中文摘要：

抗TNFαscFv在E.coliHB2151中以可溶性形式在胞周质中表达,为了进一步提高其表达量,文章通过摇瓶培养对其诱导表达条件进行了研究。将噬菌体抗体库筛选到的阳性克隆感染E.coliHB2151,分别改变诱导温度、诱导时间和诱导剂IPTG的浓度,并在培养基中加入甘氨酸和Triton X-100等添加剂,观察诱导条件的改变对scFv表达和分泌的影响。结果表明,E.coliHB2151在培养5 h对数生长期时开始诱导,在ITPG浓度为0.5 mmol/L条件下,30℃诱导20 h,胞周质中scFv的表达量比优化前增加了一倍,而且在诱导后期加入甘氨酸和Triton X-100后可增加scFv向培养基的分泌量。免疫学印迹法检测抗TNFαscFv能够与天然结构的TNFα结合,识别抗原的构象表位。该工作为其他可溶性scFv表达量的提高提供了可参考的模式。

英文摘要：

The induction conditions were studied in order to improve the soluble expression level of anti-TNFα scFv in periplasmic space of E.coli HB2151.The positive clone selected from thephage antibody library was infected into E.coli HB2151.The influences on scFv expression and secretion were observed with the alteration of induction temperature,time and IPTG concentration,as well as the addition of Glycine and Triton X-100.The binding activity of anti-TNFα scFv to antigen was determined by immunoblotting assay.Results showed that the expression level of scFv in the periplasmic space was increased one-fold after being cultured for 5 hours following with induction for 20 hours at 30 ℃ under 0.5 mmol/L IPTG.The secretion of scFv into the culture medium also increased with the addition of Glycine and Triton X-100.Immunoblotting assay showed that anti-TNFα scFv can bind to antigen TNFα with conformational epitope.This work applies a model for the improvement of soluble expression level for other kind of scFv.