This study aimed to understand gender and ethnicity differences in HIV-related stigma experienced by 1026 HIV-positive individuals living in Ontario, Canada that were enrolled in the OHTN Cohort Study. Total and subscale HIV-related stigma scores were measured using the revised HIV-related Stigma Scale. Correlates of total stigma scores were assessed in univariate and multivariate linear regression. Women had significantly higher total and subscale stigma scores than men (total, median = 56.0 vs. 48.0, p<0.0001). Among men and women, Black individuals had the highest, Aboriginal and Asian/Latin-American/Unspecified people intermediate, and White individuals the lowest total stigma scores. The gender-ethnicity interaction term was significant in multivariate analysis: Black women and Asian/Latin-American/Unspecified men reported the highest HIV-related stigma scores. Gender and ethnicity differences in HIV-related stigma were identified in our cohort. Findings suggest differing approaches may be required to address HIV-related stigma based on gender and ethnicity; and such strategies should challenge racist and sexist stereotypes.