Chromosomes sampled from seven human populations were analyzed by flow cytometry to survey normal variation in chromosome size. The populations include two African Pygmy groups, two Amerindian tribes, Druze, Khmer Cambodians, and Melanesians. Mitotic chromosomes were isolated from cultured cells and stained with Hoechst 33,258 and chromomycin A3. The relative DNA content and base-pair composition of each homolog was quantified by bivariate flow karyotyping. Significant variation in DNA content, ranging from 10-40%, was observed for chromosomes 1, 13-16, 19, 21, 22, and Y. The measurements for each population appeared to be a random sampling of the total set of 33 individuals for the majority of chromosomes. A few significant differences in the distributions of chromosomal DNA content were observed among the populations, however. The data, when combined with an earlier study of 33 unrelated individuals of unknown ethnic origin, provide a good representation of the variation in chromosome size among humans.