比较两种从噬菌体展示文库中淘选抗黄曲霉毒素B1单域重链抗体的方法,并为淘选针对小分子物质的单域重链抗体提供参考。采用固相淘选技术,以黄曲霉毒素B1人工抗原为靶分子,淘选天然单域重链抗体库,分别采用Gly-HCl法和AFB1竞争法洗脱结合噬菌体,对洗脱物进行滴度测定,以回收率和富集度为指标,对两种洗脱方法进行比较。采用phage-ELISA法鉴定噬菌体克隆,并对阳性克隆进行交叉反应分析以及序列测定。滴度测定结果显示,Gly-HCl法回收率比竞争法高5～10倍。分别随机挑取20个单克隆进行phage-ELISA鉴定,其中,Gly-HCl洗脱法未获得阳性克隆,AFB1竞争洗脱法得到8个阳性克隆。序列测定结果显示,这8个克隆均编码单域重链抗体。
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The objective of this study was tocompare the efficiency of two elute strategies in biopanning of single-domain heavy-chain antibody(HCAbs) phage display library against low molecular hapten,Aflatoxin B1(AFB1).For this,the artificial antigen AFB1-BSA and AFB1-OVA were used as ligand to screen the binding phages from an Alpaca non-immune library by antigen-coated solid phase selection.The binding phages were directly eluted with Gly-HCl or competitively eluted with AFB1 solution.The eluants were tittered separately.It was found that the recovery rate of Gly-HCl is 5 to 10 times higher than that of competition method.Twenty colons randomly picked from each eluant were characterized by phage-ELISA.Eight positive colons were isolated from the eluant by the competition method;however,no positive colons were isolated by Gly-HCl method.The sequencing data show that all the colons are encoding single-domain HCAbs.
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