A simple protocol was developed for the routine preparation of a kinetoplast DNA fraction from trypanosomatids. The digestion of this DNA with selected restriction endonucleases, followed by the electrophoretic analysis of the fragments on polyacrylamide gradient gels, yielded characteristic patterns that could be used for the intrinsic characterization of stocks (populations derived by serial passage in vivo and/or in vitro from a primary isolation, without any implication of homogeneity or characterization), strains (sets of populations originating from a group of trypanosomes of a given species or subspecies present at a given time in a given host or culture, and defined by the possession of one or more designated characters), and clones (trypanosomes derived from a single individual by binary fission) of certain pathogenic hemoflagellates.