Immunoperoxidase Demonstration of Human Serum Globulin Binding to Islet Tissue Robert L. Sorenson, Ph.D., Richard D. Shank, M.D., and Robert P. Elde, Ph.D., Minneapolis SUMMARY Sera from juvenile diabetic and nondiabetic controls were tested, using the immunoperoxidase method, for the presence of islet- binding immunoglobulins. All diabetic sera tested contained an islet-binding protein and on the average the sera were more posi- tive than age-matched controls. Normal adult sera are undifferen- tiated from juvenile diabetic sera. Most sera from children less than two years of age did not bind to islet tissue and sera from cystic fibrosis patients had a markedly diminished ability to bind to islet tissue. The binding protein appears to be an immunoglobulin which selectively reacts with elements of the islet beta cell. DIABETES 24:230-37, February, 1975. Histopathologic evidence of islet inflammatory le- sions in juvenile-onset diabetics of short duration has been demonstrated in 68 per cent of the cases investigated. 12 This "insulitis" 3 or round cell infiltra- tion has also been experimentally induced by immuniz- ing several species of animals with insulin. 4 " 7 Qualita- tively the insulitis-type lesion is undifferentiated from those found in autoimmune disorders. In addition, the leucocyte migratory inhibition test has been used to demonstrate a "specifically altered antipancreatic hypersensitivity in 30 per cent of diabetics." 8 Such observations have led to the suggestion that autoim- munity has a role in the pathogenesis of diabetes. A significant association of juvenile diabetes with other autoimmune diseases (thyroiditis, hypothyroidism and Addisonian pernicious anemia) has also been noted. 9 " 16 Attempts, however, to demonstrate 'a circulating antibody to islet tissue have been largely unsuccessful. 1718 Nonetheless, there have been indica- From the Department of Anatomy, University of Minnesota Medical School, Minneapolis, Minnesota 55455. Accepted for publication November 11, 1974. 230 tions that a circulating antibody or binding protein may be present. These include complement fixing antibodies 19 as well as the binding of gamma globulins to human beta cells in three out of five untreated diabetics. 20 Also an antibody-like activity as measured by complement consumption has been demonstrated in 58 per cent of a diabetic group and 26 per cent of a normal control group. 21 A question thus continues to exist as to the existence and nature of a circulating protein capable of binding to elements of islet beta cells. During initial phases of a study intended to search for an autoimmune compo- nent in the sera of juvenile-onset diabetics at the time of diagnosis, a generalized phenomenon of apparent serum globulin binding to islet tissue was noted. These studies are an attempt to characterize this phenome- non. METHODS Sera from seventeen newly diagnosed juvenile dia- betics (X age = 8.9 ± 4.7) were obtained prior to the initial administration of insulin. Control sera were obtained from nineteen apparently normal children aged four months to two years (X age = 1. 1 ± 0.8) and age matched control sera were obtained from thirty juveniles hospitalized for unrelated conditions (X age = 8.4 ± 5). In addition, sera from twelve patients with cystic fibrosis were obtained (X age = 12.8 ± 5) as well as thirteen normal adult specimens (X age > 21). All sera were stored in 200 /il aliquots at -20° C. until used. Routine screening of sera for binding capacity was carried out at a dilution of 1 to 3- The diluent was 0.013 M borate buffered saline pH 8.2, 0.022 M EDTA and 0.5 per cent bovine serum albumin. As a system control and for study of ligand-binding protein interaction, sera from a diabetic (J.D. 13) and a normal DIABETES, VOL. 2 4 , NO. 2 Downloaded from http://diabetesjournals.org/diabetes/article-pdf/24/2/230/348058/24-2-230.pdf by guest on 16 November 2023