A rapid and sensitive method for the identification of human tissues xenotransplanted in nude mice was developed. An in situ hybridization technique made it possible to distinguish between cells of human origin and cells of murine origin in formalin-fixed paraffin sections. High-molecular-weight DNAs extracted from human or mouse tissues were sonicated, nick-translated with 32P-dCTP, and used as hybridization probes. Dot blot hybridization of 32P-labeled probes revealed clear species-specific signals. Formalin-fixed paraffin-embedded tissue samples from repopulated tracheal transplants, containing either human tracheal epithelial cells or human renal tubular cells, were used. Cells of human and murine origin were distinguishable by in situ hybridization with sonicated DNA probes. This method has several advantages; simple preparation of probes, high sensitivity, and applicability to formalin-fixed paraffin-embedded tissue sections. In situ hybridization with sonicated DNA probes should provide a powerful tool for verifying the human origin of xenotransplanted tissues in nude mice.