交替洗脱法筛选高亲和力噬菌体肽

Selecting high affinity phage-peptide from phage display peptide libraryby alternating elution

以人胰岛素为靶蛋白从七肽展示库中筛选高亲和力噬菌体肽,在洗脱阶段采用酸性洗脱液和高浓度靶蛋白溶液进行4次交替洗脱,选择性回收高亲和力噬菌体肽。测定滴度计算回收率,ELISA法分别测定噬菌体洗脱液整体亲和力和噬菌体单克隆的结合特性并计算亲合率。洗脱步骤采用4次交替洗脱后,第二轮第4次噬菌体的回收率比第一轮增长了1 800倍,高亲和力噬菌体在洗脱液中所占比例也迅速提高,第二轮第4次洗脱液中达75％,在第三轮的各次洗脱液中几乎均达100％。建立了一种快速筛选高亲和力噬菌体肽的方法,改进后的筛选方法能使高亲和力噬菌体肽的筛选工作更为简便且效果显著。

To establish an elution strategy for selecting high affinity phage-peptide from phage display peptide library efficiently. Screening the 7-mer phage display peptide library to obtain high affinity phage-ligand for insulin. In elution step, the alternating-elution, eluting the bind phage by acid buffer and high concentration target solution alternatively, was applied. The advantage of this method was confirmed by phage recovery and the rate of high affinity clone. By using the alternating-elution, the recovery of the forth elution in the second round was 1800 times higher than the corresponding recovery in first round. The rate of high affinity phage in second round was 75%, and in third round was 100%. After improved, the new method can select the high affinity phage more simply and efficiently.