人源抗破伤风毒素单链抗体(ScFv)的原核表达、纯化及功能鉴定

实验通过DNA重组技术从一株可中和破伤风毒素的人源单克隆抗体细胞(G6)中扩增出了抗体VH、VL的基因,通过重叠PCR使连接片段与VH、VL连接成单链ScFv。经测序证实VH、VL为抗体的可变区序列,命名为ScFv-G6。将ScFv-G6连接转化PET/26b质粒,构建了抗体的表达载体,被命名为PET/26b/ScFv-G6。以该载体在大肠杆菌中分泌表达产物经Ni-亲和柱纯化后的小鼠试验证实,可抵抗破伤风毒素的攻击,表明为中和抗体。具有组织穿透力强,不易过敏,可直接靶向于毒素等特点,适合于破伤风的防治,具有重要的应用价值。

Expression, purification and functional identification of the humanized monoclonal antibody against ScFv of tetanuss toxin

The gene VH and VL,was amplified from a human monoclonal antibody line(G6) against single chain variable fragment(ScFv) of tetanus toxin through DNA recombination technology.The linker fragment was linked up with gene VH and VL into the ScFv by overlapping PCR.This sequence,by the name of ScFv-G6,ScFv-G6 was linked into the plasmid PET/26b to construct the antibody expression vector,named as PET/26b / ScFv-G6.The expression product of this vector could be secreted in E.coli and this product was purified with the Ni-affinity column.The mouse assay results showed that this product provides the protective effect against the attack of tetanus toxin.This neutralizing antibody is of the advantage that the penetrating power is strong,is not easy to induce the allergic reaction and can be directly targeted to toxin.It is suitable for the prevention and treatment of tetanus.