Little is known concerning the natural history of beta-cell autoimmunity in infants and toddlers, especially in those without a first degree IDDM relative. A population-based cohort of Colorado infants at increased IDDM risk due to their HLA genotype has been identified through a PCR-based HLA screening of cord blood and is being prospectively studied. We report the distribution of insulin (IAA), GAD65 (GAA), and ICA512 autoantibody levels in 312 children aged 9 months and in 131 children aged 15 months from this cohort, without family history of IDDM. The levels of IAA, GAA and ICA512 did not differ by the HLA genotype (DR3/4,DQB1*0302 vs. DR3/3, vs. DR2/DR4,DQB1*0302 vs. DRx/4,DQB1*0302, where x is not DR3 or DR2), by ethnicity (non-Hispanic whites vs. other ethnic groups), or by age (9 vs. 15 months). The 95th and 99th percentiles of the IAA distribution were respectively 40 and 61 nU/ml at the age of 9 months and 38 and 59 nU/ml at the age of 15 months. The 95th and 99th percentiles of the GAA distribution were respectively 0.020 and 0.046 at the age of 9 months and 0.022 and 0.098 at the age of 15 months. We propose to use IAA levels greater than 60 nU/ml and GAA index greater than 0.05 to define the presence of beta-cell autoimmunity in children younger than 2 years.