Development of flow cytometry for sorting mammalian sperm according to their sex chromosomes began in the late 1970s and early 1980s. This technology, which has recently been commercialized for bovine sperm, is based on the differences in DNA content between X- and Y-chromosome-bearing sperm. Under ideal conditions, 5000 live bovine sperm of each sex can be sorted per second at 90% accuracy. Pregnancy rates of 50% have been achieved routinely in well-managed heifers with sex-sorted, cryopreserved bovine sperm compared to 60–80% with unsexed control sperm. About 90% of offspring have been of the selected sex. Sorting sperm according to sex chromosome content is similarly successful in many other mammals including exotic species, but sorting efficiencies are somewhat less for sperm from some species. Key words: Mammals, sex chromosomes, flow cytometer, cell sorter, DNA content, X and Y sperm, Hoechst 33342