The authors have established an in vitro chemosensitivity assay for human osteosarcoma. To overcome the difficulty of maintaining primary osteosarcoma cells in culture, biopsy or surgical specimens were transplanted directly into athymic nude mice and early generation xenografts were used for in vitro scintillation assay. These xenografts enabled a large number of viable tumor cells to be obtained even from small biopsy specimens. As a result, this system afforded a workable assay in 21 (62%) of 34 osteosarcomas tested and enabled the assay of numerous drugs at 4 different concentrations. Reproducibility of the assay can be checked by using serially transplanted xenografts. Retrospectively, the in vitro results were compared with the histologic response of the resected tumor in 18 patients who had received preoperative chemotherapy. The true positive rate, true negative rate, and predictive accuracy were 40%, 100%, and 66.7%, respectively. The authors suggest that this system provides a valid tool to determine which drugs are ineffective in individual osteosarcoma patients and thus may allow more efficient therapy.