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OBJECTIVE--To investigate autoantigens in β-cells, we have used a panel of pathogenic T-cell clones that were derived from the NOD mouse. Our particular focus in this study was on the identification of the target antigen for the highly diabetogenic T-cell clone BDC-5.2.9. RESEARCH DESIGN AND METHODS--To purify β-cell antigens, we applied sequential size exclusion chromatography and reverse-phase high-performance liquid chromatography to membrane preparations of β-cell tumors. The presence of antigen was monitored by measuring the interferon-γ production of BDC-5.2.9 in response to chromatographic fractions in the presence of NOD antigen-presenting cells. Peak antigenic fractions were analyzed by ion-trap mass spectrometry, and candidate proteins were further investigated through peptide analysis and, where possible, testing of islet tissue from gene knockout mice. RESULTS--Mass-spectrometric analysis revealed the presence of islet amyloid polypeptide (IAPP) in antigen-containing fractions. Confirmation of IAPP as the antigen target was demonstrated by the inability of islets from IAPP-deficient mice to stimulate BDC-5.2.9 in vitro and in vivo and by the existence of an IAPP-derived peptide that strongly stimulates BCD-5.2.9. CONCLUSIONS--IAPP is the target antigen for the diabetogenic CD4 T-cell clone BDC-5.2.9. Diabetes 60:2325-2330, 2011, CD4 T cells are major immune players in the initiation and pathogenesis of type 1 diabetes, but the identification of the β-cell antigens that drive autoreactive T-cell responses has been [...]

A panel of [CD4.sup.+] T-cell clones has been isolated from the spleen and lymph nodes of diabetic NOD mice. These clones haven been shown to be islet-specific both in vivo [...]