An Improved Preparation of Antibody-Coated Polystyrene Beads for Sandwich Enzyme Immunoassay |
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| Authors: | Eiji Ishikawa Yoshitaka Hamaguchi Masayoshi Imagawa Mitsuo Inada Hiroo Imura Nobuhiko Nakazawa |
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| Affiliation: | 1. Department of Biochemistry , Medical College of Miyazaki, Kiyotake , Miyazaki, 889-16, Japan;2. Second Division, Department of Medicine , Kyoto University School of Medicine , Kyoto, 606, Japan;3. Radioisotope Labs. Ltd. , 3-10-5, Nihonbashi, Chuo-ku, Tokyo, 103, Japan |
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| Abstract: | An improved preparation of antibody-coated polystyrene beads for sandwich enzyme immunoassay of human thyroid-stimulating hormone (TSH) was described. Rabbit anti-TSH IgG was purified by eluting at pH 2.5 from a TSH-Sepharose column, diluted 3 or 9 fold with normal rabbit IgG and used for coating polystyrene beads by physical adsorption. In a sandwich enzyme inmunoassay of TSH using rabbit (anti-TSH) Fab-β-D-galactosidase conjugate, β-D-galactosidase activities specifically bound to thus prepared polystyrene beads in the presence of TSH was 2.8–6.3 fold higher than those bound to polystyrene beads coated with anti-TSH IgG before purification. A similar effect was observed when guinea pig anti-pork insulin IgG, rabbit (anti-human IgE) IgG and goat (anti-human IgE) IgG were treated at pH 2.5. This improvement may be based on a conformational change of Fc in IgG molecule which was caused by the treatment at pH 2.5. Other sandwich immunoassays such as fluoro-and radio-immunoassays may also be improved in the same way. (KEY WORDS: Sandwich enzyme immunoassay, TSH, insulin) |
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| Keywords: | Stabilization Enzyme immunoassay Penicillinase |
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