Large non-luteinized follicles of the marmoset monkey were cultured for up to 96 h in the presence of substances that are known to induce luteinization, i.e. LH, transforming growth factor (TGF)-beta and cyclic AMP. The state of the basal lamina, and the expression of connexin-43, alpha(2) integrin subunit and TGF-beta receptor type II (TbetaR-II) were chosen as parameters to judge the progress of luteinization. Antral follicles, cultured for 1 h, were not luteinized, as shown by an intact basal lamina, strong immunoreactivity of connexin-43 in granulosa cells, and no expression of TbetaR-II in the theca layer. After 12 h, most follicles showed a dissolution of the basal lamina, a faint reactivity of connexin-43, high expression of TbetaR-II in theca- and outer granulosa cells and high expression of alpha(2) integrin subunit in granulosa cells bordering at the basement membrane; all of which indicate luteinization. After 96 h of culture, luteal structures (e.g. corpora lutea accessoria) had developed. This was true for both non-stimulated and stimulated follicles. Our results strongly suggest that antral follicles luteinize spontaneously. The decisive determinant appears to be the follicular stage.