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Rat urinary proteins binding human IgE detected by chemiluminescence
Author(s) -
Bayard Cecilia,
Vesterberg Olof
Publication year - 1998
Publication title -
electrophoresis
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 0.666
H-Index - 158
eISSN - 1522-2683
pISSN - 0173-0835
DOI - 10.1002/elps.1150191020
Subject(s) - chemiluminescence , immunoglobulin e , blot , chemistry , albumin , gel electrophoresis , urine , chromatography , allergen , polyacrylamide gel electrophoresis , western blot , microbiology and biotechnology , biochemistry , antibody , biology , immunology , allergy , enzyme , gene
Work with laboratory animals involves the increased risk of developing an allergy. Certain proteins in male rat urine have been shown to be major allergens, i.e. , Rat n 1.01 and Rat n 1.02. Rat urinary proteins separated by sodium dodecyl sulfate‐polyacrylamide gel electrophoresis (SDS‐PAGE) were electroblotted to polyvinylidene difluoride (PVDF) membranes. Immunoblot analysis of human IgE binding components in rat urine utilizing chemiluminescence and a newly developed luminometer is described. Luminometer scanning curves of IgE reactivity to the separated rat urinary proteins are demonstrated. Rat allergic individuals showed different immune responses to the separated proteins. Rat n 1.02, also known as the major protein α 2u ‐globulin, was not always the dominant allergen. Another protein in the albumin region, 60–67 kDa, was found to be an important allergen to some rat‐sensitive subjects. Reactivity in the skin prick test with purified Rat n 1.01 and Rat n 1.02 fractions were strong. However, in dot blot under nondenaturing conditions, only weak responses were obtained to the purified rat urinary proteins except for the albumin fraction. Chemiluminescence measurements in blotting membranes of patient IgE bound to different dilutions of certain rat urine proteins revealed good quantitative relationships. Three different chemiluminescence substrates were tested. Measurement of IgE bound to individual allergens as well as the abundance and relative importance of various allergens were studied.