抗伏马菌素B_1单链抗体的原核表达及其抗体活性和稳定性分析

Prokaryotic expression of antibody single-chain variable fragment (scFv) against fumonisin B_1 and characterization of its antibody activity and stability

研究了对抗伏马菌素B1单链抗体在不同原核表达条件下表达情况及其抗体稳定性,以促进其在免疫学分析中的应用。通过构建表达载体pET22b-1D11和pMAL-1D11,分析了诱导温度、诱导剂浓度、宿主菌和高溶解性标签蛋白对单链抗体表达形式的影响;采用酶联免疫吸附实验法(ELISA)分析了单链抗体的热稳定性和有机溶剂耐受性。结果表明:条件优化后,单链抗体及其融合蛋白主要以包涵体形式表达;热处理会降低单链抗体活性甚至使其完全失活,甲醇会降低基于单链抗体的ELISA体系的灵敏度。

Prokaryotic expression under different condition and stability of antibody single-chain variable fragment（scFv） against fumonisin B1,a pernicious mycotoxic,were investigated for promoting its application in immunoassay.After construction of expression vector pET22b-1D11 and pMAL-1D11,we analyzed the influence of temperature,concentration of inducer,host bacteria,and high soluble protein tag on scFv expression form.And the thermal stability and organic solvent resistance of scFv were analyzed by enzyme-linked immunosorbent assay（ELISA）.The results showed that scFv and its fusion protein were mainly in the formation of inclusion body after optimization of the above conditions.Heat treatment could reduce or even inactivate the scFv activity,and methanol could reduce the sensitivity of scFv-based ELISA.