CP4-EPSPS夹心ELISA配对单克隆抗体的研制和生物学特性分析

Preparation and Analysis on Biological Characteristics of Monoclonal Antibodies Against CP4-EPSPS for Sandwich ELISA

为获得能够用于夹心ELISA检测的配对抗CP4-EPSPS蛋白单克隆抗体,构建CP4-EPSPS原核表达载体并转化大肠杆菌Rossetta,获得高效表达。通过对可溶性蛋白纯化,获得了高纯度目的蛋白。以纯化后的CP4-EPSPS蛋白作为抗原免疫BALB/c小鼠,经过细胞融合和筛选获得2株抗CP4-EPSPS蛋白单克隆抗体(1A5和8A3)。经鉴定,这2株抗体可以有效地识别高温变性和天然CP4-EPSPS蛋白;叠加ELISA分析表明两株抗体识别的抗原表位不同,说明两株抗体能够用于夹心ELISA检测CP4-EPSPS蛋白。

In order to obtain specific monoclonal antibody against CP4-EPSPS protein for sandwich ELISA, the CP4-EPSPS gene was cloned into a bacterial expression vector and transformed into E.coli strain Rossetta. The target protein was expressed efficiently. After purification, the target protein was obtained. Purified CP4-EPSPS was used to immunize BALB/c mice and two monoclonal antibodies （ 1A5 and 8A3 ） were generated. These antibodies can recognize nature and denatured CP4-EPSPS. Overlay ELISA showed that they recognize different epitopes on CP4-EPSPS, so they can be used in sandwich ELISA to detect CP4-EPSPS.