Abstract Bovine luteal tissue slices collected at different stages of the estrous cycle were superfused and progesterone secretion was measured over a period of 8 h. The average basal progesterone secretion by 1 g of tissue ranged between 200 and 800 ng/ml medium and remained stable for 8 to 12 h after an adaptation phase of 1 h. There was no difference in the hormone secretion of tissue slices collected at different stages of the estrous cycle. Slices of luteal tissue removed from five animals on days 12 13 of the cycle were stimulated with different doses of luteinizing hormone (LH) (0.1, 1,10,100, 1000 ng/ml). The mean progesterone secretion increased to 604 ± 45,607 ± 50, 878 ± 107, 756 ± 112, and 766 ± 117 ng/ml, respectively. The hormone secretion of control samples (0 ng LH/ml medium) was 443 ± 25 ng/ml. With an LH dosage of 10 ng/ml and more, the difference as opposed to the control values was statistically significant ( P ⩽0.01). Histological and histochemical examination of the luteal tissue prior to and after the superfusion experimental revealed degenerative alterations in the central areas of the tissue slice. Peripheral parts of the tissue, however, were not affected to the same extent. Our results suggest that superfusion within a period of 8 to 12 h represents an appropriate method for the analysis of progesterone secretion in bovine luteal tissue slices.