Single-Molecule Force Spectroscopy of DNA-Based Reversible Polymer Bridges: Surface Robustness and Homogeneity.

Single-molecule force spectroscopy, as implemented in an atomic force microscope, provides a rarely-used method by which to monitor dynamic processes that occur near surfaces. Here, a methodology is presented and characterized that facilitates the study of polymer bridging across nanometer-sized gaps. The model system employed is that of DNA-based reversible polymers, and an automated procedure is introduced that allows the AFM tip-surface contact point to be automatically determined, and the distance d between opposing surfaces to be actively controlled. Using this methodology, the importance of several experimental parameters was systematically studied, e.g. the frequency of repeated tip/surface contacts, the area of the substrate surface sampled by the AFM, and the use of multiple AFM tips and substrates. Experiments revealed the surfaces to be robust throughout pulling experiments, so that multiple touches and pulls could be carried out on a single spot with no measurable affect on the results. Differences in observed bridging probabilities were observed, both on different spots on the same surface and, more dramatically, from one day to another. Data normalization via a reference measurement allows data from multiple days to be directly compared.