Male infertility is responsible for approximately 50% of infertility worldwide. Reactive oxygen species are one of the major causes of male infertility. In this study, the effects of oxidative stress induced by tertiary-butyl hydroperoxide (TBHP) on sperm quality and testis tissue are investigated. After determination of LD50, TBHP with a concentration of 1 : 10 LD50 was injected in adult male mice strains Balb/c for two consecutive weeks. Their testis tissues were used for cell viability, histopathology analysis and ROS assay. The epididymis was also surveyed for sperm analysis by CASA system. The sperm motility, count and viability decreased in the TBHP-treated mice compared to the control mice. The flow cytometry analysis showed a significant increase in H2O2 and Symbol levels in both testis and sperm within 2 weeks after intraperitoneal injection. Body weights revealed no treatment-related effects, but atrophy of testis and a decrease of testis cells viability were observed. The results showed that exposure to TBHP could lead to morphological changes in seminiferous tubules. TBHP-induced oxidative stress caused a decrease in sperm parameters and testis cells viability. That is due to an increase level of ROS in the testis and their deleterious effects on genomic levels.