Development of the polymeric antibody specific to Salmonella as antibiotic substitution

肠炎沙门氏菌（Salmonella enteritidis， SE）是一种致病细菌，通过鸡蛋和肉经人肠道粘膜感染，引起食物中毒。我们重视利用对肠道消化酶具有耐受性的聚合抗体（IgA, IgM）作为不依赖抗生素的防御沙门氏菌感染的方法之一。本研究的目的是通过细胞融合的方法筛选SE特异性的聚合抗体，并明确其特性。采用三种不同免疫方法的小鼠脾细胞与骨髓瘤细胞融合形成杂杂瘤，通过连续克隆获得5种沙门氏菌抗原特异性单克隆抗体。采用鼻和腹膜联合免疫野生SE菌株，鼻免疫野生SE菌株获得抗SE IgG单克隆抗体2Ga80k，鼻免疫野生SE菌株获得抗SE IgA单克隆抗体NRA1、NRA2、NRA3，临床SE菌株口服感染获得抗SE igm单克隆抗体ORM1。2Ga80k为IgG2a型抗体，具有κ链，以约80 kDa的抗原和LPS识别表面糖链，并显示出阻滞50-86%粘附到Caco-2细胞的效果。ORM1为IgM型抗体，可识别42 kDa的糖链抗原，也可被沙门氏菌鞭毛抗体识别。后者表明ORM1能够识别沙门氏菌鞭毛。NRA1、NRA2和NRA3为具有α型H链结构的dIgA抗体，可识别50-80 kDa糖链抗原和LPS。观察到荧光素、异硫氰酸酯标记的ORM1、NRA1、NRA2和NRA3在SE细胞表面连接，阻断沙门氏菌对Caco-2细胞的粘附。结果表明，聚合抗SE- iga和抗SE- igm抗体对SE具有较好的粘附功能。腹膜注射杂交瘤后腹水相对产生大量单克隆抗体，产生相应种类的单克隆抗体。然而，该抗体的数量不足以实际应用。这一点将留作将来的大课题。少

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 … More 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