Measuring and Remediating Nonspecific Modifications of Gold Surfaces Using a Coupled in Situ Electrochemical Fluorescence Microscopic Methodology.

In surface-based biosensors, the nonspecific or undesired adsorption of the probe is an important characteristic that is typically difficult to measure and therefore to control or eliminate. A methodology for measuring and then minimizing or eliminating this problem on gold surfaces, readily applicable to many common surface modifications is presented. Combining electrochemical perturbation and fluorescence microscopy, we show that the potential at which the adsorbed species is removed can be used as an estimate of the strength of the adsorbate-surface interaction. This desorption potential can be easily measured through an increase in fluorescence intensity as the potential is manipulated. Furthermore, this method can be used to evaluate strategies for preventing or removing nonspecific adsorption. This is demonstrated for a wide variety of surface modifications, from strongly chemisorbed monolayers such as thiol self-assembled monolayers (SAMs) to physisorbed monolayers as well as for complex surface structures like peptide and DNA mixed-component SAMs. The use of a coadsorption strategy or small magnitude potential-step cycles was shown to significantly decrease the amount of nonspecifically or noncovalently bound probe, creating better defined surfaces.