圆形碘孢虫完整孢子ELISA的建立  

Establishment of an ELISA against mature intact Myxobolus rotundus spores

作　　者：章晋勇[1] 汪建国[1] 吴英松[2] 李明 

机构地区：[1]中国科学院水生生物研究所 [2]第一军医大学热带医学实验室

出　　处：《中国水产科学》2005年第2期133-137,共5页Journal of Fishery Sciences of China

基　　金：国家自然科学基金资助项目(30271028);中国科学院知识创新工程课题(KSCX2-1-04).

摘　　要：探索建立鱼类寄生圆形碘孢虫(MyxobolusrotundusNemezek,1911,Myxosporea,Bivalvulida)完整孢子酶联免疫吸附试验(Enzyme linkedimmunosorbentassay,ELISA)检测的可行性,对丙酮、戊二醛、甲醛、多聚甲醛、乙醇等几种常用固定剂应用于此方法的效果进行评价,初步建立了针对鱼类黏孢子虫的完整孢子ELISA检测模式。结果显示,同比其他几种固定剂,2%的多聚甲醛、0 25%的戊二醛对圆形碘孢虫完整孢子ELISA有较好的效果,经二者固定后,最低可检出10个成熟的完整孢子,这为筛选鱼类黏孢子虫孢子表面抗原分子的特异性配体,如单克隆抗体及基因工程抗体或抗体片断,分析其表面分子性质奠定了基础。同时,讨论了该方法在鱼类黏孢子虫病原检测、属种鉴定、系统发育及黏孢子虫基础生物学研究等方面的潜在应用价值。The importance of surface molecules of intact mature myxosporeans spores has been increasingly recognized in recent years.For further understanding the characterization of them,selecting the specific ligands,e.g.monoclonal antibody or antibody fragments against them and mapping candidate antigens of diagnostic and protective interest and exploring the significance of them in the relationship of parasite-host,it is need to establish a rapid and simple practical method for detecting and selecting mature intact spores.In present work,we explored the feasibility of ELISA(Enzyme-linked immunosorbent assay) against intact mature Myxobolus rotundus spores and evaluated the effect of several readily fixed reagents,including acetone,formaldehyde,ethanol,glutaraldehyde and paraformaldehyde,on the ELISA,derivated from the strategy of characterizing target molecular(receptor) of cell surface of tumours in medicine and the normal fixing methods in immunohistochemistry.The results showed that 0.25% glutaraldehyde and 2% paraformaldehyde had prefered effect than unfixed control and the other fixed reagents under investigation in present work and represented almost equal sensitivity which they could detect as low as 10 intact mature spores.The steady and reproductive of the way was demonstrated by the statistic analysis.So a simple,rapid and sensitive ELISA method were preliminarily established for detecting mature intact M.rotundus spores and the specific ligands of the surface molecules of them were selected.The tool would be able to apply to all fish parasitic myxosporean species with mature spore stage.The potential implication of the method in diagnosis of pathogen,assisting determination of specific species,phylogeny,screening the specific antigen molecules of diagnosis and protective interest,and other basic biology of fish myxosporea were also discussed.

关 键 词：完整孢子 ELISA 圆形碘孢虫 黏孢子虫 

分 类 号：S941.5[农业科学—水产养殖]