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Not AvailableCryopreservation helps in prolonging the storage of semen but freezing and thawing damage the sperm because of alteration in the structure and physiology of the sperm cell. Buffalo bull sperm were treated with different levels of cholesterol loaded cyclodextrin (CLC) and frozen in tris egg yolk glycerol diluents to determine the best suitable concentrations of CLC that benefits Buffalo sperm cryosurvival. Semen collected from Buffalo bull was diluted to 120×106 spermatozoa/ml by tris diluents and divided into four groups; control (without CLC), 0.5, 1.0 and 2.0 mg CLC/ml of diluted semen as Group I, II, III and IV respectively. At prefreeze stage, the sperm treated with 2.0 mg CLC showed significantly (P<0.01) higher values for progressive motility, viability, acrosomal integriry and HOS response. No significant difference was observed between groups I, II and III in seminal parameters at pre-freeze stage except HOS response. After thawing, significantly (P<0.01) higher percentage of progressive motility, viability, acrosomal integrity and HOS reponse was observed in spermatozoa treated with 2.0 mg CLC than any other treatment groups including control. Group I and II showed no significant (P