Selection of Highly Specific Aptamers by Graphene Oxide-SELEX to Ultrasensitive Label-Free Impedimetric Biosensor Development for Glyphosate Detection

The extensive use of the herbicide glyphosate has raised several health concerns due to the known toxicity of this chemical. The existing methods for glyphosate detection need better and more rapid accessibility for quick detection in an efficient manner. Here, we report an ultrasensitive aptamer-based electrochemical detection method for glyphosate. A high affinity single-stranded DNA (ssDNA) that binds specifically to glyphosate (GlyP) was selected and identified in vitro by the method of systemic evolution of ligands by exponential enrichment (SELEX) assisted by graphene oxide (GO), GO-SELEX. After eight GO-SELEX cycles, the ssDNA was sequenced, and the obtained aptamers were subjected to binding assays to evaluate their affinity and specificity to GlyP. The inherent GlyP–aptamer interaction was investigated by circular dichroism spectroscopy (CD). The selected sequence, denominated GlyP_1, exhibited a low dissociation constant (Kd) of 30.73 ± 1.25 nM and excellent selectivity for glyphosate. Prior to gold electrode modification, GlyP_1 was functionalized by the disulfide label and used as a recognition element for glyphosate detection. The surface modification steps for the gold electrodes to develop the aptasensor were followed up by cyclic voltammetry (CV) and electrochemical impedance spectroscopy (EIS) in the presence of a redox probe, [Fe(CN)6]3–/4–. The resulting aptasensor demonstrated an operation range from 1.16 pM to 400 pM, with a limit of detection (LOD) of 1.16 pM. Thus, in this work we report an effective sensing method that could provide a potential alternative to traditional analyses techniques for quick and easy monitoring of glyphosate in food and water.