A monoclonal antibody based enzyme-linked immunosorbent assay for the determination of GAD65, the smaller isoform of glutamic acid decarboxylase.
An enzyme-linked immunosorbent assay for GAD65, the smaller form of glutamic acid decarboxylase and an important autoantigen related to Type 1 diabetes, is described. The competitive binding assay is based on a monoclonal antibody specifically reactive with GAD65. The assay is suitable for quantification of this enzyme between 40 and 300 pg/microliter. The intraassay coefficients of variation (cv) are between 5.6% and 8.9% and the interassay cvs lie between 9.4% and 17.3%. The covalent binding of the antigen to magnetic beads as the solid phase makes the assay also applicable for quantification of GAD65 in tissue homogenates with a high concentration of detergent. The GAD65 content of islets isolated from newborn Lewis rat was detected to be 310 pg/islet. However, GAD65 was not detectable in mouse islets.
Hamann
J
J
Department of Immunochemistry, Institute of Diabetes Gerhardt Katsch, Karlsburg, Germany.
Hahmann
J
J
Strebelow
M
M
Ziegler
B
B
Ziegler
M
M
D000818:Animals; D000831:Animals, Newborn; D000911:Antibodies, Monoclonal; D001324:Autoantigens / Q000032:analysis; D001921:Brain / Q000201:enzymology*; D003922:Diabetes Mellitus, Type 1 / Q000201:enzymology / Q000276:immunology; D004797:Enzyme-Linked Immunosorbent Assay / Q000379:methods; D005260:Female; D005968:Glutamate Decarboxylase / Q000032:analysis*; D006801:Humans; D007515:Islets of Langerhans / Q000201:enzymology*; D007527:Isoenzymes / Q000032:analysis*; D051379:Mice; D008807:Mice, Inbred BALB C / Q000276:immunology; D008970:Molecular Weight; D051381:Rats; D011917:Rats, Inbred Lew; D015203:Reproducibility of Results; D012680:Sensitivity and Specificity; D013045:Species Specificity
D000911:Antibodies, Monoclonal; D001324:Autoantigens; D007527:Isoenzymes; D005968:Glutamate Decarboxylase