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从分泌抗苏丹红Ⅰ单克隆抗体(mAb)的杂交瘤细胞株SU211中克隆抗体轻链Lc基因和重链Fd基因,将Lc、Fd基因以及Linker克隆到载体pET24a(+)中,构建scFab表达载体并在E.coli BL21(DE3)获得表达,SDSPAGE和Westernblot分析表明其表达形式为包涵体,相对分子质量约为57 kD,与理论预期值相符。经复性后,ELISA鉴定结果表明复性抗体对苏丹红Ⅰ具有很高的结合特异性。本研究为新型基因工程抗体的构建以及苏丹红新型免疫检测技术的建立奠定基础。
The antibody light chain Lc gene and heavy chain Fd gene were cloned from the hybridoma cell line SU211 secreting anti-Sudan I monoclonal antibody (mAb), and the Lc, Fd gene and Linker were cloned into vector pET24a (+) to construct scFab expression The vector was expressed in E.coli BL21 (DE3). The expressed product was identified as inclusion body by SDS-PAGE and Western blot analysis. The relative molecular mass was about 57 kD, which was consistent with the expected value. After refolding, the results of ELISA showed that the refolding antibody had high binding specificity to Sudan Ⅰ. This study laid the foundation for the construction of novel genetic engineering antibodies and the establishment of Sudan’s new immunoassay technique.