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当前位置: 首页 > 文章 > 猪戊型肝炎病毒ORF2主要抗原表位区间接ELISA方法的初步建立 中国畜牧兽医 2010,37 (5) 101-104
Position: Home > Articles > Development of Indirect ELISA with Major Epitope Domain of ORF2 of SHEV China Animal Husbandry & Veterinary Medicine 2010,37 (5) 101-104

猪戊型肝炎病毒ORF2主要抗原表位区间接ELISA方法的初步建立

作  者：
闻晓波;王密;冉旭华;周恩民;李晓娟;侯喜林;朴范泽
单  位：
黑龙江八一农垦大学动物科技学院预防兽医学省重点实验室
关键词：
猪戊型肝炎病毒;ORF2;主要抗原表位区;ELISA
摘  要：
应用原核表达系统表达猪戊型肝炎病毒ORF2蛋白C端和N端的主要抗原表位区,重组蛋白命名为ORF2-C和ORF2-N,初步建立间接ELISA诊断方法。用重组蛋白ORF2-C和ORF2-N作为诊断抗原,对反应条件进行优化,初步建立ELISA诊断方法。抗原最适包被浓度ORF2-C为8μg/mL、ORF2-N为12μg/mL;血清最适稀释度均为1∶50,ORF2-C作用时间为50min、ORF2-N作用时间为90min;酶标抗体最适稀释度为1∶5000;ORF2-C判定标准:D450nm值≥0.348为阳性,D450nm值<0.348为阴性;ORF2-N判定标准:D450nm值≥0.397为阳性,D450nm值<0.397为阴性。与戊型肝炎病毒诊断试剂盒检测结果相比,阳性符合率分别为93.3%、86.7%。利用重组蛋白建立的ELISA方法特异性、敏感性和重复性均较好,且ORF2-C的检测效率明显高于ORF2-N,该方法的初步建立为进一步完善猪戊型肝炎病毒诊断方法奠定基础。
译  名：
Development of Indirect ELISA with Major Epitope Domain of ORF2 of SHEV
作  者：
WEN Xiao-bo,WANG Mi,RAN Xu-hua,ZHOU En-min,LI Xiao-juan,HOU Xi-lin,PIAO Fan-ze(Provincial Key Laboratory of Preventive Veterinary Medicine,College of Animal Science and Technology,Heilongjiang Bayi Agricultural University,Daqing 163319,China)
关键词：
swine Hepatitis E; ORF2; major epitope domain; ELISA
摘  要：
C-terminal and N-terminal major epitope domain of ORF2 of swine Hepatitis E (SHEV) were express in E.coli,denominated ORF2-C and ORF2-N respectively. Indirect ELISA assay were developed to detect antibody of swine hepatitis E virus using recombinant proteins. Recombinant proteins were used as diagnostic antigen. Indirect ELISA assay were develop for detect antibody of SHEV. This assay was optimized for ORF2-C and ORF2-N antigen coating concentration of 8 and 12 μg/mL respectively and a serum dilution of 1∶50,with a standard incubation time of 50 and 90 min respectively. ORF2-C a reading of D450 nm≥0.348 and ORF2-N a reading of D450 nm≥0.397 was scored as positive. The positive coincident rate between the fusion protein ELISA and the available Kit was up to 93.3 % and 86.7% respectively in detecting swine HEV antibody. The result showed the indirects ELISA had a better sensitivity,specificity and repeatability. ORF2-C was significantly better than the ORF2-N used as coating antigen. This research was beneficial to further development of diagnostic kit for swine hepatitis E.

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