Technetium-99m Tetrofosmin (Tc-TF) has been shown to be useful in identifying several types of tumors, such as breast, lung and thyroid cancers. However, there are no reports in the literature about the usefulness of Tc-TF liver imaging in detecting hepatocellular carcinoma (HCC). Twenty-two patients with HCC were enrolled in this study. Tc-TF liver single photon emission computed tomography (SPECT) images were performed 10 minutes after intravenous injection of 20mCi Tc-TF. All patients were pathologically investigated by liver biopsy or surgery within one week after Tc-TF liver SPECT imaging. Among the Tc-TF liver SPECT images, 20 out of 22 (90.9%) patients showed negative results without significant Tc-TF uptake in HCC, whereas only 2 (9.1%) patients showed positive results with significant Tc-TF uptake in HCC. No significant correlation between Tc-TF liver SPECT findings and sex, age, serum alpha feto-protein level, differentiation of the tumor, or status of hepatitis virus infection was found. In this preliminary report, Tc-TF liver SPECT imaging is not a sensitive method to detect HCC. However, further studies are necessary to analyze the correlation of Tc-TF liver imaging findings and the MDR gene Pgp expression in HCC.