spermatozoa motility parameters (rapid, medium slow, static, NPM and PM) and velocity. Semen was further diluted at 1:1 volume ratio with mK+ supplemented with 8% DMSO, EG and PND (fraction B), and equilibrated for additional 2h at 5°C and evaluated at 60 and 120 min for motility and velocity. Cooled semen were then transferred into 0.25-mL straws and placed into a programmable freezer. The temperature of the chamber was decreased in a stepwise manner, from 5°C to -20°C, at the rate of 1°C/min until it reached the target temperature. The straws were exposed to liquid nitrogen (LN2) vapor and then plunged into LN2 (-196°C). The semen straws were stored into LN2 tank at -196°C. After 3 months of storage, semen straws were thawed at 5°C and evaluated by CASA for spermatozoa motility and velocity. Data were analyzed by ANOVA. There were no significance differences between DMSO and EG regarding the survival and motility rate of frozen/thawed semen; however, these parameters were lower compared with the fresh semen. The PND was not a suitable cryoprotectant to cryopreserve Venda spermatozoa. In conclusion, ethylene glycol was found to be a suitable cryoprotectant to cryopreserve spermatozoa of South African Venda cocks.