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The pubertal transition has been identified as a time of risk for development of type 2 diabetes, particularly among vulnerable groups, such as African Americans (AAs). Documented ethnic differences in insulin secretory dynamics may predispose overweight AA adolescents to risk for type 2 diabetes. The objectives of this longitudinal study were to quantify insulin secretion and clearance in a cohort of 90 AA and European American (EA) children over the pubertal transition and to explore the association of genetic factors and adiposity with repeated measures of insulin secretion and clearance during this critical period. Insulin sensitivity was determined by intravenous glucose tolerance test (IVGTT) and minimal modeling; insulin secretion and clearance by C-peptide modeling; genetic ancestry by admixture analysis. Mixed-model longitudinal analysis indicated that African genetic admixture (AfADM) was independently and positively associated with first-phase insulin secretion within the entire group (P < 0.001), and among lean children (P < 0.01). When examined within pubertal stage, this relationship became significant at Tanner stage 3. Total body fat was a significant determinant of first-phase insulin secretion overall and among obese children (P < 0.001). Total body fat, but not AfADM, was associated with insulin clearance (P < 0.001). In conclusion, genetic factors, as reflected in AfADM, may explain greater first-phase insulin secretion among peripubertal AA vs. EA; however, the influence of genetic factors is superseded by adiposity. The pubertal transition may affect the development of the β-cell response to glucose in a manner that differs with ethnic/genetic background.