基因重组抗原酶联免疫法检测兔出血症病毒抗体及其标准化  

Establishment and Standardization of Indirect ELISA Using the Recombinant VP60 for the Detection of Antibodies Against Rabbit Hemorrhagic Disease Virus

作　　者：王永山[1] 陆承平[1] 周宗安[2] 陈兴祥[3] 

机构地区：[1]南京农业大学动物医学院,江苏南京210095 [2]南京军区军事医学研究所 [3]江苏省农业科学院兽医研究所,江苏南京210014

出　　处：《中国兽医学报》2004年第3期233-235,239,共4页Chinese Journal of Veterinary Science

摘　　要：以重组兔出血症病毒 (RHDV) VP6 0蛋白为抗原 ,建立了 RHDV抗体间接 EL ISA检测方法。优化的试验反应条件为 :重组 VP6 0的包被质量浓度为 1.0 mg/ L ,用 10 %牛血清封闭 ,以大肠杆菌提取物稀释被检血清以消除非特异性反应。将所建立的 EL ISA与现行血凝抑制 (HI)试验比较发现 ,不同免疫状态的兔血清的 RHDV EL ISA抗体与 HI抗体均呈正相关。对 11个 RHD免疫兔场 1130份血清样品的抗体检测表明 ,各免疫兔群血清 RHDV抗体水平不完全一致 ,D值在 1.0 9～ 1.76之间 ,显著高于非免疫兔 (0 .0 5 )及 SPF兔 (0 .0 2 ) ,低于高免兔 (2 .34)。在此基础上 ,研制了RHDV抗体酶联免疫检测试剂盒 ,测定了其主要指标 ,制定了各成分的质量控制标准 。An indirect enzyme-linked immunosorbent assay(ELISA) for the detection of antibody against rabbit hemorrhagic disease virus(RHDV) was developed using the recombinant VP60 expressed highly in E.coli as antigen.The optimized conditions for the assay were that the wells of ELISA plate were coated with recombinant VP60(1.0 mg/L),10% normal bovine serum was added as blocking agent,and the serum samples were preincubated with the extract of E.coli.The result indicated that the extract could eliminate the non-specific antibodies in the serum samples.To compare the ELISA with hemagglutination inhibition(HI),the detection of the antibody of RHDV was carried out at the same time.The antibody titers in the two methods were regression.The results of the detection of (1 130) sera from 11 farms of rabbits immunized with RHD vaccine showed that the antibody titers were different.The establishment of the detection kit facilitated the immunological detection for the antibody of RHDV.

关 键 词：基因重组抗原酶联免疫法 检测 出血症病毒 抗体 VP60重组蛋白 间接ELISA 试剂盒 

分 类 号：S852.65[农业科学—基础兽医学] Q503[农业科学—兽医学]