We evaluated a insulinoma-associated protein (IA-2) antibody assay kit using 125I-labelled recombinant IA-2. IA-2 antibodies were present in patients with early-onset type 1 diabetes mellitus (DM) at frequencies of 74%, 67%, 57%, and 50% for respective periods <1 year, 1 < or =years<2, 2< or =years<3, and 3< or =years<4 after onset. IA-2 antibody frequency was low throughout the DM course as compared with glutamic acid decarboxylase (GAD) antibody frequency. No one had IA-2 antibody, but 29% still had positive GAD antibody titers after 11 years. Of the patients with 0<years<7 duration, 42% had IA-2 Ab+/GAD Ab+, 9% IA-2 Ab+/GAD Ab-, and 24% IA-2 Ab-/GAD Ab+. Prevalence of IA-2 and GAD antibody in 1243 patients with type 2 DM were 1.5% and 3.1%, respectively, and 1.1% had both. This new IA-2 antibody kit is easy to use and provides a specific, sensitive method for making routine assays. Furthermore, the combined analysis of GAD antibody provides high detection of type 1 DM.