Glutamic acid decarboxylase autoantibody assay using I-125-labelled recombinant GAD(65) produced in yeast

We describe a new method for measuring autoantibodies (Ab) to the 65 kDa isoform of glutamic acid decarboxylase (GAD(65)). In particular, GAD(65) without the hydrophobic N-terminal region has been produced in yeast, purified, labelled with I-125 and reacted with GAD(65) Ab. Antibody bound I-125-GAD(65) is then precipitated by the addition of solid phase protein A. With the assay, GAD(65) Ab were detected in 59 of 71 (83%) islet cell antibody (ICA) positive IDDM patients and in 8 of 23 (35%) ICA negative IDDM patients (overall 67 of 94 (71%) of IDDM patients). Low concentrations of GAD(65) Ab were also detected in 2/98 (2%) healthy blood donors and 1/27 (4%) Graves' disease patients had a high level of antibody. GAD(65) Ab were not detected in any of 10 Hashimoto's thyroiditis, 20 Addison's disease or 19 myasthenia gravis sera. There was good agreement between the I-125 assay and the current reference method based on S-35-labelled full-length GAD(65) (produced by in vitro transcription/translation reaction) and solid phase protein A (r = 0.91, n = 108). Overall, our I-125 assay showed sensitivity, precision and disease group specificity at least as good as any assay so far described. These features, combined with a simple assay protocol and the convenience of I-125 counting and handling indicate that the method is suitable for routine GAD(65) Ab measurements. Copyright (C) 1996 Elsevier Science B.V.