The effects of short-term high-intensity exercise on single fiber contractile function in humans are unknown. Therefore, the purposes of this study were: (a) to access the acute effects of repeated high-intensity exercise on human single muscle fiber contractile function; and (b) to examine whether contractile function was affected by alterations in the redox balance. Eleven elite cross-country skiers performed four maximal bouts of 1300 m treadmill skiing with 45 min recovery. Contractile function of chemically skinned single fibers fromtriceps brachiiwas examined before the first and following the fourth sprint with respect to Ca2+ sensitivity and maximal Ca2+-activated force. To investigate the oxidative effects of exercise on single fiber contractile function, a subset of fibers was incubated with dithiothreitol (DTT) before analysis. Ca2+ sensitivity was enhanced by exercise in both MHC I (17%,P< 0.05) and MHC II (15%,P< 0.05) fibers. This potentiation was not present after incubation of fibers with DTT. Specific force of both MHC I and MHC II fibers was unaffected by exercise. In conclusion, repeated high-intensity exercise increased Ca2+ sensitivity in both MHC I and MHC II fibers. This effect was not observed in a reducing environment indicative of an exercise-induced oxidation of the human contractile apparatus.