人源化鼠抗人纤维蛋白单链抗体的体外成熟

In vitro maturation of a humanized mouse ScFv specific against human cross-linked fibrin

目的提高人源化鼠抗人纤维蛋白单链抗体的亲和力和特异性。方法以人源化鼠抗人纤维蛋白单链抗体为基础,通过合成部分随机化的突变引物和PCR等方法构建HCDR3和LCDR3的混合突变库,然后利用噬菌体表面呈现技术对抗体库进行筛选,并利用本实验室建立的单链抗体-碱性磷酸酶系统进行单克隆的鉴定。结果得到4株亲和力和特异性有明显改善的人源化抗体。结论构建CDR3突变库是抗体体外成熟的一个有效方法,该研究为研制低免疫原性的导向溶栓制剂奠定了基础。

Aim To increase the specificity and affi nity of a humanized mouse ScFv specific against human cross -linked fibrin.Methods Based on the sequence of the humanized ScFv,the partially randomized primers were synthesised and the Mixed HCDR3and LCDR3library was con-structed by using PCR method with partially randomized primers.The library was panned with coated antig en D -dimer,and the bioactivity of ScFv was analysed with ScFv -alkalin e phosphatase detection system which was established in our laboratory.Results Four different human-ized ScFv with better affinity and sp ecifity were identified.Conclusion Constructing the CDR3library is an e ffective way to increase the affinity and specifity of antibody.