与肺癌源性HSP70特异性结合单链融合抗体的筛选与序列分析

Identification and Sequence Analysis of Fused scFv Antibody Binding Specifically to HSP70

目的从人源性肺癌噬菌体单链抗体库中筛选与肺癌源性HSP70特异性结合的融合抗体。方法以肺癌源性的HSP70为抗原,对人源性肺癌噬菌体单链抗体库中经四轮筛选,单克隆噬菌体抗体与抗原HSP70结合经ELISA检测筛选出阳性菌株,再经PCR进一步鉴定,确定含有单链抗体基因的克隆并测序,测序结果通过GeneBank比对进行同源性分析。结果获得了1个特异性强的阳性噬菌体克隆。经DNA测序后,在Genebank中与人的免疫球蛋白库进行比对,确定为单链抗体片段。结论从人源性肺癌噬菌体单链抗体库中筛选到与HSP70特异性结合的具有功能活性的单链融合抗体,为进一步的HSP70可溶性抗体的制备以及以可溶性HSP70抗体为载体的药物导向抗肿瘤治疗奠定了基础。

Objective To identify the specific fused antibodies binding to HSP70 extracted from lung cancer by biopanning from a human scFv phage library of lung cancer. Methods HSP70 extracted from lung cancer was used as antigen to biopan human scFv phage library of lung cancer after four rounds of biopanning. High affinity and specificity positive monoclonical phage antibody was identified by ELISA. PCR was carried out for identification of the positive clones which combined with scFv gene sequence. DNA sequencing of positive clone was analyzed by Automated DNA sequencers, and their homologous sequences were searched in GeneBank. Results one positive clone was chosen and a single chain antibody with the functional variable region sequences was identified by searching GeneBank. Conclusion single chain fused antibody binding specially to HSP70 was selected from the phage library, and it lays the foundation for further research about preparation for the soluble antibody and drug target antitumor therapy.