Expression of recombinant glutamic acid decarboxylase in insect larvae and its application in an immunoassay for the diagnosis of autoimmune diabetes mellitus

摘要

S]GAD65 was assessed qualitatively by incubating GADA-positive patients' sera in the presence of 1?μM of the recombinant enzyme. All sera tested became virtually negative after incubation with antigen excess. Besides, radiometric quantitative competition assays with GADA-positive patients' sera were performed by adding recombinant GAD65 (0.62 nM-1.4?μM). All dose response curves showed immunochemical identity between proteins. In addition, a bridge-ELISA for the detection of GADA was developed using S. frugiperda-GAD65. This assay proved to have 77.3%?sensitivity and 98.2% of specificity. GAD65 could be expressed in insect larvae, being S. frugiperda the best choice due to its high yield and purity. The development of a cost effective immunoassay for the detection of GADA was also afforded.