The biochemical composition of milt and the effect of its varying quantity on fertility and hatchability of Clarias gariepinus (African catfish) eggs were investigated. The study aimed at determining the right quantity of milt that can be used to fertilize certain quantity of eggs. The study further examined the effect of varying volume of milt on the water quality during the incubation of C. gariepinus eggs in order to provide baseline information on chemical composition, which can be used as a basis for sperm-mediated gene transfer. Result obtained from this study showed that 0,035ml of milt gave the best hatchability 2041±90.9, while 0.175ml of milt gave the least hatchability 1341± 384.2 .There was a positive significant correlation between the quantity of milt used and hatchability (P< 0.05) and there was a significant difference among the treatments and within the hour of the experiment (P< 0.05). The investigation revealed that the quantity of milt used for artificial reproduction in fish significantly affect water quality used for incubation (P