HIV-related stigma inhibits optimal HIV prevention and treatment among African-Americans. Regional differences in HIV/AIDS prevalence may be related to stigma among young African-Americans. Baseline data (N = 1,606) from an HIV prevention intervention were used to investigate regional differences in HIV-related stigma and knowledge among African-American adolescents in four midsized cities in the Northeastern and Southeastern US. Analyses indicated greater HIV-related stigma among adolescents from the Southeast relative to adolescents from the Northeast (F = 22.23; p < 0.0001). Linear regression indicated a negative relationship between HIV stigma and HIV knowledge (b = −0.65; p < 0.0001). Addressing HIV/AIDS in high prevalence locales should include efforts to reduce HIV-related stigma.