Biol. Pharm. Bull. 29(7) 1480—1482 (2006)

were found in human sera for the first time in 1993. The prevalence of biotin-binding immunoglobulin was higher in patients with atopic dermatitis, other dermatitis, allergic disorders, and autoimmune disease. Recently, biotin-binding IgG was detected in a multiwell microplate format for the first time. We developed a quantitative assay for biotinbinding IgG without any purification of samples using F(ab’)2anti-human IgG-coated multiwell microplates. 5) We showed that the ratio of biotin molecules to protein molecules (biotin–protein ratio) in biotinylated IgG in human sera affected the detection limit in this assay. The slope of doseresponse curve of the assay decreased as the biotin–protein ratio of standards decreased and the observed values of biotinylated IgG in human sera depended on the biotin–protein ratio of standards. Thus it was very important to measure the biotin–protein ratio of biotinylatd immunoglobulins which were used as standards in the assay. In this study, we investigated a synthesis method for biotinylated human immunoglobulins, how to determine the biotin–protein ratio of the biotinylated proteins, and their stability.