Development of glutamic acid decarboxylase 65 (GAD65) autoantibody assay using biotin-GAD65 fusion protein.

Research output: Contribution to journalArticle

Abstract

We evaluated a biotin-glutamic acid decarboxylase 65 (GAD65)-based enzyme-linked immunosorbent assay (B-ELISA) to detect GAD65 autoantibodies (GAD65Ab) in 78 sera from individuals with newly diagnosed type 1 diabetes. The GAD65Ab index of patients with type 1 diabetes (mean value of GAD65Ab index of 1.891) was significantly higher than those in 50 sera from healthy control group (mean value of 0.068). The intra- and inter-assay coefficients of variation (CV) were calculated to be 1.042 and 10.703%, respectively. The specificity of the B-GAD65 ELISA was comparable to the standard radioimmunoassay (RIA) which is routinely used in the laboratory. We describe the optimal conditions of the binding kinetics from each assay-step for the detection of GAD65Ab using the WHO standard serum 97/550 as a model autoantibody serum. We concluded that incubation times of 15, 90, and 90 min for step 1 (pre-incubation of Biotin14-GAD65 with serum), step 2 (binding the Ab/Ag complex to NeutrAvidin plate)

Details

Authors
  • Dong Luo
  • Chad N Rogers
  • Jordan T Steed
  • Lisa K Gilliam
  • Christiane S Hampe
  • Åke Lernmark
External organisations
  • University of Washington, Seattle
Research areas and keywords

Subject classification (UKÄ) – MANDATORY

  • Endocrinology and Diabetes
Original languageEnglish
Pages (from-to)97-104
JournalJournal of Biotechnology
Volume111
Issue number1
Publication statusPublished - 2004
Publication categoryResearch
Peer-reviewedYes
Externally publishedYes