【摘要】 将表达类志贺毒素II型变异体B亚单位融合蛋白的菌株ppSLT-IIeB在含氨苄青霉素的2×YTA培养基中培养,至对数生长期时加入IPTG诱导,诱导的重组菌超声波裂解后,经聚丙烯酰胺凝胶电泳证明其已表达融合蛋白。用谷胱苷肽活化的Sepharose4B亲和层析柱纯化融合蛋白,测定纯化蛋白的浓度,免疫BALB/C小鼠,细胞融合后经ELISA检测,得到一株阳性杂交瘤细胞,命名为7C3-1,制备的单抗腹水ELISA效价为210,Westernblet结果表明此单抗仅与融合蛋白反应,而与载体蛋白不反应。利用此单抗检测18株水肿病菌株,发现有13株产类志贺毒素II型变异体。更多还原

【Abstract】 The toxin SLTIIe was difficult to purify,so we expressed fusion protein GSTSLTIIeB in recombinant E.coli ppSLTIIeB.The fusion protein purified by Glutathione Sephrose 4B as antigen immunized BALB/C mice. GST expressed in recombinant E.coli ppGEX6P1 as negative antigen and SLTIIeB as positive antigen absorbed in ELISA plate. SP2/0 myelomacells and spleen cells of the immunized BALB/C mice were fused by PEG2000. The hybridoma culture supernatants were screened for antiSLTIIeB antibodies by indirectELISA and cytotoxicity neutralization. One monoclonal antibody 7C31 was detected and only reacted with a 33 000 fusion protein band in Western blot. The Mab as probe had detected 18 strains of ED by DotELISA, which showed 13 strains were positive.更多还原