Ecological Studies on Food-Bacteria from Food and Environments

食品和环境中食品细菌的生态学研究

基本信息

批准号：
09460146
负责人：
UEMURA Takashi
金额：
$ 5.57万
依托单位：
Coolage of Agriculture, Osaka Prefecture University
依托单位国家：
日本
项目类别：
Grant-in-Aid for Scientific Research (B)
财政年份：
1997
资助国家：
日本
起止时间：
1997 至 1998
项目状态：
已结题

项目摘要

(1) To develop a rapid and specific method to detect and/or identify enterohemorrhagic Escberichia coli O 157 : H7, two mouse monoclonal antibodies (MCA) were prepared. Specificities of these two MAbs (1D9 and 3E8) were determined by flow cytometry method (FCM). 3E8 and 1D9 were found to react with E.coli O157 : H7, Citrobacter freundii and Salmonella group N (O : 30), but not with E bermannii With a mixture containing strains of E coil O157 : H7 and E coli O6 : Hl, two different peaks appeared in FCM with MCA, whereas a single peak appeared with polyclonal rabbit antiserum. From these findings, FCM with MCA is suggested to be a rapid, specific, and useful method to detect and identify strain(s) of E celi O157 : H7 in food ingredients (Ref.#1).(2) FCM with fluorescent-labeled anti-CPE antibody was applied to develop a rapid, specific, and convenient method to detect enterotoxin (CPE) exposed on the surface of spores of Clostridium perfringens. The results obtained indicate that FCM can specifically detect CPE exposed on C perfringens spores for a short time. Thus, FCM is found to be a rapid, specific, and convenient assay method for detection of CPE exposed on C perfringens spores, suggesting that it will be hopefully useful to diagnose food poisoning caused by C perfringens (Ref.#2).(3) Sandwich enzyme-linked immunosorbent assay (ELISA) was developed for the quantitative estimation of C perfringens enterotoxin (CPE) using* monoclonal* and* polyclonal* antibodies* as* capturing* and* detecting* antibodies* respectively.* The dose- dependent relationship between absorbance at 405 nm and concentration of purified CPE was obtained over the range of 0.5 ng/ml-8.0 ? g/ml. The sandwich ELISA is found to detect crude CPE in culture and CPE in 10% fecal extracts. This method is a convenient, rapid and sensitive for specific detection of GPE(Ref#3).

(1)为了开发一种快速、特异的检测和/或鉴定肠出血性大肠杆菌O 157 : H7的方法，制备了两种小鼠单克隆抗体(MCA)。这两种 MAb（1D9 和 3E8）的特异性通过流式细胞术（FCM）测定。发现3E8和1D9与大肠杆菌O157:H7、弗氏柠檬酸杆菌和N组沙门氏菌(O:30)反应，但不与含有大肠杆菌O157:H7和大肠杆菌O6:Hl菌株的混合物反应。在具有 MCA 的 FCM 中出现两个不同的峰，而在多克隆兔抗血清中出现单个峰。根据这些发现，FCM 与 MCA 被认为是一种快速、特异且有用的方法，用于检测和鉴定食品成分中的大肠杆菌 O157 : H7 菌株（参考文献#1）。(2) FCM 与荧光-应用标记的抗 CPE 抗体开发了一种快速、特异、便捷的方法来检测产气荚膜梭菌孢子表面暴露的肠毒素 (CPE)。所得结果表明FCM可以特异性检测短时间暴露在产气荚膜梭菌孢子上的CPE。因此，FCM 被发现是一种快速、特异且方便的检测产气荚膜梭菌孢子上暴露的 CPE 的检测方法，这表明它有望有助于诊断产气荚膜梭菌引起的食物中毒（参考文献#2）。(3 ) 开发了夹心酶联免疫吸附测定 (ELISA)，用于使用*单克隆*和*多克隆*抗体*作为*捕获*和*检测*来定量估计产气荚膜梭菌肠毒素 (CPE) * 405 nm 处的吸光度与纯化 CPE 浓度之间的剂量依赖性关系在 0.5 ng/ml-8.0 范围内获得。克/毫升。夹心 ELISA 可检测培养物中的粗 CPE 和 10% 粪便提取物中的 CPE。该方法方便、快速、灵敏，可特异性检测GPE(Ref#3)。