人源性抗HBsAg单链抗体在大肠埃希菌中分泌表达条件的研究

目的 研究抗HBsAS单链抗体A-15在大肠埃希菌中的高效表达,增加该抗体在培养基中的产量.方法 改变工程菌的诱导时机、诱导剂浓度和诱导时间,以观察这些因素对单链抗体表达的影响.另外,加入不同浓度的蔗糖、甘氨酸和Triton X-100,以考察三种化学物质对单链抗体在培养基中分泌量的影响.结果 工程菌培养4 h后,加终浓度0.5 mmol/L IPTG诱导表达8 h为抗HBsAS单链抗体A-15较佳的表达条件.终浓度0.3 mol/L蔗糖,2%甘氨酸和1%Triton X-100同时加入诱导时的培养基中,可使培养基中表达的单链抗体亲和活性分别增加16.78倍.经测定抗HBsAg单链抗体A-15在培养基中最终表达量为7.4 mg/L.结论 抗HBsAg单链抗体A-15的分泌表达条件得到优化,表达量明显提高,这为更有效地制备该抗体用于其结构和功能研究提供了一个切实可行的方法.

Optimization of human anti-HBsAg scFv secretary expression in Escherichia coll

Objective To explore the conditions for high expression of anti-HBsAg scFv A-15 in E. coli, increase the production of the scFv in the culture medium. Methods By changing induction occasion, concentration of inductor IPTG and induction time, influence of various conditions on expression of anti-HBsAg scFv A-15 was analyzed through ELISA. In addition, the effects of sucrose, glycine and Triton X-100 at different concentrations on the scFv excretion into culture medium was evaluation. Results The optimal expression conditions were as follows: the induction was started after culturing for 4 h, the concentration of IPTG was 0.5 mmol/L,and the induction lasted for 8 h .The scFv affinity in culture medium with 0.3 mol/L sucrose,2% glycine, 1% Triton X-100,16.78-fold higher, respectively than that without the three chemicals. The final yield of anti-HBsAg scFv A-15 was estimated to be 7.4 mg/L. Conclusion The conditions for production of anti-HBsAg scFv A-15 were optimized,which provides a practical method for more efficient production of the scFv in E. coli for further studying structure and function.