The aim of our study was to optimize the cytokinesis-blocked (CB) micronucleus (MN) assay for the evaluation of radiation induced chromosomal damage in endothelial cells (EC) in vitro. After irradiation of confluent monolayers of rat and bovine aortic EC, with various doses of 250 kV X rays (0 - 3 Gy), the cells were trypsinized, resuspended in medium with cytochalasin B and then replated. After 3 days of growth they were again trypsinized and after fixing and staining of the cells, 1000 CB cells were scored for MN. The MN dose-response curves showed a rapid increase in the MN yield after low doses (<0.25 Gy) of irradiation. This points to the high radiosensitivity of EC, with rat EC being more radiosensitive than bovine EC. A further slow increase (<1 Gy) was observed after the initial fast increase. The occurrence of a fast and a slow component can be attributed to differences in radiosensitivity of EC in different stages of the cell cycle. For doses higher than 1 Gy, no further increase occurred due to severe damage at the spindle apparatus, as a result of which the division of many cells was inhibited.