^99mTc-HYNIC-Annexin V was prepared for apoptosis radionuclide imaging in vivo. The expression plasmid of human Annexin V was constructed and expressed in E. coli.. HYNIC was prepared and used as bifunctional chelating agent for ^99mTc labeling. The radiochemical purity, radiolabeling yield and stability of ^99mTc-HYNIC-Annexin V were studied. The coding sequence of human Annexin V gene was successfully cloned into the expression vector, and highly expressed in E. coli. A protein with a molecular weight of about 36kD could be induced and confirmed by Western Blotting and SDS-PAGE. Annexin V could be labeled with ^99mTc using HYNIC, and showed high radiochemical purity of more than 90%. The radiolabeling yield could reach more than 90%. The reaction condition was moderate. It suggested that ^99mTc-HYNIC-Annexin V may be a promising agent for apoptosis imaging and clinical application.