The potential effects of the phytoestrogen genistein (GEN) on the immune system were evaluated in both F(0) (dams) and F(1) generations of Sprague-Dawley rats exposed to a soy-free diet containing low (L: 25 ppm), middle (M: 250 ppm), and high (H: 1250 ppm) levels of GEN. In dams, exposure to GEN from Gestation Day 7 to Postpartum Day 51 (totally 65 days) produced a significant increase in NK cell activity (M and H), while a decrease in the percentage of helper T cells (H). In F(1) males, exposure to GEN gestationally, lactationally, and through feed from Postnatal Days 22 to 64 (total 78 days) produced an increase in the relative weights (% body) of spleen (L and H) and thymus (L). Furthermore, exposure to GEN increased the number of splenic B cells (H), T cells (L, M, and H), and T-cell subsets (L, M, and H). Although GEN decreased the percentages of splenic NK cells (L, M, and H), no effect on the activity of NK cells was observed. In F(1) females, exposure to GEN produced a decrease in terminal body weight (H), with an increase in the relative weight of spleen (L, M, and H). Exposure to GEN also increased the number of splenic B cells (L), macrophages (L and M), T cells (H), helper T cells (L and H), and cytotoxic T cells (M and H). Additionally, exposure to GEN increased the percentages of T cells (M and H), helper T cells (H), and cytotoxic T cells (M and H). Moreover, the spleen IgM antibody-forming cell response to sheep red blood cells was enhanced (H), although the percentages of B cells were decreased (M and H). No effect on the activity of NK cells was observed; however, the percentages of splenic NK cells were decreased by GEN (L and H). In conclusion, these results demonstrate that exposure to GEN can modulate the immune responses in Sprague-Dawley rats. Furthermore, the sexual dimorphic effects of GEN in F(1) male and female rats suggest that there may be interactions between GEN and the responses modulated by sex hormones.