A flow microfluorometer was used to measure metaphase chromosomes in suspension at rates up to 100,000 per min. Chromosomes from cells of the Chinese hamster M3-1 cell line were isolated, stained for DNA with the fluorescent dye ethidium bromide, and analyzed for DNA content. Nine distinct peaks were resolved that correspond well with independent chromosomal DNA measurements made with a high-resolution scanning cytophotometer. Chromosomes were sorted from each peak by an electronic cell sorter. Visual examination of each fraction indicated the purity of the sorted chromosomes. This novel technology allows separation of purified populations of individual chromosomes suitable for biochemical and biological characterizations.