D- and z-values for a mixture of four Listeria monocytogenes strains originating from the roe of different fish species were determined in rainbow trout (Oncorhynchus mykiss) roe. The D60- and D63-values obtained were 1.60 and 0.44 min, respectively, and the z-value was 5.36°C accordingly. In pilot-scale experiments, rainbow trout roe (100 g) was vacuum packaged into glass jars and pasteurized both at 62 and 65°C for 10 min. These treatments were enough to destroy 108 CFU/g of L. monocytogenes cells, which was the highest possible Listeria cell count to grow in roe. On the basis of the determined z-value and calculation of pasteurization values, these experimental pasteurizations were found to theoretically destroy at least 45 log units of L. monocytogenes cells in rainbow trout roe. In addition, these pasteurization treatments did not significantly affect the sensory quality of the roe. The sensory quality of pasteurized vacuum-packaged rainbow trout roe stored at 3°C was evaluated as good after 6 months of storage and not statistically different from the control that was frozen from the same roe lot as the pasteurized roe samples. Pasteurization of rainbow trout roe was proven to be an appropriate method for ensuring product safety with regard to L. monocytogenes and to stabilizing the sensory and microbial quality of roe. However, the safety risk caused by spore-forming bacteria still exists in pasteurized roe. Therefore, it has to be stored below 3°C.