He Xiang, Yang Jin-ju, Liu Ying, Liu Xiao-lan, Chen Yong, Gao Jian-en, Sun Qi-hong
Department of Immunology, Beijing Institute of Radiation Medicine, Beijing 100850, China.
Xi Bao Yu Fen Zi Mian Yi Xue Za Zhi. 2007 Sep;23(9):847-9.
To identify the specificity of mAb DBD02, we developed and optimized a new method by biopanning of T7 Select human liver cDNA phage display library.
After two rounds of biopanning, eluted phages were subjected to Dot blot using mAb DBD02 as primary antibody. The positive PFUs (plaque forming unit) which recognized by DBD02 were further confirmed by Western blot, PCR and sequencing.
The antigen recognized by DBD02 was identified as alcohol dehydrogenase. And the epitope for mAb DBD02 was further mapped within a peptide of 22 amino acids (QDYKKPIQEVLKEMTDG-GVDFS).
Our data indicate that biopanning of T7 phage display library by mAbs is time, cost, and labor-saving and specific tool for protein antigen identification.
为鉴定单克隆抗体DBD02的特异性，我们通过对T7 Select人肝脏cDNA噬菌体展示文库进行生物淘选，开发并优化了一种新方法。
经过两轮生物淘选后，使用单克隆抗体DBD02作为一抗，对洗脱的噬菌体进行斑点印迹。通过蛋白质印迹、聚合酶链反应和测序进一步确认被DBD02识别的阳性噬菌斑形成单位（PFU）。
DBD02识别的抗原被鉴定为乙醇脱氢酶。单克隆抗体DBD02的表位进一步定位在一个22个氨基酸的肽段（QDYKKPIQEVLKEMTDG-GVDFS）内。
我们的数据表明，利用单克隆抗体对T7噬菌体展示文库进行生物淘选是一种省时、省钱、省力且用于蛋白质抗原鉴定的特异性工具。
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