Electrochemical sensor for detection of unmodified nucleic acids

We have developed a nucleic acid (NA) sensor based on mediated electrochemical oxidation of guanine residues. In this method, oligonucleotide probes are bound to a tin-doped indium oxide (ITO) electrode through a self-assembled phosphonate monolayer. The end carboxyl moiety of the monolayer is activated with carbodiimide and reacted with the amine group of a C6 alkyl linker which has been added to the 5′-end of the oligonucleotide probe. Upon hybridization of the complementary target NA, the hybrid is detected using a redox-active mediator, tris(2,2′-bipyridyl) ruthenium(II). We speculate that the monolayer does not impede electron-transfer since it contains many defect sites when assembled on a polycrystalline ITO surface. These defect sites are accessible to the mediator, but not to NA or proteins. The electrocatalytic current was a linear function of the amount of guanine bound at the electrode surface, with a detection limit of 120 amoles of guanine cm−2 at 0.28 cm2 ITO electrodes.