Recent advances in the chemistry of radiolabeling with 99m Tc such as use of the 99m Tc tricarbonyl and 99m Tc-HYNIC cores have revived interest in 99m Tc-labeling of small biomolecules and further investigation as potential radiopharmaceuticals. Isoniazid, a drug commonly used for treatment of tuberculosis, has been chosen for the present study. Three distinct strategies were utilized to radiolabel isoniazid with 99m Tc. In the direct labeling protocol, the hydrazino amide functional group of isoniazid was used for 99m Tc-labeling in the HYNIC sense using tricine and EDDA as co-ligands. The other strategies of 99m Tc-labeling involved the derivatization of isoniazid to its N, N-diacetic acid derivative, which in turn was either used as a tridentate ligand for labeling with the [ 99m Tc(CO) 3 (H 2 O) 3 ] + synthon or directly labeled by the conventional route wherein 99m Tc is in the +3 oxidation state. The complexes prepared in >95% yields were characterized by paper chromatography, thin layer chromatography and HPLC. Comparison of the three approaches showed that maximum specific activity and stability was obtained in the 99m Tc-isoniazid derivative synthesized via the tricarbonyl method. However, in vitro cell-binding studies indicated that none of the 99m Tc-isoniazid complexes prepared had any appreciable uptake in Mycobacterium tuberculosis.