Evaluation of the formation of a junctional DNA nanostructure through annealing curve analysis.
The accessibility and binding affinity of DNA are two key parameters affecting the hybridization efficiency in surface-based biosensor technologies. Better accessibility will result in a higher hybridization efficiency. Often, mixed ssDNA and mercaptohexanol monolayers are used to increase the hybridization efficiency and accessibility of surface-bound oligonucleotides to complementary target DNA. Here, no mercaptohexanol monolayer was used. We demonstrate by differential microcantilever deflection measurements at different pH that the hybridization efficiency peaks between pH 7.5 and 8.5. At low pH 4.5, hydration and electrostatic forces led to tensile surface stress, implying the reduced accessibility of the bound ssDNA probe for hybridization. In contrast, at high pH 8.5, the steric interaction between neighboring ssDNA strands was decreased by higher electrostatic repulsive forces, bending the microcantilever away from the gold surface to provide more space for the target DNA. Cantilever deflection scales with pH-dependent surface hybridization efficiency because of high target DNA accessibility. Hence, by changing the pH, the hybridization efficiency is adjusted.