DETECTION OF GAD(65) ANTIBODIES IN DIABETES AND OTHER AUTOIMMUNE-DISEASES USING A SIMPLE RADIOLIGAND ASSAY

Autoantibodies to glutamic acid decarboxylase (GAD) are frequent at or before the onset of insulin-dependent diabetes mellitus (IDDM). We ha ve developed a simple, reproducible, and quantitative immunoprecipitat ion radioligand assay using as antigen in vitro transcribed and transl ated [S-35] methionine-labeled human islet GAD(65). By using this assa y, 77% (77 of 100) of serum samples from recent-onset IDDM patients we re positive for GAD(65) antibodies compared with 4% (4 of 100) of seru m samples from healthy control subjects. In competition analysis with unlabeled purified recombinant human islet GAD(65), binding to tracer was inhibited in 74% (74 of 100) of the GAD(65)-positive IDDM serum sa mples compared with 2% of the control samples. The levels of GAD antib odies expressed as an index value relative to a standard serum, analyz ed with or without competition, were almost identical (r = 0.991). The intra- and interassay variations of a positive control serum sample w ere 2.9 and 7.6%, respectively (n = 4). The frequency of GAD antibodie s was significantly higher with IDDM onset before the age of 30 (80%, 59 of 74) than after the age of 30 (48%, 10 of 21) (P < 0.01). The pre valence of islet cell antibodies showed a similar pattern relative to age at onset. Because simultaneous occurrences of multiple autoimmune phenomena are common, we analyzed sera from patients with other autoim mune diseases. The frequency of GAD antibodies in sera positive for DN A autoantibodies (8% [2 of 25] and 4% [1 of 25] in competition analysi s) or rheuma factor autoantibodies [12% (4 of 35) and 3% (1 of 35) in competition analysis] was not different from that in control samples. In contrast, in sera positive for ribonucleoprotein antibodies the fre quency of GAD antibodies was significantly increased (73% [51 of 70] a nd 10% [7 of 70] in competition analysis [P < 0.025]). In conclusion, even large numbers of serum samples can now be tested for GAD(65) anti bodies in a relatively short time, allowing screening of individuals w ithout a family history of IDDM for the presence of this marker.