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Staphylococcus aureus is a pathogen frequently associated with milk. Pulsed UV-light is a novel technology which can be used for inactivation of this pathogen in milk in a short time. Pulsed UV-light damages DNA of the bacteria by forming thymine dimmers which leads to bacterial death. This study investigated the efficacy of the pulsed UV-light system for continuous milk treatment for inactivation of S. aureus. The effect of sample distance from the UV-light source, number of passes, and flow rate were investigated. A response surface method was used for design and analysis of the experiments. Milk was treated at 5, 8, or 11 cm distance from UV-light strobe at 20, 30, or 40 ml/min flow rate and treated up to three times by recirculation of milk to find the effect of number of passes on inactivation of pathogenic microorganisms. Log10 reductions varied from 0.55 to 7.26 log10 CFU/ml. Complete inactivation was obtained in two cases and in most conditions, growth was not observed following an enrichment procedure. This research demonstrates that pulsed UV-light has a potential to be used for inactivation of S. aureus in milk.