A sensitive enzyme-linked immunosorbent assay for the quantitation of antigen-specific murine immunoglobulin E |
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| Authors: | P N Hill F T Liu |
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| Affiliation: | Department of Molecular Immunology, Scripps Clinic and Research Foundation, La Jolla, CA 92037, U.S.A. |
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| Abstract: | An enzyme-linked immunosorbent assay (ELISA) for the quantitation of 2,4-dinitrophenyl (DNP)-specific murine immunoglobulin (Ig) E is described. The assay uses beta-galactosidase, which is conjugated to goat anti-rabbit gamma-globulin (GARG) via a method using a mild heterobifunctional cross-linking reagent, m-maleimidobenzoyl-N-hydroxy-succinimide ester (MBS). The assay has a sensitivity for detection of about 200 pg/ml of anti-DNP IgE. Analyses of murine serum samples using this ELISA correlate well with those obtained using the passive cutaneous anaphylaxis (PCA) reaction in rats. With the use of automated 96-well reader, data acquisition is rapid and, therefore, this ELISA is ideal for analyses of large numbers of samples. The assay can be easily modified for the measurement of other Ig classes and of IgE of other antigen specificities. |
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| Keywords: | BSA bovine serum albumin DNP 2,4-dinitrophenyl DTNB dithiodinitrobenzoic acid ELISA enzyme-linked immunosorbent assay GARG goat anti-rabbit γ-globulin Ig immunoglobulin MBS 2-ME 2-mercaptoethanol NBS newborn bovine serum NMS normal mouse serum ONPG PBS phosphate-buffered saline pH 7.2 0.01 M sodium phosphate buffer 0.15 M NaCl PCA passive cultaneous anaphylaxis RAME rabbit anti-mouse IgE RIA radioimmunoassay |
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