汉坦病毒核衣壳蛋白mAb B11模拟结合肽的筛选和鉴定

Screening of mimotopes of HFRSV with mAb B11 by phage display peptide library

目的 应用噬菌体展示肽文库筛选可与肾综合征出血热病毒mAbB11特异性结合的短肽。方法 采用Protein A亲和纯化的单抗为筛选配基 ,对噬菌体展示的随机 12肽文库进行生物亲和淘选 ;夹心ELISA、竞争ELISA鉴定筛选克隆的结合特性 ,并进一步对阳性克隆进行序列测定和分析。结果 通过 4轮淘选 ,ELISA测定显示筛选到的噬菌体短肽多数能与B11单抗特异性地结合 (12 15 ) ;序列分析表明 8个阳性克隆氨基酸序列分为 4组 ;同源性分析显示核心序列可能为 (M F)HR(H T)X(H W )或 (R F)HX(H T)P(W M) (L Y)。

Objective To obtain the 12 mer phage displayed mimotopes of mAb B11 specific to hemorrhagic fever with renal syndrome virus. Methods A group specific mAb B11 against nucleocapsids of both HTNV and SEOV, which possesses no evident neutralizing titer but can enhance the protective activities of other mAbs, was used as selective molecule. A 12 mer phage peptide library was bio panned for 4 rounds, and the result was confirmed by sandwich ELISA, competition ELISA and DNA sequencing. Results After 4 rounds of effective screening, 12 positive clones screened out of 15, and amino acid sequences of 8 positive clones revealed 2 consensus motifs:(M/F)HR(H/T)X(H/W)or(R/F)HX(H/T)P(W/M)(L/Y). Conclusion The peptides displayed by phage can mimic the epitope of HFRSV antigen, which provides important basis for preparing more effective epitope based vaccine and specific diagnosis reagent.