This study investigated the effect of Matrigel on the differentiation and proliferation of luteinized granulosa cells in vitro. Human luteinized granulosa cells (LGC), obtained after the oocyte retrieval from the IVF process were cultured either on plastic or on Matrigel. Phase-contrast microscopy showed that LGC were flat and attached on the surface of poly-D-lysine while they formed three dimensional cell aggregates when cultured on Matrigel. Bromodeoxiuridine (BrdU) labeling followed by immunocytochemistry for BrdU and 3b-hydroxysteroid dehydrogenase (HSD) demon-strated that BrdU-labeld cells were shown in both of the 3b-HSD positive and negative granulosa cells when cells were cultured on Matrigel and on poly-D-lysine. Progesterone secretion was increased until culture day 4, and then slightly decreased. The difference in progesterone secretion between the cells cultured on Matrigel and those on poly-D-lysine was increased, as the culture day was increased. The results suggested that Matrigel provides a better culture environment for LCG and maintains proliferation of LCG.