双向电泳结合蛋白质印迹技术分析猪囊尾蚴乳酸脱氢酶  

Analysis of lactate dehydrogenase expression in Taenia solium cysticerci by two dimensional electrophoresis and Western-blotting

作　　者：方文[1] 包怀恩[2] 肖靓靓[2] 牟荣[2] 

机构地区：[1]贵阳医学院医学检验系,贵阳550004 [2]贵阳医学院寄生虫学教研室,贵阳550004

出　　处：《黑龙江畜牧兽医》2011年第8期8-10,共3页Heilongjiang Animal Science And veterinary Medicine

基　　金：国家自然科学基金项目(30660168)

摘　　要：为了研究猪囊尾蚴乳酸脱氢酶(LDH)的表达,试验选择四川省雅江县呷拉乡猪带绦虫病患者,给其口服槟榔-南瓜子驱虫,收集、制备虫卵悬液(8万个/mL),再给3头20日龄三元杂交乳猪猪灌胃虫卵悬液,每头1 mL,40 d后收集、制备猪囊尾蚴蛋白,进行双向电泳(2-DE)分析,将凝胶蛋白斑点转移至聚偏氟乙烯膜(PVDF膜),用自制的大鼠抗猪带绦虫LDH血清作为一抗、健康大鼠血清作为阴性对照进行蛋白质印迹(Western-blotting)分析。结果表明:双向电泳凝胶共检测到(207±9)个蛋白质斑点,相对分子质量(Mr)为14 400～94 000,等电点(pI)为3.0～10.0;试验组特异性抗原抗体阳性杂交斑点为1个,阴性对照未见阳性杂交斑点;将Western-blotting检测的抗原抗体阳性杂交斑点与原双向电泳凝胶斑点进行比对,找到对应蛋白斑点,经ImageMaster 2D Platinum 5.0软件分析后初步确定该蛋白斑点的pI/Mr为7.03/35 368,与猪带绦虫LDH的pI/Mr理论推导值接近,说明猪囊尾蚴表达LDH。To analyze the lactate dehydrogenase （LDH） expression of Taenia solium cysticerci, the experiment used the technology of two -dimensional eleetrophoresis and Western - blotting to study it. The total proteins of Taenia solium eystieerci acquired 40 days after infecting pigs were separated by two - dimensional electrophoresis, and Western - blotting were performed to find out LDH after the protein spots on the gel transferred to PVDF membrane. The results showed that there are （ 207 ±9 ） protein spots of Taenia solium cysticerci in 2 - DE gel stained by Coomassie brilliant blue ,their relative molecular mass （Mr） were 14 400 - 94 000 ,their isoeleetric points（pI） were 3.0 - 10.0. Western - blotting showed that there was 1 specific antigen spot with anti - LDH mixed sera of the experimental rats and non - specific spot with the mixed sera of the healthy rats. The spot in the Western - blotting corresponded to the protein spot in 2 - DE gel. The pI/Mr of the protein （7.03/35 368 ） in 2 - DE gel was similar to that of Taenia solium LDH deduced theoretical values. It indicates that the Taenia solium cysticerci express the lactate dehydrogenase.

关 键 词：猪带绦虫 囊尾蚴 乳酸脱氢酶 双向电泳 蛋白质印迹 

分 类 号：S852.734[农业科学—基础兽医学]