| Co-Investigator(Kenkyū-buntansha) |
KONISHI Yoshiko National Institute of Health Sciences, Division of Microbiology, Group Reader, 衛生微生物部, 室長 (10195761)
AMETANI Michiko The Institute of Physical and Chemical Research, Research Center for Allergy and Immunology, Researcher, 免疫アレルギー研究センター, 研究員 (80240688)
AZUMA Norihiro Utsunomiya University, Faculty of Agriculture, Professor, 農学部, 教授 (30151062)
天野 冨美夫 国立感染症研究所, 細胞化学部, 主任研究員 (90142132)
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| Budget Amount *help |
¥14,300,000 (Direct Cost: ¥14,300,000)
Fiscal Year 2003: ¥1,900,000 (Direct Cost: ¥1,900,000)
Fiscal Year 2002: ¥2,400,000 (Direct Cost: ¥2,400,000)
Fiscal Year 2001: ¥2,400,000 (Direct Cost: ¥2,400,000)
Fiscal Year 2000: ¥7,600,000 (Direct Cost: ¥7,600,000)
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| Research Abstract |
Salmonella enteritidis (SE) is a pathogenic germ, which infect via man's intestinal tract mucosa with chicken egg and meat, and causes food poisoning. We paid our attention to using the polymeric antibody (IgA, IgM) which shows tolerance to digestive enzyme in intestine as one of the methods of defending Salmonella infection without being dependent on antibiotics. The purpose of this research is selecting polymeric antibody specific to SE by cell fusion method, and clarifying the characteristics.
Five kinds of monoclonal antibodies specific to Salmonella antigen were produced by successive cloning from hybridoma made through cell fusion between spleen cells of the mouse, which were immunized by three different methods, with a myeloma cell. Anti-SE IgG monoclonal antibody, 2Ga80k, was produced by combined use of nasal and peritoneal immunization of wild strain of SE, three kinds of anti-SE IgA monoclonal antibodies, NRA1, NRA2, and NRA3, by nasal immunization of wild strain of SE, and an
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ti-SE-IgM monoclonal antibody, ORM1, by the oral infecting method of clinical strain of SE. 2Ga80k was IgG2a type of antibody, which has κ chain, recognized the surface sugar chain with about 80 kDa of antigen and LPS, and showed the effect, which obstructed about 50-86% adhesion into a Caco-2 cell. ORM1 was IgM type of antibody, recognized sugar chain antigen of 42 kDa, which was also recognized by antibody to flagellum of Salmonella. The latter suggest that ORM1 can recognize flagellum of salmonella. NRA1, NRA2, and NRA3 showed to be the dIgA antibody having α type H chain structurally, and recognized sugar chain antigen of 50-80 kDa and LPS. It was observed that ORM1, NRA1, NRA2 and NRA3, which were labeled with fluorescein, isothiocyanate was connected in cell surface of SE, and that they obstructed adhesion of a salmonella to the Caco-2 cell. It was shown the polymeric anti-SE-IgA and anti-SE-IgM antibodies have superior adhesion function for SE. Large quantities of monoclonal antibodies were comparatively produced from ascites after peritoneal injection of the hybridoma, which produced the respective kind of monoclonal antibody. However, the amount of this antibody was not enough to use practically. This point will be left as a big subject of the future. Less
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