Human islet cell autoantibodies specifically bind cloned rat islet cells

Sera from newly diagnosed type I (insulin dependent) diabetic patients and control subjects were tested by indirect immunofluorescence on a particular subclone of RINm5F cells for the presence of cytoplasmic antibodies. Cell cultures after biochemical fixation and frozen sections of cell sheets obtained after incubating confluent monolayers with dispase were used alternatively as substrate. Eight out of ten ICA positive sera previously tested on frozen sections of human pancreas bind RIN cells when fixed with acetone-ethanol whereas no staining was obtained after treating the cells with paraformaldehyde or glutaraldehyde. Four normal sera did not bind RIN cells under these conditions. Alternatively, frozen sections of cell sheets appeared to be a more sensitive substrate as all ICA positive sera tested reacted with cell cytoplasm. In addition, an immunoradiometric assay was developed using microtitre plate wells coated with Nonidet P40 solubilized RIN antigens. Coated wells were incubated with ICA positive and ICA negative diabetic sera along with sera from normal individuals. Antigen antibody complexes were detected by the binding of 125I-Protein A. A good correlation was found between binding values and results obtained by indirect immunofluorescence on frozen sections of human pancreas. Although this study indicates that RIN cells express a specific cytoplasmic antigen to which ICA positive sera bind, more information is required concerning the exact nature of this antigen before such assay can be reliably used for the detection of ICA.