To investigate whether insulin antibody (IAB) formation is associated with the appearance of islet cell cytoplasmic antibodies (ICA), islet cell surface antibodies (ICSA) and insulitis Balb/c Bln mice were immunized with porcine insulin in combination with or without Freund's adjuvant. The animals received 8 i.p. injections and were followed up to 150 days for the development of antibodies and insulitis. Mice immunized with insulin in CFA developed IAB as well as ICSA. Mice only receiving Freund's adjuvant emulsified in saline also developed ICSA. ICA were not detectable. Inflammatory infiltrates were found in the exocrine pancreatic parenchyma but not in islets. The results show that nonspecific stimulation of the immune system and the application of insulin as antigen leads to both the formation of ICSA and IAB, while insulitis was not detectable.