A novel gene, TSG23/Tsg23, was identified by comparing the expression profiles of human adult and fetal testis using Affymetrix Genechips. RT-PCR analysis from multiple human and mouse tissues indicated TSG23/Tsg23 mRNA was mainly expressed in the testis. In situ hybridization revealed that TSG23/Tsg23 mRNA was located in spermatocytes and round spermatids of the seminiferous tubules in human and mouse testis. To further confirm the result from RT-PCR, the antibody for human TSG23 was generated against the protein encoded by the gene. Western blot analysis demonstrated that TSG23 was mainly expressed in human testis, with a molecular weight of about 23 kDa. Immunohistochemistry showed that TSG23 was predominantly located in spermatocytes and round spermatids, consistent with the results from in situ hybridization. In order to explore the function of TSG23 in spermatogenesis, the study compared the expression of TSG23 in the testis from fertile persons and from patients with azoospermia. The results showed that there was less expression in patients with obstructive azoospermia compared with fertile persons, and no detectable TSG23 at mRNA and protein levels in patients with non-obstructive azoospermia. The expression of Tsg23 mRNA was considerably decreased in a time-dependent manner in the testis of an azoospermic mouse model induced by Busulfan. These data suggest that TSG23/Tsg23 is involved in human and mouse spermatogenesis.