中文摘要：

为了深入研究抗猪流感病毒（SIV）单链抗体（scF v）的功能,以及抗体的放大生产和应用,在获得特异性抗体SIV scF v编码基因的基础上,建立该单链抗体的原核高效表达体系,并对表达产物进行复性纯化和活性测定。通过PCR从重组噬粒中扩增抗scF v基因,将其克隆到原核表达载体p ET28a（＋）中后,转化大肠杆菌BL21（DE3）感受态细胞并诱导表达。SDS-PAGE和Western blot分析表明重组scF v主要以包涵体形式表达,经过复性和纯化可获得可溶性的重组scF v,分子质量约为33 ku。间接ELISA和阻断ELISA表明,复性后的scF v与包被在酶标板上的SIV具有特异的结合反应,且具有浓度依赖性。重组scF v可以与其他2株SIV国内分离株特异性结合,提示重组scF v可能针对的是该病毒的保守区域。本研究成功获得了具有中和H1N1猪流感病毒活性的重组抗SIV scF v,为深入研究其功能以及进行大规模生产应用提供依据。

英文摘要：

In this study,a novel single-chain variable fragment（ scF v） antibody against swine influenza virus（ SIV） was prepared and characterized. The scF v gene was amplified from the recombinant phagemid by PCR. DNA fragment encoding anti-SIV scF v was cloned into the pE T-28a（ ＋） expression vector,and 6His-tagged scF v antibody was expressed as inclusion bodies in Escherichia coli BL21（ DE3）. After refolded by protein refolding kit,the solubilized His-tagged scF v proteins were purified using Ni2＋- Sepharose column chromatography.The SDS-PAGE and Western blotting analyses showed that the recombinant scF v antibody was about 33 ku. The scF v was further characterized by indirect ELISA and blocking ELISA,showing that the affinity and specificity of scF v were fully renatured and could bind SIV. Indirect ELISA also showed that the scF v had the specific binding activity with the other two strains of H1N1 influenza virus,suggesting that this antibody may bind to the conserved region of this virus. The findings lay the foundation for further application of scF v.