Vitamin A deficiency is associated with increased risks of vertical transmission of HIV-1 (HIV) and of disease progression and mortality among HIV-infected adults. The study's objectives were to describe serum Vitamin A concentrations (Vitamin A levels) among HIV-infected children in a prospectively-followed North American cohort (the NICHD IVIG Clinical Trial), to examine changes in Vitamin A levels, and to investigate the relationships between Vitamin A levels and morbidity and mortality. Blood was collected from children at baseline and at three month intervals throughout the study. Serum samples were stored at -70°C at a central repository for 4 to 7 years until retrieved for Vitamin A assay. Samples were hexane-extracted and assayed by HPLC. The rate of change in Vitamin A levels, calculated by fitting a linear regression model, was expressed as μg/dl/year. Among 376 children enrolled in the trial, 207 had one or more serum samples available for testing; a total of 861 serum samples were analyzed. The median Vitamin A level at baseline was 31.0 μg/dl (range: < 10 μg/dl to 98 μg/dl). In a multivariate analysis, only race/ethnicity was marginally associated with baseline Vitamin A level. The rate of change in Vitamin A levels was examined for the 180 children with more than one serum sample available for analysis, and did not vary significantly by any factor other than baseline Vitamin A level. Baseline Vitamin A level was not associated with measures of morbidity, including incidence of infections, growth failure, CD4+ percent decline below 15%, increases in serum HIV RNA levels above either 105 or 106 copies/ml, or acute care hospitalization. Neither baseline Vitamin A level nor the rate of change of Vitamin A levels was associated with mortality. This study provides the first descriptive and analytic assessment of Vitamin A levels and changes in these levels in a prospectively-followed cohort of HIV-infected children in North America. Vitamin A was not associated with morbidity or mortality among these children with relatively normal Vitamin A levels.