北京地区牛传染性鼻气管炎血清学调查及其抗体检测间接ELISA方法的建立
发布时间：2018-07-13 15:14
【摘要】：为了解北京地区奶牛IBR流行情况,用IDEXX IBRgE抗体检测试剂盒检测来自北京不同地区的2582份牛血样。结果显示,调查的地区均有IBRV抗体阳性牛存在,其中昌平23.6%(245/1037),房山100%(50/50),延庆50.63%(202/399),顺义60%(9/15),丰台30.03%(164/546),密云43.46(232/535),总阳性率为34.93%(902/2582)。血清学调查结果表明,目前北京地区IBR流行情况严重,应尽快采取相应的措施。为建立一种检测IBRV的方法,试验将IBRV接种MDBK细胞,以浓缩纯化的IBRV作为包被抗原,建立了间接ELISA检测方法。通过棋盘法确定了抗原的最佳包被浓度和血清的最佳稀释倍数。抗原包被浓度20μg/mL,4℃过夜;PBST洗涤;3%BSA的PBST作为封闭液,37℃封闭2h;血清用PBST作1:80稀释,100μL/孔,37℃度作用1h;HRP-兔抗牛IgG用3%BSA的PBST作1:20000倍稀释,37℃作用1h;TMB底物室温避光显色10min。阴、阳性血清临界值为0.240。特异性试验结果表明,IBRV阳性血清被检为阳性,BVDV、牛白血病病毒、副结核病病毒阳性血清均被检为阴性。敏感性试验结果表明,IDEXX试剂盒中的标准阳性血清1:640倍稀释时仍能检出阳性抗体。与中和试验相比,特异性为100%(23/23),敏感性为82.76%(24/29)。批内和批间重复性试验的变异系数均小于10%,表明该方法重复性好。与IDEXX试剂盒相比,阳性符合率为87.7%(228/260),阴性符合率为98%(196/200)。以上结果表明所建立的间接ELISA方法可用于IBRV感染的监测,并替代目前市场上昂贵的IBR进口检测试剂盒。
[Abstract]:In order to understand the prevalence of IBR in dairy cows in Beijing, 2582 bovine blood samples from different areas of Beijing were detected with IDEXX IBRgE antibody test kit. The results showed that there were 23.6% (245r1037) in Changping, 100% (50 / 50) in Fangshan, 50.63% (2022 / 399) in Yanqing, 60% (9 / 15) in Shunyi, 30.03% (164 / 546) in Fengtai, 43.46 (232r535) in Miyun, the total positive rate was 34.93% (902 / 2582). The results of serological investigation showed that the epidemic situation of IBR in Beijing area was serious at present, and corresponding measures should be taken as soon as possible. In order to establish a method for detecting IBRV, an indirect Elisa method was established by inoculating IBRV into MDBK cells and using concentrated and purified IBRV as encapsulated antigen. The best coating concentration of antigen and the best dilution multiple of serum were determined by chessboard method. The concentration of antigenic coating was 20 渭 g / mL ~ (4 鈩，
本文编号：2119839
[Abstract]:In order to understand the prevalence of IBR in dairy cows in Beijing, 2582 bovine blood samples from different areas of Beijing were detected with IDEXX IBRgE antibody test kit. The results showed that there were 23.6% (245r1037) in Changping, 100% (50 / 50) in Fangshan, 50.63% (2022 / 399) in Yanqing, 60% (9 / 15) in Shunyi, 30.03% (164 / 546) in Fengtai, 43.46 (232r535) in Miyun, the total positive rate was 34.93% (902 / 2582). The results of serological investigation showed that the epidemic situation of IBR in Beijing area was serious at present, and corresponding measures should be taken as soon as possible. In order to establish a method for detecting IBRV, an indirect Elisa method was established by inoculating IBRV into MDBK cells and using concentrated and purified IBRV as encapsulated antigen. The best coating concentration of antigen and the best dilution multiple of serum were determined by chessboard method. The concentration of antigenic coating was 20 渭 g / mL ~ (4 鈩，
本文编号：2119839
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