Evaluation of a high IgE-responder mouse model of allergy to bovine beta-lactoglobulin (BLG): development of sandwich immunoassays for total and allergen-specific IgE, IgG1 and IgG2a in BLG-sensitized mice

An animal model of food allergy represents an important tool for studying t he mechanisms of induction and repression of an allergic reaction, as well as for the development of an immunotherapy to prevent or minimize such an a dverse reaction. IgE and IgG1 (Th2 response) vs. IgG2a (Th1 response) are g ood markers for the induction of an allergic response in mice. Nevertheless , while the total serum concentrations of these isotypes are easy to measur e using classical sandwich immunoassays, this is not the case for allergen- specific isotypes. To develop an animal model of allergy to bovine beta-lac toglobulin (BLG), we set up quantitative assays for total and for allergen- specific IgE, IgG1 and IgG2a. Microtiter plates coated either with anti-iso type antibodies (Abs) or with allergen were used for Ab capture, while anti -isotype Fab' fragments coupled to acetylcholinesterase were used for visua lization. These assays of anti-BLG specific Abs are original in two ways, F irst, assay calibration is performed using anti-BLG specific mAbs, thus all owing good quantification of the different isotypes and subclasses of serum antibodies. Second, the detection of all anti-BLG specific Abs, i.e., thos e recognizing both the native and denatured forms of the protein, is achiev ed through indirect coating of BLG using biotin-streptavidin binding. The p resent assays are quantitative, specific to the isotype (cross-reactivity < 0.5%), very sensitive (detection limit in the 10 pg/ml range), and reprodu cible (coefficient of variation less than 10%). Applied to the humoral resp onse in mice sensitized with BLG adsorbed on alum, these assays proved to b e a very useful tool for monitoring high IgE-responder mice following BLG i mmunization, and for an immunotherapy directed at polarizing the immune res ponse. (C) 2000 Elsevier Science B.V. All rights reserved.