The ability to segregate simultaneously occurring sounds is fundamental to auditory perception. Many studies have shown that musicians have enhanced auditory perceptual abilities; however, the impact of musical expertise on segregating concurrently occurring sounds is unknown. Therefore, we examined whether long-term musical training can improve listeners' ability to segregate sounds that occur simultaneously. Participants were presented with complex sounds that had either all harmonics in tune or the second harmonic mistuned by 1%, 2%, 4%, 8%, or 16% of its original value. The likelihood of hearing two sounds simultaneously increased with mistuning, and this effect was greater in musicians than nonmusicians. The segregation of the mistuned harmonic from the harmonic series was paralleled by an object-related negativity that was larger and peaked earlier in musicians. It also coincided with a late positive wave referred to as the P400 whose amplitude was larger in musicians than in nonmusicians. The behavioral and electrophysiological effects of musical expertise were specific to processing the mistuned harmonic as the N1, the N1c, and the P2 waves elicited by the tuned stimuli were comparable in both musicians and nonmusicians. These results demonstrate that listeners' ability to segregate concurrent sounds based on harmonicity is modulated by experience and provides a basis for further studies assessing the potential rehabilitative effects of musical training on solving complex scene analysis problems illustrated by the cocktail party example.