Commercially available human chromosome (HSA) painting probes were hybridized on river buffalo (Bubalus bubalis, 2n = 50) chromosomes by using FISH and R-banding techniques. Clear hybridization FITC-signals revealed extensive conservation of human chromosome regions in this species and demonstrated that human chromosome probes primarily paint euchromatic regions (R-bands). The present results are discussed in the light of previous gene mapping data obtained in river buffalo and ZOO-FISH data in cattle, and in relation to the standard bovine chromosome nomenclatures. In particular, HSA 8, HSA 10, HSA 11, and HSA 16+7 paint, respectively, BBU 1p, BBU 4p, BBU 5p, and BBU 24, which are homoeologous, respectively, to cattle chromosomes 25, 28, 29 and 27. Thus, these river buffalo chromosome arms can serve as markers to resolve discrepancies in the nomenclature of cattle and related species.