title: A reverse-sandwich ELISA for IgG antibody to a pollen allergen.
abstract: We have developed a reverse-type sandwich ELISA for measurement of IgG (+IgA) antibody to a major allergen of Sugi (Japanese cedar) pollens. In this assay, microplate wells were coated with the allergen proteins that had been dispersed in the presence of 0.5 mol/L NaCl and 20 micrograms/ml of bovine serum albumin, followed by addition of test serum, and then the biotinylated allergen. Beta-galactosidase-conjugated streptavidin was used for detection of the captured allergen, 4-methylumbelliferyl-beta-D-galactoside was used as the enzyme substrate, and fluorescence intensity of the product, 4-methylumbelliferone, was measured by a fluorometric reader. This assay was found to detect a total activity of IgG, IgA, and IgE antibodies present in the serum of the patients with pollinosis. IgG (+IgA) antibody was determined after absorption of the serum with anti-IgE Sepharose gel. Sera from individuals who were unexposed to the pollens demonstrated no nonspecific reactions in this assay. This method may be useful as a simple technique for monitoring the efficacy of immunotherapy that is expected to elicit IgG blocking antibody and also for large-scale seroepidemiologic studies on the prevalence of non-IgE antibodies in a general population.
authors.affiliation: Department of Microbiology, Institute of Public Health, Tokyo, Japan.
authors.forename: H
authors.initials: H
authors.lastname: Miyazawa
authors.forename: S
authors.initials: S
authors.lastname: Inouye
authors.forename: M
authors.initials: M
authors.lastname: Sakaguchi
authors.forename: K
authors.initials: K
authors.lastname: Koizumi
mesh_terms: D000485:Allergens / Q000276:immunology*; D000818:Animals; D000906:Antibodies / Q000032:analysis*; D002850:Chromatography, Gel; D004797:Enzyme-Linked Immunosorbent Assay / Q000379:methods*; D006801:Humans; D006967:Hypersensitivity / Q000276:immunology*; D007073:Immunoglobulin E / Q000032:analysis; D007074:Immunoglobulin G / Q000032:analysis*; D011058:Pollen; D011817:Rabbits
chemical_list: D000485:Allergens; D000906:Antibodies; D007074:Immunoglobulin G; D007073:Immunoglobulin E