This study was carried out to investigate effects of copper sulphate (CuSO4) additive to semen extenders on sperm parameters: progressive motility, viability, membrane integrity and DNA damage, after semen dilution and cryopreservation. Semen samples of 5 buffalo bulls of 3-5 years old were collected at 5 different occasions during the autumn 2011. A total number of 25 samples were used in each examination. Sperm progressive motility and viability were measured at 0 (T0), 60 (T1) and 120 (T2) min after diluting semen in tris-citric acid extender containing 0 (control), 0.004, 0.008, 0.016, 0.032 and 0.064 mg L-1 CuSO4. Later, semen was diluted in a tris-citric acid-egg yolk-glycerol extender containing the same amounts of CuSO4, cooled to 4 ˚C and kept refrigerated for 4 hr to equilibrate, sperm progressive motility, viability, membrane integrity and DNA damage were estimated. Then, semen was packed in 0.5 mL French straws and frozen in liquid nitrogen. Later, the frozen semen was thawed in 37 ˚C water bath for 30 sec, and the same parameters as well as total antioxidant capacity (TAC) of the frozen-thawed semen were estimated. The results showed that copper additive at the rate of 0.032 mg L-1 gives a better protection of sperms through the process of dilution, equilibration and freeze-thawing than that in control and other Cu concentrations, while 0.064 mg L-1 CuSO4 had deleterious effect on the sperm.