中文摘要：

目的制备抗淋巴样增强因子-1（LEF-1）羧基端86个氨基酸（Lct86）特异性单克隆抗体，并初步明确其表位，为进一步研究LEF-1 C端特异序列的功能奠定基础。方法从Jurkat细胞株中扩增LEF-1 C端特异序列cDNA，并克隆入PQE40、pET32a原核表达载体；分别转化入M15、B121（DE3）感受态菌，IFIG诱导表达；纯化PQE40-Lct86融合蛋白作为免疫原免疫Balb／c小鼠制备单克隆抗体，以pE332a-Lct86融合蛋白鉴定单克隆抗体特异性。LEF-1 C端特异序列不同长度的基因片段定向克隆入PQE40，鉴定各片段的表达，利用Western blotting初步鉴定单克隆抗体的表位。结果成功构建了PQE40-Lct86、pE332a-Lct86重组原核表达载体；纯化获得PQE40-Lct86融合蛋白；得到一株特异性的抗LEF-1 C端单克隆抗体TMAb1，证实其表位位于437aa～454aa。结论成功制备了一株特异性的抗LEF-1 C端单克隆抗体TMAb1，并鉴定其抗原识别表位位于437aa～454aa，为进一步研究LEF-1 C端的功能奠定基础。

英文摘要：

Objective To prepare monoclonal antibody against LEF-1 carboxyl terminus 86 amino acids （Lct86）, and identify the antigenic epitope recognized by this monoclonal antibody. Methods LEF-1 carboxyl terminus cDNA was amplified from Jurkat cell line, and then cloned into expression vector pE-332a and PQE40. The recombinants were transformed into BI21 （DE3） and M15 respectively, and then induced with IPTG. Balb/c mice were immunized with purified PQF40-Lct86 and the specificity of monoclonal antibody was tested with pE332a-Lct86. Furthermore, different length gene fragments of LEF-1 carboxyl terminus were subcloned in-frame into PQE40. These construct plasmids were transformed into M15 and induced to expression, and then were used to identify antigenic epitope of LEF-1 carboxyl terminus recognized by the monoclonal antibody. Results PQF40-Lct86 and pE332a-Lct86 recombinant plasmids were constructed. After purification, we obtained purified PQE40-Lct86 and had one specific monoclonal antibody against LEF-1 carboxyl terminus（TMAb1 ）, which could recognize the antigenic epitope （437aa- 454aa）. Conclusion One specific monoclonal antibody （TMAb1） which can recognize antigenic epitope （437aa- 454aa） of the LEF-1 carboxyl terminus is successfully prepared and would be useful to explore the biological role of the LEF-1 carboxyl terminus.