Kheyar A, Martin S, St-Laurent G, Timoney P J, McCollum W H, Archambault D
Département des Sciences Biologiques, Université du Québec à Montréal, Canada.
Clin Diagn Lab Immunol. 1997 Nov;4(6):648-52. doi: 10.1128/cdli.4.6.648-652.1997.
To provide a convenient and sensitive method for the detection of equine arteritis virus (EAV)-specific serum antibodies, we developed an immunoblot assay employing the EAV nucleocapsid (N) and membrane (M) proteins expressed in a procaryotic expression vector (pMAL-c2) for the production of recombinant maltose-binding (MBP) fusion proteins (MBP-N and MBP-M). The antigenic reactivity of the recombinant fusion proteins and their Xa factor cleavage EAV products was confirmed by immunoblot using horse antisera to EAV. Some horse sera, however, showed immune reactivity to the MBP fusion partner protein. Based on a total of 32 horse sera analyzed for the presence of EAV antibodies by immunoblot, using the MBP-N or -M fusion proteins and the Xa factor cleavage EAV products, and in the serum neutralization test, there was 100% concordance between the assays. Sera from horses experimentally infected with EAV were reactive in the immunoblot test with both the MBP-N and the MBP-M fusion proteins by day 14 after EAV exposure. The reactivity continued to the end of the experiment at day 145 after infection. This immune reactivity correlated with the detection of neutralizing antibodies in the serum samples. Based on these findings, the recombinant N and M proteins might be useful for serodetection of EAV-infected animals.
为了提供一种方便且灵敏的检测马动脉炎病毒（EAV）特异性血清抗体的方法，我们开发了一种免疫印迹分析方法，该方法采用在原核表达载体（pMAL-c2）中表达的EAV核衣壳（N）蛋白和膜（M）蛋白来生产重组麦芽糖结合（MBP）融合蛋白（MBP-N和MBP-M）。通过使用马抗EAV血清进行免疫印迹，证实了重组融合蛋白及其Xa因子切割的EAV产物的抗原反应性。然而，一些马血清对MBP融合伴侣蛋白表现出免疫反应性。基于通过免疫印迹分析的总共32份马血清中EAV抗体的存在情况，使用MBP-N或-M融合蛋白以及Xa因子切割的EAV产物，并结合血清中和试验，这些检测方法之间的一致性为100%。在EAV暴露后第14天，实验感染EAV的马的血清在免疫印迹试验中与MBP-N和MBP-M融合蛋白均发生反应。这种反应性在感染后第145天实验结束时仍持续存在。这种免疫反应性与血清样本中中和抗体的检测相关。基于这些发现，重组N蛋白和M蛋白可能对EAV感染动物的血清学检测有用。
