抗传染性支气管炎病毒M蛋白单克隆抗体的制备及鉴定

Preparation and Identification of Monoclonal Antibodies Against M Protein of Avian Infectious Bronchitis Virus

将传染性支气管炎病毒M41毒株通过接种鸡胚的方法扩繁,从含病毒的尿囊液中提取总RNA,RT-PCR扩增M基因,构建原核表达载体pGEX-6p-1-M,导入大肠杆菌Transetta(DE3)内诱导表达。将纯化后的重组M蛋白作为免疫原免疫BALB/c小鼠,通过杂交瘤技术制备能稳定分泌抗M蛋白单克隆抗体的杂交瘤细胞。通过小鼠体内诱生腹水的方法大量制备抗M蛋白的单克隆抗体,用辛酸/硫酸铵方法纯化腹水,并对纯化的单克隆抗体进行了鉴定。

The infectious bronchitis virus(IBV)M41 strain was propagated in chicken embryos.Total RNA was extracted from the allantoic fluid.M gene was amplified by RT-PCR to construct the prokaryotic expression vector pGEX-6p-1-M,through which the recombinant M protein was expressed in E.coli Transetta(DE3).BALB/c mice were immunized with the purified recombinant M protein.Anti-M protein monoclonal antibody hybridoma cells were prepared by hybridoma technique.Anti-M protein monoclonal antibodies were prepared by in vivo induction ascites method.Ascites was purified by caprylic acid-ammonium sulfate method,and the purified monoclonal antibody was identified.