从噬菌体表面展示肽库中筛选边缘无浆体膜表面蛋白5单克隆抗体识别的抗原表位

目的从噬菌体表面展示肽库中筛选边缘无浆体膜表面蛋白5(Membrane surface protein5,MSP5)单克隆抗体识别的抗原表位。方法用MSP5单克隆抗体1D8作为靶标,对噬菌体展示随机12肽库进行筛选,通过ELISA和竞争抑制ELISA鉴定筛选克隆的结合特性,并提取阳性克隆的单链DNA,进行测序分析。结果从表面展示随机肽序列的噬菌体文库中筛选到与MSP5单抗1D8特异结合的噬菌体克隆,其一致序列为LING。竞争抑制试验表明,含特异序列的克隆能与MSP5重组蛋白抗原竞争。结论初步确定MSP5单克隆抗体1D8的抗原表位为线性表位,为进一步研究其在边缘无浆体诊断及新型疫苗研制中的作用奠定了基础。

Screening of An Antigen Epitope Recognized by Monoclonal Antibody against Membrane Surface Protein 5 of Anaplasma marginale from Phage Display Peptide Library

Objective To screen the antigen epitopes recognized by the monoclonal antibody(McAb)against membrane surface protein(MSP)5 of Anaplasma marginale from phage display peptide library.Methods Phage display random 12-peptide library was screened using McAb 1D8 of MSP5 as a target,and the screened clones were analyzed for binding ability by ELISA and competitive inhibition ELISA,based on which ssDNAs were extracted from the positive clones for sequencing.Results The phage clone showing specific binding to McAb 1D8 of MSP5,with a consensus sequence LING,was obtained.Competitive inhibition ELISA demonstrated competition of the phage clone with recombinant MSP5 antigen.Conclusion The antigen epitopes of McAb 1D8 against MSP5 of A.marginale was preliminarily proved as linear,which laid a foundation of further study on its role in the diagnosis of A.marginale infection and preparation of novel vaccine.