The lateral charge transport in films comprising metal complexes bound to DNA monolayers on Si electrodes was measured by scanning electrochemical microscopy (SECM) in the steady-state feedback mode. Single-stranded (ss) DNA monolayers covalently bonded to the Si surface were prepared by automated solid-phase synthesis on hydroxyl-terminated n-alkyl monolayers at atomically flat Si(1 1 1)-H surfaces. Duplex (ds) DNA films were produced by hybridisation to the ssDNA monolayers. Our previous STM imaging investigations showed these dsDNA films exhibit considerable alignment, but the ssDNA films were observed by AFM to be rather disordered. Using solutions of Fe(CN)(6)(4-), IrCl63-, Ru(bipy)(3)(2+), Co(bipy)(2)(3+) and Ru(NH3)(6)(3+) as redox mediators, we compared the rate of charge transport in dsDNA films to that in ssDNA films. Lateral charge transport on the substrate - the source of the SECM feedback - depends on the self-exchange rate between the surface and solution as well as the rate of diffusion of charged, surface-bound species. Several mechanisms of lateral charge transfer (physical surface diffusion, electron hopping between DNA-bound redox species, charge injection into Si and DNA-mediated long-range electron transfer) were explored as possible explanations for the positive feedback observed in Ru(bipg)(3)(2+) and Ru(NH3)(6)(3+) solutions. While Ru(bipy)(3)(3+) was found to inject holes into the silicon valence band across the organic monolayer, the fast charge transport in Ru(NH3)(6)(3+)/dsDNA films (effective diffusion coefficient, (2.2 +/-0.3) x 10(-5) cm(2) s(-1)) was attributed to a combination of physical diffusion of the ruthenium centres on the surface and charge injection into the Si electrode. For analysis of the SECM feedback experiments with lateral charge transport coupled to electron transfer to dissolved mediators, an analytical approximation was developed and validated by comparison with the results of finite difference simulations. This model successfully accounts for the variation in the SECM feedback with the concentration of the mediator in the solution. (c) 2007 Elsevier B.V. All rights reserved.