2型猪链球菌谷氨酸脱氢酶单克隆抗体的制备与鉴定

目的制备抗猪链球菌（Streptococcus suis,S.suis）谷氨酸脱氢酶（glutamate dehydrogen-ase,GDH）的单克隆抗体。方法分别以2型S.suis全菌细胞和重组GDH蛋白为免疫原免疫BALB/c小鼠;采用淋巴细胞杂交瘤技术将免疫后的小鼠脾细胞与SP2/0骨髓瘤细胞融合;酶联免疫吸附试验（ELISA）筛选分泌抗GDH单克隆抗体的杂交瘤细胞株并克隆化;免疫印迹（Western blot）鉴定GDH单克隆抗体的特异性;制备小鼠腹水并进行抗体效价测定。结果细胞融合后经ELISA筛选获得分泌的阳性细胞株,经过有限稀释克隆化筛选,获得4株能稳定分泌抗GDH单克隆抗体的杂交瘤细胞株。单抗腹水的抗体效价均在1∶102400以上。结论本研究获得了4株能稳定分泌抗体且其分泌的抗体能够与GDH蛋白进行特异性反应的单克隆抗体细胞株,所获得特异性单克隆抗体能用于猪链球菌结构和功能的研究,也能够进一步用于猪链球菌感染血清学诊断方法研究。

Preparation and identification of monoclonal antibodies against GDH of Streptococcus suis serotype 2 highly virulent strain isolated from china

Objective To prepare monoclonal antibodies（McAbs）against glutamate dehydrogenase（GDH）protein of Streptococcus suis（S.suis）.Methods S.suis isolates and the GDH recombinant protein were used as antigen to immunize female BALB/c mice respectively.The immunized splenic cells of mice were fused with murine myeloma SP2/0 cells.The hybridoma cell strains secreting McAb against GDH were screened with enzyme linked immunosorbent assay（ELISA）.The specificity of the secreted GDH McAbs was confirmed by Western blot.The ascites were harvested and the antibody titer was measured.Results Four hybridoma cell lines secreting McAb against GDH were obtained by fusion of mouse myeloma cells SP2/0 with splenic cells from BALB/c mice immunized with the purified expression product of GDH.Titers of McAbs in ascites were higher than 1∶102 400.Conclusion The McAbs against GDH were successfully prepared and identified,which would be useful for the detection of S.suis infection.