A novel electrochemical micromethod for the investigation of the interactions between DNA and non-electroactive species is described. The method was developed using the system of double-stranded DNA (dsDNA) modified gold electrodes (dsDNA/Au), a synthesized water-soluble C60 derivative as a model, and [Co(phen)3]3+/2+ (phen=1, 10-phenanthroline) as an electroactive indicator. Electrochemical studies with dsDNA-modified gold electrodes suggest that the C60 derivative can interact strongly with dsDNA, with binding sites of the major groove of the double helix and phosphate backbone of dsDNA, a binding constant of (1.6

The interactions of benzyl viologen (BV) with single-and double-stranded calf-thymus DNA immobilized onto gold electrodes have been studied by electrochemical methods. Benzyl viologen interacts electrostatically with both doublestranded (ds) and single-stranded (ss) DNA, and the strength of the interactions is dependent on ionic strength (μ). The dicationic form (BV2+) binds to dsDNA 9 times more strongly than the singly reduced form, BV·+, in a pH 7.4 Tris-HCl buffer solution at μ=8.4mM. BV2+ binds to ssDNA 5 times more strongly than the BV·+ form. From measurements atμ=8.4mM, a binding constant (K2+) of 2.0 ((0.2)×104M-1 and a binding site size (s) of 1 base pair were obtained, respectively, for dsDNA. For ssDNA, at the same ionic strength, the values obtained for K and s were 3.6 ((0.4) ×104M-1 and 2 nucleotides, respectively. The amount of BV bound, whether to dsDNA or ssDNA, decreased with increasing ionic strength. Whereas the binding rate of BV to both dsDNA and ssDNA immobilized onto gold electrodes is relatively low, once immobilized, it dissociates rapidly away from the electrode surface. The electron-transfer rate constant for BV is moderately fast at both dsDNA- and ssDNA-modified gold electrodes. The application of benzyl viologen as an electroactive indicator capable of differentiating between surface-immobilized single- and double-stranded DNA in denaturation/regeneration cycles has been explored.

The immobilization of thiol-derivatized DNA on a Au (111) single crystal surface by self-assembly has been investigated by electrochemical scanning tunneling microscopy (EC-STM). Continuous potential-dependent orientation changes of double-stranded oligodeoxynucleotides (ODN) have been observed in a certain potential range from 200 to 600mV (versus SCE). It is suggested that the DNA duplexes stand straight on the gold surface at potentials negative of the potential of zero charge (pzc) and then lay down on the surface when the potential shifts positively. These results are in agreement with the expectation based on the Coulombic interaction consideration between negatively charged DNA helices and gold surface. As the applied potential shifts positively, the surface charge changes from negative to positive, that is, the Coulombic force between negatively charged DNA helices and gold surfaces changes from repulsion to attraction. However, for the single-stranded oligodeoxynucleotides, no distinct changes in the surface structure were observed with the applied potential.

Gold nanoparticle/streptavidin conjugates covered with 6-ferrocenylhexanethiol were attached onto a biotinylated DNA detection probe of a sandwich DNA complex. Due to the elasticity of the DNA strands, the ferrocene caps on gold nanoparticle/streptavidin conjugates are positioned in close proximity to the underlying electrode modified with a mixed DNA capture probe/hexanethiol self-assembled monolayer and can undergo reversible electron-transfer reactions. A detection level, down to 2.0pM (10amol for the 5μL of sample needed) for oligodeoxynucleotide samples was obtained. The amplification of the voltammetric signals was attributed to the attachment of a large number of redox (ferrocene) markers per DNA duplex formed. The ferrocene oxidation current increased with the target concentration and began to level off at a target concentration of 10nM. An Excellent linearity was found within the range between 6.9 and 150.0pM and reasonable relative standard deviations (between 3.0 and 13.0%) were obtained. The amenability of this method to the analyses of polynucleotides (i.e., PCR products of the pre-S gene of hepatitis B virus in serum samples) was also demonstrated. The method is shown to be simple, selective, reproducible, and costeffective and does not require labeling of the DNA targets.

Gold electrodes were modified with DNA by adsorption. The DNA-modified electrodes were electrochemically characterized with Co(bpy)3+, a electroactive DNA-binding complex, as an indicator. It is interesting that the pair of redox peaks of Co(bpy)3+ split into two pairs at dsDNA-modified gold electrodes. One pair of peaks shifts negative, and the peak current increases notably; another pair of peaks shifts positive. These suggest that dsDNA has been immobilized onto gold electrode surfaces and the layer of dsDNA on the surfaces can bind Co(bpy)3+ in two different ways. Gold electrodes can be modified also with ssDNA by adsorption but only one pair of peaks of Co(bpy)3+ appears at ssDNA-modified gold electrodes. The amount of Co(bpy)3+ enriched by the layers of dsDNA or ssDNA adsorbed at gold electrodes was estimated from the peak charge of Co(bpy)3+ reduction at the electrodes obtained by CV. The stability of the DNA-modified electrodes was investigated. The DNA modification layer on gold surfaces is unstable to alkali and to heat, but stable to acid solutions and very stable in long stock in a dry. state. A comparison of modifications of gold, platinum and glassy carbon with DNA was carried out.