Despite theoretical support linking sexual selection for elaborate coloration and behavioral isolation, few empirical examples clearly demonstrate that the sexually dichromatic signals responsible for behavioral isolation between species are, or have been, subject to sexual selection within species. The present study investigates sexual selection on male nuptial coloration in Etheostoma barrenense, a darter species for which male color has been shown in a previous study to contribute to behavioral isolation from a sympatric congener. Through the use of motorized model fish, we demonstrate that female E. barrenense discriminate between the orange and red body hues of conspecific males and exhibit an association preference for orange over red models. Combined with the results from a previous study, these data provide evidence for a link between sexual selection and behavioral isolation and by extension for a role of sexual selection in speciation. A post hoc analysis of the colors modeled in darter visual space suggests that quantifying signal values as they are perceived, rather than expressed, provide a promising way of linking sexual selection and behavioral isolation.