[1]王 磊,常继涛,于 力*.基于牛呼吸道合胞体病毒融合前F蛋白的间接ELISA方法的建立与应用[J].中国预防兽医学报,2019,(11):1126-1130.[doi:0.3969/j.issn.1008-0589.201903004]
 WANG Lei,CHANG Ji-tao,YU Li*.Establishment and application of an indirect ELISA using Prefusion-F protein antigen for the detection of antibodies against bovine respiratory syncytial virus[J].Chinese journal of preventive veterinary medicine,2019,(11):1126-1130.[doi:0.3969/j.issn.1008-0589.201903004]
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基于牛呼吸道合胞体病毒融合前F蛋白的间接ELISA方法的建立与应用()
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《中国预防兽医学报》[ISSN:1008-0589/CN:23-1417/S]

卷:
期数:
2019年11
页码:
1126-1130
栏目:
诊断和检测技术
出版日期:
2019-12-25

文章信息/Info

Title:
Establishment and application of an indirect ELISA using Prefusion-F protein antigen for the detection of antibodies against bovine respiratory syncytial virus
文章编号:
1008-0589(2019)11-1126-05
作者:
 

王 磊常继涛于 力*

 (中国农业科学院哈尔滨兽医研究所 兽医生物技术国家重点实验室/牛羊传染病研究创新团队,黑龙江 哈尔滨 150069)
Author(s):
 

WANG Lei CHANG Ji-tao YU Li*

 (Division of Livestock Infectious Disease, State Key Laboratory of Veterinary Biotechnology, Harbin Veterinary Research Institute,
Chinese Academy of Agricultural Sciences, Harbin 150069, China)
关键词:
牛呼吸道合胞体病毒融合前的F蛋白间接ELISA 抗体检测
Keywords:
 Bovine respiratory syncytial virus (BRSV)  prefusion-F protein  indirect ELISA  antibody detection
分类号:
S852.65
DOI:
0.3969/j.issn.1008-0589.201903004
文献标志码:
A
摘要:
为确定牛呼吸道合胞体病毒(BRSV)在我国的流行情况,本研究以BRSV融合前F蛋白作为包被抗原,经各项条件优化后建立了检测BRSV抗体的间接ELISA方法:确定抗原最佳包被浓度为2 mg/L、待检血清的最佳稀释度为1∶100、最佳封闭液为3 %明胶、羊抗牛IgG-HRP的稀释度为1:8 000,抗体阴阳性临界值为0.204。特异性试验结果显示,该ELISA方法与阿卡斑病毒(AKAV)、牛轮状病毒(BRV)、蓝舌病病毒(BTV)、口蹄疫病毒(FMDV)、牛副流感病毒3型(BPIV3)、牛病毒性腹泻病毒(BVDV)以及牛传染性鼻气管炎病毒(IBRV)的阳性血清无交叉反应。敏感性试验结果显示,当BRSV阳性血清稀释倍数达1∶6 400时仍为阳性结果,表明该方法具有较高的敏感性。重复性试验结果显示,ELISA组内、组间最大变异系数分别为6.80 %和8.80 %,表明该方法的重复性良好。对195份临床牛血清样品检测结果显示,BRSV抗体阳性率为61.0 %;对其中的143份血清同时进行病毒中和试验检测,结果显示二者符合率为89.5 %。上述结果表明本研究建立的间接ELISA方法可以用于牛群中BRSV抗体的检测,为BRSV的血清流行病学调查提供可靠的技术手段。
Abstract:
An indirect ELISA was established for monitoring the prevalence of bovine respiratory syncytial virus (BRSV) infection in China using prefusion-F protein as a coating antigen. A total of 195 serum samples from different regions in China were detected by indirect ELISA. The reaction conditions were optimized, including 2 mg/L purified prefusion-F protein as coating antigen, 1:100 dilution of testing serum, 3% gelatin as the best sealing solution and 1:8 000 dilution of IgG-HRP conjugated goat anti-bovine IgG, and the cut off-value was determined as OD450nm 0.204. The ELISA established in this study was tested for specificity of detecting antibodies against BRSV and no cross-reaction was detected with positive sera against Akabane virus (AKAV), Bovine rotavirus (BRV), Bluetongue virus (BTV), Foot-and-mouth disease virus (FMDV), Bovine parainfluenza virus type 3 (BPIV3), bovine virus diarrhoea virus (BVDV) or Bovine rhinotracheitis virus (IBRV). Sensitivity test results showed that when the dilution ratio of BRSV antibody-positive serum reached 1:6,400, it was still a positive result, indicating that the method had a high sensitivity. The inter- and intra-assay in ELISA demonstrated that the coefficient of maximum variation was 6.80% and 12.2% respectively, indicating that the repeatability of this indirect ELISA is good. In the detection of 195 serum samples, the positive rate of ELISA was 61.0% (119/195). Of this, 143 serum samples were tested simultaneously by virus neutralizing test. The coincidence rate of the ELISA with the neutralizing test was 89.5%. These results indicated that the developed ELISA method could be used for the detection antibodies against BRSV and provide a reliable method for the epidemiological investigation of BRSV.

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(本文编辑:李 娜;

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更新日期/Last Update: 2019-12-09