摘要
为建立牛冠状病毒(BCV)检测方法,利用构建的pET30a-N重组质粒高效表达了BCV重组N蛋白,western blot检测该重组蛋白具有很好的免疫活性。以纯化的蛋白作为包被抗原,通过方阵试验确定了抗原的最适包被浓度为1.75μg/mL,血清最佳稀释倍数为1∶200,酶标二抗最佳稀释倍数为1∶8000,建立了检测BCV抗体的间接ELISA方法。用该法对黑龙江一些地区采集到的256份牛血清样品进行检测,结果阳性率为65.23%,与病毒中和试验方法的检测结果符合率达95.31%。本研究建立的间接ELISA方法为BCV的检测和区域流行病学调查提供了一种快速简便的血清学诊断方法。
An indirect ELISA was established for the rapid detection of specific bovine coronavirus （BCV） antibodies in cattle using a recombinant N protein as an antigen. The recombinant N protein was expressed from recombinant plasmid pET30a-N in Escherichia coll. Western blot confirmed that the expressed fusion protein could specifically react with antiserum against BCV. This assay was optimized using 1.75 μg/mL N protein as coat antigen, 1：200 dilution of testing serum and 1：8 000 dilution of HRP-labeled goat anti-bovine IgG. Test against 256 serum samples from Heilongjiang province by this assay detected 65.23 % positive, which showed 95.31% agreement with VN test.
出处
《中国预防兽医学报》
CAS
CSCD
北大核心
2009年第8期618-622,共5页
Chinese Journal of Preventive Veterinary Medicine
基金
黑龙江农垦总局攻关课题(HNKXIV-08-07)
