A simple electrochemical immunosensor for sensitive detection of transgenic soybean protein CP4-EPSPS in seeds.

Soybean is the most produced crop in Argentina, and 99 % corresponds to genetically modified soybean. One of the main produced varieties is Roundup Ready® soybean (RR), which was modified to express the enzyme CP4 5-enolpyruvylshikimate 3-phosphate synthase (CP4 EPSPS), which confers resistance to glyphosate, the main herbicide worldwide used. The possible impact of genetically modified organisms (GMO) has generated public concerns, thus increasing interest in the development of GMOs detection devices. In this work, an electrochemical immunosensor for CP4 EPSPS detection in soybean seeds was obtained, by using a gold electrode modified with an anti-CP4 EPSPS polyclonal antibody produced in our laboratory. The presented immunosensor resulted in a simple, low-cost, fast, and reproducible device. Also, labeling and/or signal amplification system was not necessary, since the sensor showed high sensibility with a low detection limit (lower at 0,038 % RR soybean, 38 ng mL-1 CP4 EPSPS).

Polyclonal antibody production anti Pc_312-324 peptide. Its potential use in electrochemical immunosensors for transgenic soybean detection.

A new polyclonal antibody that recognizes the CP4 5-enolpyruvylshikimate-3-phosphate synthase (CP4-EPSPS), which provides resistance to glyphosate in soybean (Roundup Ready®, RR soybean), was produced. New Zealand rabbits were injected with a synthetic peptide (Pc_312-324, (PEP)) present in the soybean CP4-EPSPS protein. The anti-PEP antibodies production was evaluated by electrophoresis (SDS-PAGE) and an enzyme-linked immunosorbent assay (ELISA) was developed in order to study their specificity. The ELISA showed that the polyclonal antibody was specific to PEP. In addition, the anti- PEP was immobilized onto a gold disk electrode and the antigen-antibody interaction was evaluated using cyclic voltammetry (CV) and electrochemical impedance spectroscopy (EIS). Moreover, the EIS showed that the electron transfer resistance of the modified electrode increased after incubation with solutions containing CP4-EPSPS protein from RR transgenic soybean, while no changes were detected after incubation with no-RR soybean proteins. These results suggest that the CP4-EPSPS was immobilized onto the electrode, due to the specific interaction with the anti-PEP. These results show that this antigen-antibody interaction can be detected by electrochemical techniques, suggesting that the anti-PEP produced can be used in electrochemical immunosensors development to quantify transgenic soybean.