The effect of in-vitro culture on the motility and morphology of fresh and frozen-thawed human testicular spermatozoa obtained from obstructive azoospermic patients and on the motility of testicular spermatozoa obtained from nonobstructive azoospermic patients was evaluated, The outcome of intracytoplasmic sperm injection (ICSI) with fresh and frozen-thawed human testicular spermatozoa was studied, The results showed that significant improvement of sperm morphology and motility was observed in culture of fresh (n = 17) and frozen-thawed (n = 15) testicular sperm samples obtained from patients with obstructive azoospermia, The motility of cultured testicular spermatozoa reached a peak at 72 h without the need for special media, In six of 20 samples obtained from patients with non-obstructive azoospermia, improvement of sperm motility was observed, When only non-motile testicular spermatozoa mere cultured, they all remained non-motile (n = 9). In patients with obstructive azoospermia, fertilization rates of 80 and 81% were obtained using ICSI with fresh and frozen-thawed testicular spermatozoa respectively, Clinical pregnancies were observed in four out of nine patients with fresh testicular spermatozoa and two out of five patients after using frozen-thawed spermatozoa, When fresh testicular spermatozoa obtained from patients with non-obstructive azoospermia were used for ICSI, the fertilization rate was 68% and two out of seven patients achieved clinical pregnancies, In conclusion, the morphology and motility of fresh and frozen-thawed testicular spermatozoa in patients with obstructive azoospermia can be significantly improved after in-vitro culture, The outcome of in-vitro culture of testicular spermatozoa in patients with non-obstructive azoospermia is unpredictable. In-vitro culture of non-motile testicular spermatozoa is not successful so far, The outcome of ICSI with fresh and with frozen-thawed testicular spermatozoa was similar.