传染性法氏囊病病毒VP4蛋白单克隆抗体的制备及鉴定

Preparation and identification of monoclonal antibodies against Infectious Bursal Disease Virus (IBDV) VP4

摘要:

传染性法氏囊病病毒 (IBDV) 蛋白VP4在抑制宿主免疫应答中起重要作用，为制备IBDV VP4的单克隆抗体，以实验室保存的融合蛋白His-VP4免疫BALB/c小鼠，经过细胞融合、筛选、亚克隆后获得4株能稳定分泌抗VP4的单抗杂交瘤细胞株，分别命名为3B3、3H11、4C8和4G6，经间接ELISA测定4株单抗的亲和力解离常数分别为4.61×10–11、1.71×10–10、4.26×10–11和5.02×10–11，均为高亲和力抗体。4株单抗的重链类型分别为IgG1、IgG1、IgG2b和IgG1。进一步以Western blotting鉴定，该4株单抗均能特异地识别IBDV的VP4蛋白，间接免疫荧光和Western blotting试验表明4株单抗均能识别IBDV感染DF-1细胞后产生的VP4蛋白。该单抗为检测IBDV以及研究IBDV VP4的生物学作用奠定了基础。

Abstract:

Infectious bursal disease virus (IBDV) VP4 plays an important role in immunosuppression of host. In order to develop monoclonal antibodies (McAbs) against VP4, we vaccinated BALB/c mice with His-VP4, screened and subcloned positive clones. We established 4 hybridoma cell lines that stably secreted McAbs against VP4 and named these cell lines 3B3, 3H11, 4C8 and 4G6, respectively. We tested the dissociation constant (Kd) of these McAbs, and found that their Kds were 4.61×10–11, 1.71×10–10, 4.26×10–11, 5.02×10–11, respectively. The isotypes of these McAbs were determined to be IgG1, IgG1, IgG2b and IgG1. These McAbs specifically bound to VP4 in IBDV infected DF-1 cells as demonstrated by Western blotting analysis and fluorescence antibody assay. These McAbs would help to detect IBDV infection and to analyze the biological activities of IBDV VP4.