Electrochemical investigation of DNA-modified surfaces: From quantitation methods to experimental conditions

We have thoroughly compared cyclic voltammetry and chronocoulometry, the two commonly used electrochemical techniques in quantitative determination of DNA surface density on gold electrodes. The electrostatic binding of redox cations, such as [Ru(NH3)6]3+, to the anionic DNA phosphate backbone allows the determination of the number of phosphate residues and thereby the surface density of the oligonucleotide strands. The results obtained by the two methods agreed well with each other for thiol-terminated DNA immobilized on gold electrodes via Au–S linkages and for unmodified DNA physically adsorbed on the surface. An improved procedure has been developed for the preparation of DNA-modified gold chips with a relatively high surface coverage. The density of thiol-terminated single-stranded DNA probes was determined to be (2.2 ± 0.3) × 1013 molecules/cm2 upon reaching adsorption equilibrium, which was in good agreement with that obtained by phosphor imaging of 32P-radiolabeled DNA.