The purpose of this study was to investigate the correlation between histopathological changes and 201Tl accumulation in tumor following chemotherapy in the evaluation of chemotherapeutic effect. Walker 256 carcinoma implanted in the right thighs of Wistar rats was treated by the intravenous injection of adriamycin (ADR). 201Tl scintigraphy was performed before and 12, 24, 72, and 96 hours after chemotherapy. The count ratio of tumor to normal muscle (T/N ratio) was measured by gamma camera. Tumor growth rate was measured and histopathological examination of the tumor after 201Tl scintigraphy was performed. The tumors of all rats disappeared within 2 weeks after chemotherapy, but the T/N ratio did not change. Histopathological findings showed a decrement of tumor cells, small necrotic regions, and remarkable lymphocyte infiltration in the tumor bed in the healing stage. After 201Tl injection, lymphocytes isolated from tumor tissue showed noted 201Tl accumulation. These results suggest that the T/N ratio did not decrease after ADR chemotherapy because of the accumulation of 201Tl in lymphocytes. We concluded that the use of 201Tl may underestimate the therapeutic effect of ADR.