动物源性人兽共患细菌病防控生物新制剂的研究Ⅰ．分泌抗大肠杆菌O157：H7单克隆抗体杂交瘤细胞株的建立  

Novel Biological Products for Prevention and Control of Bacterial Zoonoses Ⅰ. Establishment of the Hybridoma Producing Monoclonal Antibodies against Enterohemorrhagic E. coli O157∶H7

作　　者：夏兴霞[1] 刘洁[1,2] 王永山[1] 诸玉梅[1] 欧阳伟[1,2] 何孔旺[1] 张雪寒[1] 

机构地区：[1]江苏省农业科学院兽医研究所 农业部动物疫病诊断与免疫重点开放实验室 国家兽用生物制品工程技术研究中心,江苏南京210014 [2]南京农业大学动物医学院,江苏南京210095

出　　处：《江苏农业学报》2009年第2期291-295,共5页Jiangsu Journal of Agricultural Sciences

基　　金：国家支撑计划资助项目(2007BAD40B01)

摘　　要：将E．coli O157：H7（EHEC—O157：H7）用甲醛灭活、超声破碎，免疫BALB／c小鼠，制备免疫脾细胞；与SP2／0骨髓瘤细胞融合后，获得7株分泌抗E．coli O157：H7单克隆抗体（mAbs）的杂交瘤细胞株，分别命名为F5、B8、E7、G9、G11、F1和C3，Ig亚类分别是IgG2ak、IgG2aK、IgG1K、IgMK、IgMh、IgMK和IgG2aK。用间接ELISA检测，E7和F1细胞培养上清液的效价为1．6×10^3，腹水效价均为1×10^11；C3、F5、B8、G9和G11细胞培养上清液的效价为1×10～2×10^2，腹水效价均为1×10^3。相加ELISA结果显示，F1、G9和G113株mAbs对应相同的抗原表位，而其它4株对应不同的抗原表位。用间接ELISA分析特异性，7株mAbs与某些其他血清型的大肠杆菌有交叉反应。Westernblotting表明，c3、E7和F13株mAbs在蛋白分子量28×10^3处有特异性条带，而其它4株mAbs则未显示蛋白带。Seven hybridomas secreting monoelonal antibodies （mAbs） against enterohemorrhagic E. coil O157：H7 （EHEC-O157： H7）were established by fusing SP2/0 with spleen cells from BALB/c mice immunized with formaldehydekilled and ultrasonically-broken E. coli O157： H7, naming E7, F1, C3, F5, BS, G9 and G11. The Ig isotypes of mAbs were IgG2aK, IgG2aK, IgG1K, IgMK, IgMk, IgMK and IgG2ak, respectively. The ELISA antibody titers of E7 and F1 were 1.6 × 10^3 and 1 ×10^11 in supernatant and ascitic fluid respectively, the others were 1 × 10 ^-2 × 10^2 in supernatant,and 1 × 10^3 in the ascitic fluids. Additivity ELISA indicated that F1, G9 and G11 were corresponding to the same antigen epitope, but the other 4 mAbs were different epitopes. The 7 mAbs could react with some other kinds of bacteria in the specific analysis by indirect ELISA. In the western-blotting, mAbs C3, E7 and F1 displayed the stained protein band of approximate 28 × 10^3 ,but the other 4 mAbs didn＇t show any stained protein band.

关 键 词：肠出血性大肠杆菌O157∶H7 单克隆抗体(mAb) 生物制剂 人兽共患病 

分 类 号：Q785[生物学—分子生物学]