Type 1 diabetes (T1D) is an autoimmune disease that occurs in genetically susceptible individuals. Regulatory T cells (T_regs) have been shown to be defective in the autoimmune disease setting. Thus, efforts to repair or replace T_regs in T1D may reverse autoimmunity and protect the remaining insulin-producing β cells. On the basis of this premise, a robust technique has been developed to isolate and expand T_regs from patients with T1D. The expanded T_regs retained their T cell receptor diversity and demonstrated enhanced functional activity. We report on a phase 1 trial to assess safety of T_reg adoptive immunotherapy in T1D. Fourteen adult subjects with T1D, in four dosing cohorts, received ex vivo–expanded autologous CD4⁺CD127^lo/−CD25⁺ polyclonal T_regs (0.05 × 10⁸ to 26 × 10⁸ cells). A subset of the adoptively transferred T_regs was long-lived, with up to 25% of the peak level remaining in the circulation at 1 year after transfer. Immune studies showed transient increases in T_regs in recipients and retained a broad T_reg FOXP3⁺CD4⁺CD25^hiCD127^lo phenotype long-term. There were no infusion reactions or cell therapy–related high-grade adverse events. C-peptide levels persisted out to 2+ years after transfer in several individuals. These results support the development of a phase 2 trial to test efficacy of the T_reg therapy.