Homophobia and heterosexism in women’s collegiate athletics has been predominantly researched using qualitatively methodology (e.g., Blinde & Taub, 1992; Kauer & Krane, 2006; Krane 1996, 1997; Krane & Barber, 2003). Few researchers have examined the heterosexist attitudes from a quantitative perspective and few with a sport—specific questionnaire. The researcher examined whether on-going evidence of reliability and validity for the Heterosexist Attitudes in Sport—Lesbian scale (Mullin, 2013) questionnaire would be demonstrated. Female collegiate athletes (N = 239) from the mid-Atlantic region completed the HAS-L as well as a battery of questionnaires. Approximately 4 weeks later, participants completed the HAS-L again. The HAS-L was significantly related with the Attitudes toward Lesbians subscale of the Attitudes toward Lesbians and Gay Males—Short scale (Herek & McLemore, 2011) and other identified correlates of heterosexism, suggesting evidence of some criterion-related and convergent validity. Internal consistency and test-retest estimates ranged from .50–85 and .64–.91, respectively. The findings demonstrate good evidence of reliability and validity for the Cognitive/Affective subscale of the HAS-L. More research is necessary to better examine the behavioral subscales. Future researchers should consider making revisions of the Avoidance of the Lesbian Label subscale to achieve acceptable levels of reliability and validity.