Bombesin (BNN)-like peptides have very high binding affinity for the gastrin-releasing peptide (GRP) receptor. The goal of the current study was to optimize the labeling conditions of a new Tc-99m-radiolabeled BNN-like peptide based on the bifunctional chelating ligand HYNIC using different co-ligands (EDDA and tricine). The radiolabeling conditions (pH, amount of co-ligand, amount of stannous chloride, temperature and reaction time) for newly-formed Tc-99m-tricine-HYNIC-Q-Litorin and Tc-99m-EDDA-HYNIC-Q-Litorin were optimized and evaluated by RHPLC and RTLC. Radiochemical yields for Tc-99m-tricine-HYNIC-Q-Litorin and Tc-99m-EDDA-HYNIC-Q-Litorin were 98.0 +/- A 1.7 and 97.5 +/- A 2.5%, respectively. When EDDA was used as co-ligand, the labeling of Tc-99m-EDDA-HYNIC-Q-Litorin was optimal in the following reaction mixture: HYNIC-peptide: EDDA: 10 mu g/5 mg, pH 3, SnCl2 concentration: 12 mu g/0.1 mL, reaction temperature: 100 A degrees C, reaction time: 15 min. Besides, the optimum conditions were HYNIC-peptide:tricine: 10 mu g/50 mg, pH 5, SnCl2 concentration: 12 mu g/0.1 mL, reaction temperature: 100 A degrees C, reaction time: 15 min for preparing Tc-99m-tricine-HYNIC-Q-Litorin. The manufactured Tc-99m-HYNIC-Q-Litorin conjugates may offer new possibilities for imaging cancer cells expressing bombesin receptors.