两种来源的IBDV抗原制备单克隆抗体的比较

目的]比较以原核表达蛋白与纯化病毒制备单克隆抗体的特性。方法]RT-PCR扩增IBDVVP2基因,通过原核表达系统表达目的蛋白,并亲和纯化重组蛋白。尿囊液扩增IBDV,并通过超速离心纯化病毒。分别用纯化的重组VP2蛋白和IBDV免疫Balb/c小鼠,ELISA筛选杂交瘤细胞株。结果]获得了2株分泌VP2蛋白单克隆抗体的杂交瘤细胞株,腹水ELISA效价均为1∶2×104;4株分泌IBDV单克隆抗体的杂交瘤细胞株,腹水ELISA效价分别为1∶2×106,1∶6×104,1∶1×105,1∶4×103。所有单克隆抗体均与其制备用免疫原反应,不与空载体或其他病毒抗原反应。经10～20次传代,仍保持稳定的效价。结论]纯化的原核表达VP2蛋白和IBDV均能诱导小鼠产生免疫应答;用原核表达的VP2蛋白作为免疫原,可以获得特异的VP2抗体。

Comparison of Infectious Bursal Disease Monoclonal Antibodies Prepared with Two Different Immunogens

Objective]To compare the characteristics of monoclonal antibodies against infectious bursal disease virus (IBDV) prepared with two different immunogens,VP2 protein expressed by prokaryotic system and purified IBDV.Method]The first immunogen was produced by RT-PCR amplification of IBDV VP2 gene,prokaryotic expression and purification by affinity chromatography.The second immunogen was IB-DV purified by ultracentrifugation.Balb/c mice were immunized with the two immunogens,respectively,to prepare monoclonal antibodies.Positive hybridized cells were screened by ELISA.Result]Two cell lines secreting antibodies against IBDV VP2 protein were obtained,and their ELISA titers were 1∶2×104.Four cell lines secreting antibodies against IBDV were produced,and their ELISA titers were 1∶2×106, 1∶ 6×104,1∶1×105 and 1∶4×103,respectively.All monoclonal antibodies specifically bound to their own immunogen but did not react with other viruses or proteins.After 10-20 passages,these cell lines still secreted antibodies stably.Conclusion]The monoclonal antibodies prepared with recombinant IBDV VP2 protein or purified IBDV can induce immune response in mice,and VP2 specific monoclonal antibodies can be obtained with VP2 protein expressed by prokaryotic system as immunogen.