The creation of gold surfaces modified by single- or double-stranded DNA self-assembled monolayers (SAMs) is shown to produce heterogeneous surface packing densities through the use of electrochemical studies coupled with fluorescence imaging. The modified surfaces created by direct adsorption of thiolate DNA [followed by passivation with mecaptohexanol (MCH)] resulted in regions covered by a monolayer of DNA SAM and other regions that were coated by large particles of DNA. The difference in fluorescence intensity measured from these regions was dramatic. More importantly, a regional variance in fluorescence intensity in response to electrochemical potential was observed: the large aggregates showing a significantly different modulation of fluorescence intensity than the monolayer-coated regions. Electrochemical desorption and detection of the fluorescently tagged DNA provided clear evidence of a complete surface modification. These studies have implications for biosensor/biochip development using DNA SAMs. A modification in the method used to produce the DNA SAMs resulted in a significantly different surface with much fewer aggregates and more significant electromodulation of the fluorescence intensity, though at much lower DNA surface density (ca. 1% of maximum theoretical coverage). This method for forming the modified surfaces has clear advantages over the currently accepted practice and emphasizes the importance of studying the nonaveraged nature of the sensor surface using in situ imaging tools like electrofluorescence microscopy.

The electrochemical properties of phospholipid monolayers of dioleoyl phosphatidylcholine (DOPC) spread from the gas/solution interface on mercury surfaces are quantitatively investigated in this paper. These layers display multiple states which interconvert through phase transitions characterized by two sharp capacitive peaks. Potential pulse techniques (chronocoulometry) were used to quantitatively investigate the properties of the DOPC monolayer on a mercury electrode. Charge density and the resulting film pressure due to DOPC spreading at the Hg/solution interface were determined. Results indicate that the lipid layers are displaced from the mercury surface at negative potentials in excess of −1.8 V. The potential of maximum film pressure or stability of the lipid monolayer and the shift in the potential of zero charge due to lipid transfer to the mercury surface were estimated as −0.4 and +0.435 V versus Ag/AgCl (saturated KCl), respectively. The similarity of the DOPC monolayer properties on mercury to the insoluble surfactant monolayer properties on single-crystal gold electrodes is noted. The spread DOPC layer and specifically the first phase transition was further characterized utilizing Tl+ and Cd2+ reduction. From potentials of −0.65 V to the potential coincident with the first phase transition, the permeability of the layer to these metal ions increases with an increase in the applied negative potential. The second phase transition represents a process involving the growth and coalescence of defects.

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The reductive desorption of a self-assembled monolayer (SAM) of a fluorescent thiol molecule (BodipyC10SH) from Au was characterized using electrochemistry and epi-fluorescence microscopy. Molecular luminescence is quenched near a metal surface, so fluorescence was only observed for molecules reductively desorbed and then separated from the electrode surface. Fluorescence imaging showed that reductive desorption was selective, with desorption occurring from different regions of the Au electrode depending on the extent of the negative potential excursion. When desorbed, the molecules were sufficiently mobile, diffusing away from the electrode surface, thereby preventing oxidative readsorption. At sufficiently negative desorption potentials, all of the thiol was desorbed from the electrode surface, resulting in fluorescence at the air/solution interface. The selective removal of the thiol monolayer from distinct regions was correlated to features on the electrode surface and was explained through potential-dependent interfacial energies. This in situ electrofluorescence microscopy technique may be useful in sensor development.