基于不同重组小反刍兽疫病毒N蛋白建立的双抗原S-ELISA抗体检测方法的比较研究

A comparative study on double-antigen S-ELISA for the detections of antibodies based on different recombinant N protein of peste des petits ruminants virus

为探索建立小反刍兽疫病毒抗体双抗原夹心ELISA方法的原料,本研究成功使用pET-30a与p ET-32a两个不同原核载体分别表达与纯化了小反刍兽疫病毒核衣壳蛋白N抗原。使用不同原核载体表达的N抗原组合建立的双抗原夹心ELISA抗体检测方法、相同原核载体表达的N抗原组合建立的双抗原夹心ELISA抗体检测方法、N抗原包被建立的抗体间接ELISA检测方法以及血清中和试验方法分别对已知背景的小反刍兽疫抗体阳性血清与小反刍兽疫抗体阴性血清进行检测。综合考虑方法的敏感性、特异性和稳定性,最终选择用不同原核载体表达的pET-30a-(N)与pET-32a-(N)抗原组合建立的小反刍兽疫病毒抗体双抗原夹心ELISA检测方法。

To establish a double-antigen S-ELISA method for the detection of peste des petits ruminants virus antibodies,the nucleocapsid protein(N antigen) were successfully expressed and purified with two different prokaryotic vectors(p ET-30 a and p ET-32 a).The double-antigen S-ELISA antibody detection method based on the combination of N antigens expressed by different prokaryotic vectors,the double-antigen S-ELISA antibody detection method based on the combination of N antigens expressed by the same prokaryotic vectors,the indirect ELISA method based on N antigen coating and the serum neutralization test method were completed to detect the positive and negative serum of peste des petits ruminants with known background respectively. Considering the sensitivity,specificity and stability of the methods,a double-antigen S-ELISA method for the detection of peste des petits ruminants virus antibody was established by combining p ET-30 a-(N) and p ET-32 a-(N) antigens expressed in different prokaryotic vectors.