The role of balancing selection in maintaining genetic variation for fitness is largely unresolved. This reflects the inherent difficulty in distinguishing between models of recurrent mutation versus selection, which produce similar patterns of inbreeding depression, as well as the limitations of testing such hypotheses when fitness variation is averaged across the genome. Signatures of X-linked overdominant selection are less likely to be obscured by mutational variation because X-linked mutations are rapidly eliminated by purifying selection in males. Although models maintaining genetic variation for fitness are not necessarily mutually exclusive, a series of predictions for identifying X-linked overdominant selection can be used to separate its contribution from other underlying processes. We consider the role of overdominant selection in maintaining fitness variation in a sample of 12 X chromosomes from a population of Drosophila melanogaster. Substantial variation was observed for male reproductive success and female fecundity, with heterozygous-X genotypes exhibiting the greatest degree of variance, a finding that agrees well with predictions of the overdominance model. The importance of X-linked overdominant selection is discussed along with models of recurrent mutation and sexually antagonistic selection.