Regulatory T-cells (Tregs) play a critical role in maintaining dominant peripheral tolerance. Previous characterizations of Tregs in type 1 diabetes have used antibodies against CD4 and α-chain of the interleukin-2 receptor complex (CD25). This report extends those investigations by the addition of a more lineage-specific marker for Tregs, transcription factor forkhead box P3 (FOXP3), in subjects with type 1 diabetes, their first-degree relatives, and healthy control subjects. With inclusion of this marker, two predominant populations of CD4⁺CD25⁺ T-cells were identified: CD4⁺CD25⁺FOXP3⁺ as well as CD4⁺FOXP3⁻ T-cells expressing low levels of CD25 (CD4⁺CD25^LOWFOXP3⁻). In all study groups, the frequency of CD4⁺CD25⁺FOXP3⁺ cells was age independent, whereas CD4⁺CD25^LOWFOXP3⁻ cell frequencies strongly associated with age. In terms of additional markers for delineating cells of Treg lineage, FOXP3⁺ cells were CD127⁻ to CD127^LOW whereas CD25⁺ cells were less restricted in their expression of this marker, with CD127 expressed across a continuum of levels. Importantly, no differences were observed in the frequency of CD4⁺CD25⁺FOXP3⁺ T-cells in individuals with or at varying degrees of risk for type 1 diabetes. These investigations suggest that altered peripheral blood frequencies of Tregs, as defined by the expression of FOXP3, are not specifically associated with type 1 diabetes and continue to highlight age as an important variable in analysis of immune regulation.