Ren Fangli, Wang Ruoke, Zhang Yanquan, Liu Chunxiao, Wang Yinyin, Hu Jim, Zhang Linqi, Chang Zhijie
1 State Key Laboratory of Biomembrane and Membrane Biotechnology, School of Medicine, School of Life Sciences, Tsinghua University , Beijing, China .
Monoclon Antib Immunodiagn Immunother. 2014 Dec;33(6):401-8. doi: 10.1089/mab.2014.0043.
CREPT (cell-cycle related and expression-elevated protein in tumor), a novel gene also called RPRD1B and C20ORF77, was recently identified to promote tumorigenesis through up-regulation of the expression of genes related to cell cycle. The previous study demonstrated that CREPT is highly expressed in a variety of tumors and enhances the expression of Cyclin D1 by promoting the formation of a chromatin loop. To study the correlation of CREPT expression with clinical factors in different tumors, we generated a monoclonal antibody (3E10) using purified recombinant human GST-CREPT protein as an antigen. In this study, we characterized the specificity of the monoclonal antibody and cloned the gene encoding the antibody for preparation of industrial production. Our results showed that the monoclonal antibody 3E10 was sensitive and specific to recognize human endogenous CREPT protein. We have mapped the epitope of the antibody and cloned the variable region sequence of the gene encoding the antibody. We confirmed that the cloned gene produced an equivalent antibody as that produced by the original hybridoma. This study provided a basis for large-scale production of the CREPT antibody, which will be useful for the study of the role of CREPT in different tumors.
CREPT（肿瘤中与细胞周期相关且表达上调的蛋白），一种也被称为RPRD1B和C20ORF77的新基因，最近被发现可通过上调与细胞周期相关基因的表达来促进肿瘤发生。先前的研究表明，CREPT在多种肿瘤中高表达，并通过促进染色质环的形成来增强细胞周期蛋白D1的表达。为了研究CREPT表达与不同肿瘤临床因素的相关性，我们以纯化的重组人GST-CREPT蛋白为抗原制备了单克隆抗体（3E10）。在本研究中，我们鉴定了该单克隆抗体的特异性，并克隆了编码该抗体的基因以用于工业化生产。我们的结果表明，单克隆抗体3E10对识别人类内源性CREPT蛋白敏感且特异。我们已定位了该抗体的表位，并克隆了编码该抗体的基因的可变区序列。我们证实，克隆基因产生的抗体与原始杂交瘤产生的抗体等效。本研究为大规模生产CREPT抗体提供了基础，这将有助于研究CREPT在不同肿瘤中的作用。
