Objectives: To longitudinally follow a cohort of adolescents with human immunodeficiency virus (HIV) and to investigate long-term antiretroviral therapy adherence and factors associated with adherence. Design, Setting, and Patients: Adolescents infected with HIV (N=231; mean age, 18.4 years; 72.7% female; 74.9% African American) from 13 cities throughout the United States were assessed at 3-month intervals. Main Outcome Measures: Self-reported adherence measures were validated by comparison with HIV-1 RNA viral load, and behavioral factors that may be associated with antiretroviral therapy adherence were assessed. Results: At the initial visit, approximately 69% of the adolescents reported being adherent to antiretroviral therapy. Adolescents in the later HIV disease stage were less likely to be adherent compared with those in the earlier disease stage. Less alcohol use and being in school were associated with adherence by adolescents on weekends and over the preceding month. Longitudinal adherence was investigated among 65 subjects initially adherent with available information for at least 4 consecutive visits. The median time to nonadherence was 12 months, and failure to maintain adherence was significantly associated with younger age and depression. Among adolescents who attained an undetectable viral load, only about 50% maintained an undetectable viral load for the year. Conclusions: These findings indicate an urgent need for better interventions to assist adolescents with HIV in adhering to their medication regimens. Adolescents with advanced disease are likely to need more intervention. New treatments recently found effective for adolescent depression may assist in improving adherence for a majority of adolescents with HIV. ©2005 American Medical Association. All rights reserved.