The chemokine CXCR4 receptor is over-expressed in a wide variety of tumors. In this study, AMD3100, which was a prototype non-peptide antagonist of CXCR4 receptor, was labeled with 99mTc. 99mTc–AMD3100 was verified by thin layer radio chromatography (TLRC). The tumor-localizing properties of 99mTc–AMD3100 were evaluated and proved in mice bearing Hep-G2 tumor xenograft. 99mTc–AMD3100 was a promising, novel receptor-specific radiopharmaceutical with potential application in the imaging of human tumors.