Among food-borne pathogens, Listeria monocytogenes is more radiation resistant than gram-negative bacteria of the genera Salmonella and Vibrio. This study was designed to determine if initial cell concentration and/or temperature at the time of irradiation influences the radiosensitivity of L. monocytogenes. Concentrations of 103, 106, and 109 CFU (colony-forming units)/ml of L. monocytogenes Scott A were suspended in tryptic soy broth and exposed to 0 to 5 kGy of gamma radiation (1.25 MeV) at 20, 4, and −80°C. Survivors were enumerated and irradiation D-values were calculated using regression analysis and total-dose methods. A 103 CFU/ml population was destroyed with a 6 CFU/ml was significantly lower (P < 0.05) than those (0.58 and 0.62 kGy) at 20° and 4°C, respectively. However, the −80°C D-value was not significantly different (0.61 kGy) when calculated by the total dose required to eliminate all recovery. At 109 CFU/ml, a D-value (calculated by both methods) of 0.42 kGy was obtained at both 4° and −80°C, which was significantly lower (P < 0.05) than 0.50 kGy for 20°C suspensions. The temperature of irradiation only influenced the radiosensitivity of L. monocytogenes at 109 CFU/ml.