抗甲状腺刺激性免疫球蛋白单链抗体噬菌体抗体库的初建

PRELIMINARY CONSTRUCTION OF PHAGE ANTIBODY LIBRARY OF ScFv AGAINST TSI

目的 建立抗甲状腺刺激性免疫球蛋白单链抗体噬菌体抗体库,为进一步获得功能性抗体可变区基因,制备单链抗体和单链抗体-辣根过氧化物酶融合蛋白,用于甲状腺自身免疫性疾病的治疗和检测打下基础。方法 利用RT-PCR技术从分泌抗甲状腺刺激性免疫球蛋白的杂交瘤细胞中扩增出轻重链可变区基因,与噬粒表达载体p3SCMH连接,转化大肠杆菌,以甲状腺刺激性免疫球蛋白阳性患者血清IgG为抗原对表达的噬菌体抗体进行筛选。结果 经过三轮“吸附-洗脱-富集”筛选过程,洗脱下来的噬菌体呈现明显富集,收获率增加了40倍。结论 初步建立了抗甲状腺刺激性免疫球蛋白单链抗体的噬菌体抗体库。

The study is for the purpose of constructing phage antibody library of ScFv against TSI and thereafter, to expess ScFv antibody and ScFv - HRP fused protein, which have potential application in the therapy and diagnosis of autoimmune thyroid disease. Methods: RT - PCR reactions were applied to amplify immunoglobin VH and VL genes from ybridomas secreting specific McAb against human TSI. Phage antibody library was generated with the VH and VL genes using p3SCMH as vector. Three rounds of panning against coated TSI were completed. Results: After the third round selection, the enrichment of eluted phage particles was observed by improving the rate of obtained phages approximately 40 folds. Conclusion: The phage antibody library of ScFv against TSI was generated successful-ly.