Novel enzyme immunoassay of anti-insulin igG in human serum

✍ Scribed by Takeyuki Kohno; Eiji Ishikawa; Satoru Sugiyama; Syuji Nakamura

• Publisher: John Wiley and Sons
• Year: 1988
• Tongue: English
• Weight: 564 KB
• Volume: 2
• Category: Article
• ISSN: 0887-8013
• DOI: 10.1002/jcla.1860020106

✦ Synopsis

A novel enzyme immunoassay of anti-insulin IgG in human serum is described. A serum sample containing anti-insulin IgG was treated with dextran-charcoal at pH 6.0 to remove endogenous insulin and subsequently incubated with dinitrophenyl biotinyl nonspecific rabbit IgG-insulin conjugate. The reaction mixture was further incubated with a rabbit (antidinitrophenyl bovine serum albumin) IgG-coated polystyrene ball to trap the complex formed between anti-insulin IgG and the conjugate. After washing to eliminate nonspecific IgG in the test serum, the polystyrene ball was incubated with dinitrophenyl-L-lysine to elute the complex. The eluate was incubated with an avidin-coated polystyrene ball. Finally, the amount of human anti-insulin IgG in the complex trapped onto the avidin-coated polystyrene ball was measured by incubation with rabbit (antihuman IgG (?-chain)) Fab'-peroxidase conjugate. This enzyme immunoassay was 1,000-fold more sensitive than the conventional enzyme immunoassay, in which an insulinbovine serum albumin-coated polystyrene ball was incubated with a serum sample containing anti-insulin IgG and subsequently with rabbit (antihuman IgG (ychain)) Fab'-peroxidase conjugate. The principle of the novel enzyme immunoassay can be used to more sensitively measure antibodies for most kinds of haptens and antigens than the conventional enzyme immunoassay.