Islet cell autoantigen 69 kDa (ICA69) has been reported as a polypeptide antigen expressed in pancreatic beta cells, and autoimmunity against this antigen has been associated with insulin-dependent diabetes mellitus. We have studied the cell type specificity and ontogeny of ICA69 gene expression in man. The ICA69 gene was expressed in all adult human tissues. The level of expression was three-to five-times higher in the pancreas than in the brain, liver, intestine, kidney, spleen, lung or adrenal glands. Pancreatic ICA69 expression increased with age, adult levels being five times higher than the levels present at 13 weeks of gestation. Total RNA from four separate preparations of isolated human islets revealed levels of ICA69 mRNA similar to those found in the pancreas as a whole, although another islet antigen, glutamic acid decarboxylase 65, was highly enriched in the islets. In situ hybridization and immunohistochemical staining of sections of the fetal and adult pancreas revealed expression of the ICA69 gene and protein throughout the acinar, ductal, and islet tissue, but not in the mesenchyme. Analysis of ICA69 mRNA levels in human cell lines indicated expression in neural, endothelial and epithelial cells, but not in fibroblasts. In conclusion, ICA69, although highest in the pancreas, is widely distributed in other human tissues, excluding connective tissue. Within the human pancreas, ICA69 is not enriched in the islets or in the beta cells.