Inactivation of the p53 tumor suppressor gene has been implicated in the pathogenesis of numerous human cancers, including osteosarcomas. Appendicular osteosarcomas of the dog appear to be a good model for their human equivalent with regard to biologic behavior, epidemiology and histopathology. We individually screened exons 5-8 of the p53 gene for mutations in 15 canine appendicular osteosarcomas using 'Cold' SSCP to compare the role of this gene in human and canine osteosarcoma tumorigenesis. Seven of the tumors (47%) exhibited point mutations, with one tumor possessing two mutations within different exons. Of these, seven were missense mutations and the eighth was a 'silent' mutation potentially affecting the exon 6-7 splicing region. Five of the missense mutations were located in highly conserved regions IV and V, while another corresponded with the highly conserved codon 220 mutational hotspot located outside the conserved domains. The locations and types of mutations were nearly identical to those reported in human cancer. These findings provide strong evidence of the involvement of p53 mutations in the development of canine appendicular osteosarcomas. Canine osteosarcomas appear to be a promising model for their human equivalent on a clinical, pathologic, and molecular level.