Insulin is a beta-cell-specific and pivotal autoantigen in type 1 diabetes (1) and is produced after proteolysis of its prohormone proinsulin, which is a potential target of autoimmunity. T-cell studies in the nonobese diabetic (NOD) mouse (2, 3) and in prediabetic relatives of patients with type 1 diabetes (4, 5) indicate the existence of proinsulin-specific immunodominant epitopes localized in the region between the C-peptide and the A--chain, whereas immune intervention with proinsulin in the NOD mouse can prevent diabetes onset. These studies have led to the proposal of proinsulin as a primary target of autoimmunity associated with type 1 diabetes. At the humoral level, proinsulin-specific autoantibodies have also been reported in humans (6, 7). Our own data show, however, that proinsulin autoantibody (PAA) and insulin autoantibody (IAA) levels correlate well in newly diagnosed patients, but IAAs are more sensitive and specific for type 1 diabetes (7). Furthermore, IAA or PAA could be completely displaced with either insulin or proinsulin in the majority of patients, suggesting that the main epitopes recognized by these antibodies at diabetes onset are on the insulin molecule. In the present study, we investigated whether antibodies to proinsulin-specific epitopes either preceded or were a more relevant finding early in the development of diabetes-associated autoimmunity.