抗鸡传染性支气管炎病毒单克隆抗体的制备及其特性的研究

Studies on the Monoclonal Antibodies against Avian Infectious Bronchitis Virus

本实验将鸡传染性支气管炎病毒（澳大利亚Ｔ株）进行纯化，制成抗原，应用杂交瘤技术建立了５株分泌抗鸡传染性支气管炎病毒的杂交瘤细胞株，其免疫球蛋白的亚类分别属于ＩｇＧ１、ＩｇＧ３、ＩｇＭ。杂交瘤细胞分泌抗体的能力稳定，长期传代对其抗体分泌能力没有影响。５株单克隆抗体与其它１２株鸡传染性支气管炎病毒标准株之间存在不同程度的交叉反应，但对鸡新城疫病毒、传染性法氏囊病病毒、产蛋下降综合征病毒及禽流感病毒均无交叉反应。用此杂交瘤细胞制备腹水的ＥＬＩＳＡ效价为１０－６～１０－７，抗体对热的稳定性有一定的差别。抗体纯化后，经ＳＤＳ－ＰＡＧＥ凝胶电泳表明具有单克隆抗体的特征，经快速蛋白分析鉴定其纯度为８９．４９％。竞争ＥＬＩＳＡ法证明其中４株的单抗与抗原结合位点并不完全相同。采用单抗夹心ＥＬＩＳＡ法，对抗原的检测方法进行了初步探索。

Five monoclonal antibodies(McAbs) were produced against T strain of Australia of Avian Infectious Bronchitis Virus (IBV) using hybridoma technique.After verification by calculating chromosome and testing of antibodies secreting stability,five strains of specific antibodies producing hybridoma were successfully established.Their isotype were IgG 1、IgG 3、IgM. Varied degrees of cross reaction were noted between the McAbs and other 12 IBV standard strains.However,no cross reaction was observed between the McAbs and Newcastle disease virus,infectious bursal disease virus,avian influenza virus(N 9H 2),egg drop syndrome virus respectively. After BALB/c mouse peritoneal cavity inoculation with the hybridoma cell,the monoclonal antibodies titer of ascites were variable amongst 10 6 to 10 7. Competitive Enzyme linked immunosorbent assay (ELISA) unveiled that four McAbs were specifically combined with three epitope sites of the T strain of IBV.Western blotting indicated that the epitopes recongized by McAbs B3 4 was present on Nprotein of IBV.