猪戊型肝炎ELISA诊断试剂盒的研制及应用  

Preparation and Application of Swine Hepatitis E Virus Diagnostic ELISA Kit

作　　者：刘启文[1] 李震[2,3] 赵凯[4,3] 于瑞嵩[2,3] 唐雪明[4,3] 朱宏[4,3] 吴潇[4,3] 谭芙蓉[4,3] 王金斌[4,3] 陶世如[4,3] 蒋玲曦[4,3] 

机构地区：[1]上海师范大学生命与环境科学学院,上海200234 [2]上海市农业科学院畜牧兽医研究所,上海201106 [3]上海市农业遗传育种重点实验室,上海201106 [4]上海市农业科学院生物技术研究所,上海201106

出　　处：《畜牧兽医学报》2010年第11期1435-1441,共7页ACTA VETERINARIA ET ZOOTECHNICA SINICA

基　　金：上海市科学技术委员会重大项目(07DZ19058);上海市自然科学基金(09ZR1428000)

摘　　要：本试验旨在建立有效的针对猪戊型肝炎病毒(HEV)的诊断技术及试剂盒。作者建立了应用猪HEVORF3筛选出来的一段序列合成肽swHEV11检测HEV血清抗体的间接ELISA诊断方法,确定的抗原最适包被浓度为3μg·mL-1;血清最适稀释度为1∶10,作用时间为30min;酶标抗体最适稀释度为1∶12000,作用时间为60min。用HEV ORF3合成肽swHEV11研制的猪HEV ELISA诊断试剂盒,与北京万泰试剂盒检测比较,符合率为73.4%(205/279),灵敏度和特异性分别为73.3%(200/273)和83.3%(5/6)。该试剂盒批内重复的变异系数<10%,批间变异系数<20%。结果表明,该方法的建立和所研制的猪戊型肝炎诊断试剂盒灵敏度高,特异性强,稳定性合格,为猪戊型肝炎病毒抗体的检测和流行病学的调查提供了一种简便快速的血清学诊断方法。The objective of this study was to establish effective diagnostic technique and diagnostic kit for swine hepatitis E virus（HEV）.An indirect ELISA assay was developed for detection of swine HEV using synthetic peptide swHEV11 screening from one sequence of the open reading frame（ORF） 3 of swine HEV.This assay was optimized for an antigen coating concentration of 3 μg·mL-1 and a serum dilution of 1：10,with a standard incubation time of 30 min.The optimal dilution and reaction time of enzyme labeled antibody was 1：12 000 and 60 min,respectively.When the kit was applied to detect HEV antibodies in sera of pigs,the results were quite consistent with the HEV diagnostic kit from Wantai.The conformity rate reached 73.4%（205/279）,the sensitivity rate and specificity rate reached 73.3%（200/273） and 83.3%（5/6）,respectively.The coefficient of variation（CV） was less than 10% in the same batch,and less than 20% in different batches.The results indicated that the application of indirect ELISA and the HEV ELISA kit is specific,sensitive and stable,and may provide a simple and rapid method for monitoring serum antibodies against HEV infection and seroepidemiological surveys of HEV infection.

关 键 词：猪戊型肝炎病毒 合成肽 间接ELISA 检测 诊断试剂盒 

分 类 号：S854.43[农业科学—临床兽医学] S858.285.3[农业科学—兽医学]