This study assessed the influence of gender on attitudes about bisexuals. A total of 164 heterosexual female and 89 heterosexual male undergraduates completed the Biphobia Scale (Mulick & Wright, 2002), rewritten to refer to bisexual men and bisexual women and thus re-named the Gender-Specific Binegativity Scale. A mixed-design ANOVA revealed an interaction between rater's sex and target's sex: women equally accepted bisexual men and bisexual women, but men were less accepting of bisexual men than bisexual women. A mediation analysis indicated the relationship between rater's sex and greater acceptance of bisexual women was partially explained by eroticization of female same-sex sexuality. Finally, participants also responded to two open-ended items, which provided additional information about the content of binegativity: participants described male bisexuals negatively, as gender-nonconforming, and labeled them "really gay," whereas participants described female bisexuals positively, as sexy, and labeled them "really heterosexual." These findings suggest multiple underlying beliefs about bisexuals that contribute to binegativity, particularly against bisexual men. Results also confirm the importance of considering gender (of both the target and the rater) when assessing sexual prejudice.