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目的 制备丙型肝炎病毒 (HCV )非结构蛋白NS5 (NS5 )的抗独特型单链可变区抗体scFv(抗 IdscFv) ,为研制HCVNS5的抗 IdscFv疫苗奠定基础。方法 采用噬菌体表面展示技术 ,将HCVNS5单克隆抗体固相包被于Nunc板 ,从噬菌体单链可变区抗体库中经过 5轮“吸附 洗脱 扩增”筛选过程 ,随机挑选 80个克隆 ,利用酶联免疫吸附试验 (ELISA)、交叉反应和竞争抑制实验 ,对其进行免疫学检测 ,获得与HCVNS5单克隆抗体结合活性较强的抗 IdscFv阳性克隆 ,并对HCVNS5特异性抗 IdscFv的编码序列进行序列测定分析。结果 筛选得到的HCVNS5抗 IdscFv片段由 786bp组成 ,具有结合HCVNS5单克隆抗体的生物学活性和特异性。结论 用噬菌体抗体库技术能够成功地获得HCVNS5的抗 IdscFv。本实验结果为开展用抗 IdscFv防治丙型肝炎的研究创造了条件。
Objective To prepare anti-idiotype single chain variable region antibody scFv (anti-IdscFv) of hepatitis C virus (HCV) nonstructural protein NS5 (NS5), which lays a foundation for the development of anti-IdscFv vaccine against HCV NS5. Methods The phage display technique was used to coat the HCV NS5 monoclonal antibody on the Nunc plate. After five rounds of “elution-elution amplification” from the phage library of single-chain variable region antibody, 80 clones were selected at random. ELISA, cross-reaction and competition inhibition experiments were carried out, and immunological detection was carried out to obtain anti-IdscFv positive clones with strong binding activity to HCVNS5 monoclonal antibody, and the coding sequence of HCVNS5 specific anti-IdscFv Sequence analysis. Results The screened HCV NS5 anti-IdscFv fragment consisted of 786 bp with the biological activity and specificity of binding to HCVNS5 monoclonal antibody. Conclusion The anti-IdscFv of HCVNS5 can be successfully obtained using the phage antibody library technology. The experimental results for the prevention and treatment of hepatitis C with anti-IdscFv create the conditions for the study.