Antigen Epitope Analysis and Establishment of Rapid ELISA for Phosphinthricin Acetyltransferase
LIANG Yu-Xin1,3, LI Zhong-Peng1, ZHANG Chun-Yu1, HOU Ji-Chao1,3, LI Xiao-Yu1,*, WANG Yong-Zhi1,2,*
1 Institute of Plant Protection, Jilin Academy of Agricultural Sciences, Gongzhuling 136100, China;
2 Institute of Plant Protection, Chinese Academy of Agricultural Sciences, Beijing 100193, China;
3 College of Plant Protection, Jilin Agricultural University, Changchun 130118, China
Abstract:With the continuous expansion of genetically modified soybean market, Chinese citizens have become more and more disputable about the food safety and environmental risks of genetically modified soybean (Glycine max). More and more countries require labeling of genetically modified food, and some countries also have requirements on the content of genetically modified ingredients. Therefore, in order to establish a rapid and effective detection method for Bar-transgenic soybean quantitative detection, 18 strains of phosphinothricin acetyltransferase (PAT) monoclonal antibody (MAb) with good specificity were prepared in mice (Mus musculus) immunized with purified PAT protein, and a panel of monoclonal antibodies (9F5 and 3G12) that can be used for sandwich enzyme linked immunosorbent assay (ELISA) were screened using the segmented expression peptides of PAT expressed in Escherichia coli, and a rapid quantitative double antibody sandwich ELSIA (DAS-ELISA) for PAT was established based on this panel of monoclonal antibodies. ELISA plate coated with the capture antibody 9F5, and co-incubated with detector antibody 3G12 (2 μg/mL) labeled with HRP for 30 minutes at 37 ℃, color rendering at room temperature and away from light for 15~20 minutes. The detection limit was 1.69 ng/mL and detection range of PAT protein was 0.5~8.0 μg/mL. The coefficient of variation in the plate and between plates was less than 10%. With high sensitivity and stability, the detection could be completed in 50 minutes. This method provides technical support for detection of Bar-transgenic soybean.
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