南方农业学报2012，Vol.43Issue(9)：1395-1399,5.DOI:10.3969/j:issn.2095-1191.2012.09.1395
抗大片吸虫分泌排泄抗原单克隆抗体制备及其生物学特性鉴定
Preparation of monoclonal antibodies against excretory-secretory antigen of Fasciola gigantica and identification of its biological characteristics
摘要
Abstract
[Objective]Fasciola gigantica excretory and secretory antigen monoclonal antibodies were prepared for further study on diagnosis and control of Fasciola gigantica diseases. [ Method ]The monoclonal antibodies of Fasciola gigantica excretory and secretory antigen were made by hybridoma technology and its biological characteristics were identified by ELISA, thiocyanate elution. [ Result ] After indirect ELISA selection and three times of subcloning, five hybridoma ceD lines (6D3, 6B4, 7D2, 7D1, and 7D4) of monoclonal antibodies against excretory-secretory antigen of Fasciola gigantic were obtained. The numbers of chromosome of hybridoma cell lines, ranging from 90 to 110, were larger than that of the parent cells. The prepared five McAbs were found to be the isotopes of Ig G2b, Ig M,Ig M,Ig Gl and Ig M, respectively, with κ-light chain. The supernate antibody liters of 6D3, 6B4, 7D2, 7D1, and 7D4 were 1:400, 1:3200, 1:25600, 1:6400, and 1:6400, respectively; the ascites titers of five hybridoma cell lines were 1:104, 1:104, l:105, l:107, and 1:106, respectively. Epitope mapping indicated that 6D3 and 7D1, 6D3 and 7D2, 7D1 and 7D2, 7D4 and 7D1, and 7D4 and 7D2 acted on different epitopes; 6D3 and 6B4 probably acted on the same epitopes, or had steric hindrance on the same epitope; 7D4 and 6D3, 6B4 and 7D4, 6B4 and 7D1, and 6B4 and 7D2 might have steric hindrance. Relative affinity sequence of the five lines was 6B4>7D4>7D1 >6D3>7D2. All of the five cell lines could excrete monoclonal antibodies steadily. [ Conclusion ] It was concluded that the produced monoclonal antibodies against excretory-secretory antigen could be applied in the follow-up research of Fasciola gigantica diagnosis and immune mechanism.关键词
大片吸虫/分泌排泄抗原/单克隆抗体/生物学特性/鉴定Key words
Fasciola gigantica/ excretory-secretory antigen (ES Ag)/ monoclonal antibody/ biological characteristics / identification分类
农业科技引用本文复制引用
李晓栩,王华俊,林贤,陈汉忠,曹池,唐大运,孟盟,刘明如..抗大片吸虫分泌排泄抗原单克隆抗体制备及其生物学特性鉴定[J].南方农业学报,2012,43(9):1395-1399,5.基金项目
国家自然科学基金项目(30660140) （30660140）
广西教育厅资助项目(200620) （200620）
