Immunolocalization of factor VIII (a specific endothelial cell marker) and heparin-binding (fibroblast) growth factors (HBGF)-1 and HBGF-2, along with quantitative image analysis, were used to evaluate vascular development and distribution of HBGF in the bovine corpus luteum (CL) at three stages (early, middle, and late) of the estrous cycle. Luteal vascularity increased from the early to the middle stage and then declined to the late stage. During the early stage, most of the microvessels were present in the cores of the tissue infoldings (presumably thecal-derived areas), but numerous capillary sprouts could be seen invading the parenchymal areas. During the middle stage, capillary density was so great that most parenchymal cells were in contact with one or more capillaries. At the late stage, relatively few capillaries were present in the parenchymal areas, whereas larger microvessels were prominent throughout the CL. Throughout the estrous cycle, HBGF-1 and HBGF-2 were present primarily in the cytoplasm of large and small luteal cells. For each stage of the estrous cycle, HBGF-2 staining was greater than that of HBGF-1. Relatively high levels of HBGF-2, but not HBGF-1, were also present in connective tissue tracts. In addition, HBGF-1 and HBGF-2 appeared to co-localize in some luteal cells. Although the distribution of HBGF-1 was homogeneous throughout the CI, that of HBGF-2 was heterogeneous. For HBGF-1, staining increased from the early to the middle stage but remained unchanged from the middle to the late stage. In contrast, HBGF-2 staining was greatest in the middle stage and similar in the early and late stages. These data are the first report of HBGF-1 and HBGF-2 immunolocalization in bovine luteal tissues throughout the estrous cycle, and demonstrate that HBGF-2 staining follows a pattern similar to that of luteal vascular development.