抗平颏海蛇短链神经毒素全人源单克隆抗体的筛选、制备及生物活性研究

摘要

Objective To prepare human anti-postsynaptic neurotoxin monoclonal antibody from phage antibody library using recombined postsynaptic short-chain neurotoxins ofLapemis curtus.Methods The three postsynaptic neurotoxins were expressed inEscherichia coli and phage antibodies against neurotoxins were screened. The obtained scFvs were further constructed to full antibodies. The antigen binding ability, biochemical and pharmacokinetic characteristics of the depurated antibodies were evaluated, and the anti-toxin effects of the antibody drugs were verifiedin vivo.Results Two positive scFvs with specific binding ability to all the three neurotoxins were obtained after 4 rounds of panning, and then the full antibodies were generated, expressed and purified. Antibody binding specificity was further confirmed. Pharmacokinetics of these two antibodies, SM-SD-911 and SM-SD-861 were similar to a conventional IgG molecule. SM-SD-911 and SM-SD-861 also showed strong antitoxin effectin vivo. Conclusionfull human anti-postsynaptic neurotoxin antibody has been successfully obtained by using recombinant neurotoxin technology and a large phage antibody library which may achieve clinical efficacy in navy medical applications.%目的 重组表达3种平颏海蛇短链神经毒素,利用噬菌体抗体库筛选制备全人源抗毒素单克隆抗体并评估其生物活性.方法 以重组表达纯化的神经毒素蛋白作为抗原,从自主构建的噬菌体抗体库中筛选获得噬菌体抗体,经序列测定确定其抗体类型后构建完整抗体.利用真核表达系统表达纯化抗体后鉴定其抗原结合活性、生化特性及药代动力学特性.在体内验证抗体药物的抗毒素作用.结果 经过4轮筛选,获得2株能同时结合3种神经毒素的噬菌体单链抗体,并将其制备成完整的抗体,进一步证实所获得的抗体可特异性结合3种抗原.2株全人源抗体均可拮抗神经毒素对昆明小鼠的致死作用.结论 利用重组神经毒素和噬菌体抗体库技术成功获得了特异性抗平颏海蛇短链神经毒素的全人源治疗性抗体.