In this study, the effects of freezing of testicular tissue by slow freezing on the proliferation rate, colonization and viability of spermatogonial stem cells (SSCs) were evaluated. The testicular tissue of the freezing group was freezed by slow-freezing method, and the fresh group remained intact. The results showed that the cell proliferation rates of fresh group [1.48 ± 0.02] and [1.77 ± 0.02] compared to the frozen-thawed group [1.39 ± 0.02] and [1.64 ± 1.64] in the 7th and 14th days, were significantly higher. There was significant difference between the number of colonies in the 7th and 14th days. In the first day after cell isolation, cell viability of the fresh group [95 %] was significantly higher than the frozen-thawed group [87.7 %]. Also, in the 7th and 14th day, cells viability fresh group were significantly more than frozen-thawed group. These results suggest that slow freezing of testicular tissue reduces the proliferation, colonization and survival of SSCs.