Interaction between nile blue and immobilized single- or double-stranded DNA and its application in electrochemical recognition

The interactions of nile blue (NB) with calf-thymus double-stranded DNA (dsDNA) in solution, single-stranded DNA (ssDNA) and dsDNA adsorbed on gold electrodes were studied electrochemically. The binding of NB with dsDNA showed a mechanism containing an electrostatic interaction with the binding constant of (5.9 ± 0.2) × 104 M−1 in 0.01 M pH 7.4 Tris-HCl. The presence of adsorbed dsDNA resulted in a great increase in the peak currents of NB in comparison with those obtained at a bare or ssDNA adsorbed gold electrode. The electron transfer rate constants of nile blue adsorbed on bare gold and bound to ss- and dsDNA/Au electrodes were 1.4 ± 0.1, 1.2 ± 0.02 and 2.9 ± 0.2 s−1, respectively. Nile blue showed an electrostatic action with adsorbed ssDNA and an action with adsorbed dsDNA by the mechanism containing both electrostatic and intercalative bindings. The Langmuir adsorption constants of NB at ssDNA/Au and dsDNA/Au electrodes were (1.6 ± 0.2) × 105 and (4.2 ± 0.6) × 105 M−1, respectively. The difference between interactions of NB with adsorbed ss- and dsDNA has been used for hybridization recognition of hepatitis B virus DNA fragment. The interactions between NB and covalently immobilized hepatitis B virus ssDNA fragment and its hybridization product showed the mechanisms similar to those of adsorbed calf-thymus DNA.