论文部分内容阅读
目的建立抗伏马菌素B1(FB1)的单克隆杂交瘤细胞株,制备抗FB1的单克隆抗体(McAb)。方法采用FB1-KLH偶联物小剂量长周期免疫BALB/c小鼠后,采用细胞融合方法获得分泌抗FB1的杂交瘤细胞株,采用多次亚克隆的方法建立了4株稳定分泌抗FB1抗体的杂交瘤细胞株。腹水诱生法获得大量McAb,采用抗体亚类测定试剂盒鉴定抗体的亚类,SDS-PAGE测定抗体分子量,McAb的特异性和灵敏度用间接竞争抑制ELISA。结果免疫小鼠血清检测结果显示经过5次免疫后血清的效价稳定在1×10-6,经过4次亚克隆后建立4株稳定分泌抗FB1抗体的杂交瘤细胞株,获得腹水,纯化抗体,抗体亚类属于IgG2a类,轻链为κ,轻链和重链的相对分子质量分别是55000和32000,ELISA检测抗体特异性显示可与FB1发生特异性反应,采用此抗体建立间接竞争抑制ELISA方法的线性范围在2～500ng/ml。结论制备了特异性和灵敏度较高的抗FB1单克隆抗体,为建立伏马菌素免疫学检测方法打下良好基础。
Objective To establish a monoclonal hybridoma cell line resistant to fumonisin B1 (FB1) and prepare a monoclonal antibody (McAb) against FB1. Methods BALB / c mice were immunized with FB1-KLH conjugate at low dose and long period. Hybridoma cell lines secreting anti-FB1 were obtained by cell fusion method. Four strains of stable secreting anti-FB1 antibody Of hybridoma cell lines. A large number of McAbs were obtained by ascitic fluid induction. Antibody subclass was identified by using antibody subclass determination kit. The molecular weight of antibody was determined by SDS-PAGE, and the specificity and sensitivity of McAb were tested by indirect competitive inhibition ELISA. Results The results of sera from immunized mice showed that the titer of serum was stable at 1 × 10-6 after 5 immunizations and 4 strains of hybridoma cell lines stably secreting anti-FB1 antibody were established after 4 subclones to obtain ascites and purified antibody , The antibody subclass belongs to IgG2a class, the light chain is κ, the relative molecular masses of light chain and heavy chain are 55000 and 32000, respectively. The specificity of ELISA detection antibody shows that it can react specifically with FB1. The antibody was used to establish indirect competitive inhibition ELISA The linear range of the method is 2 ~ 500ng / ml. Conclusion The anti-FB1 monoclonal antibody with high specificity and sensitivity was prepared and laid a good foundation for the establishment of immunological detection of fumonisin.