Improved Thermal Stability and Homogeneity of Low Probe Density DNA SAMs Using Potential-Assisted Thiol-Exchange...

Methods for producing DNA SAM-based sensors with improved thermal stability and control over the homogeneity of low DNA probe density will enable advanced sensor development. The thermal stability of low-coverage DNA SAMs was studied for surfaces prepared using potential-assisted thiol exchange (E_dep) and compared to DNA SAMs prepared without control over the substrate potential (OCP_dep). Both surface preparation methods were studied using in situ fluorescence microscopy and electrochemistry with fluorophore or redox-modified DNA SAMs on a single-crystal gold bead electrode. Fluorescence microscopy showed that the influence of the underlying surface crystallography was important in both cases. The highest thermal stability was realized for square or rectangular surface atomic structure (e.g., surfaces from 110 to 100). The 111 and related surfaces were the least thermally stable. The low DNA coverage surfaces prepared by E_dep had better thermal stability and higher DNA probe mobility as compared to OCP_dep-prepared surfaces with the similar coverage. These results were correlated with methylene blue redox-tagged DNA probes, which directly measured the average DNA coverage. Both methods indicated that E_dep DNA SAMs were more uniformly distributed across the electrode surface, while the surfaces prepared via OCP_dep assembled into clusters with reduced mobility. The potential-assisted thiol-exchange approach to preparing low-coverage DNA SAMs was shown to quickly create modified surfaces that were consistent, had mobility characteristics which should yield superior DNA hybridization efficiencies, and having greater thermal stability which will translate into a longer shelf-life.