Nineteen “female” Holstein-Friesian calves born co-twin with a bull were purchased shortly after birth. One subsequently was judged to be a normal female with 0 % XY (100 % XX) karyotypes in cultures of blood lymphocytes. However, one typical freemartin also had 0 %XY (100 % XX) karyotypes in 300 cells when examined at three ages. Overall, 18 freemartins examined at 1, 24, and 52 weeks of age averaged 60.7 %, 57.9%, and 55.5 % XY cells, respectively (P > 0.05). No systematic relationship was found between the proportion of XY cells and the abnormalities of the reproductive organs, as judged by vaginal depth and clitoral development at 1, 24, and 52 weeks, and examination of the reproductive organs per rectum at 54 weeks of age. The clitoris of untreated freemartins and those treated with estrone or estradiol postnatally did not enlarge. Testosterone treatment caused enlargement. None of the hormones affected vaginal depth. These studies provide convincing evidence that the blood lymphocyte XX:XY karyotype ratio (1) is not related to the degree of masculinization and (2) is stabile in the postnatal freemartin. Thirdly, the freemartin reproductive organs do not respond to common sex-steroid hormones postnatally other than the clitoral response to testoster