The brain areas perfused by superselective intra-arterial (i.a.) chemotherapy were assessed using single photon emission computed tomography (SPECT) with technetium-99m-hexamethyl-propyleneamine oxime (99mTc-HMPAO). A superselective catheter was introduced into the anterior, middle, or posterior cerebral artery of patients with malignant glioma for i.a. chemotherapy. 99mTc-HMPAO was subsequently injected via the same catheter used for chemotherapy, and a higher dose of 99mTc-HMPAO was injected intravenously to obtain adequate background brain images. Comparison of the SPECT images with magnetic resonance images could confirm complete perfusion of the tumor tissue. In two patients with malignant glioma, regions of interest were selected in the peritumoral brain area and a reference brain area, and the radioactivity was measured. The concentration of 99mTc-HMPAO was about 50 times higher in tissue perfused by superselective injection into anterior or middle cerebral artery compared to intravenous injection. 99mTc-HMPAO SPECT is readily available in many institutions and the information provided is useful for planning more effective and safe i.a. chemotherapy.