Facilitating Nanomechanical Measurements on Physisorbed Biopolymers with Automated On-the-Fly Monitoring of Single-Molecule Force Curves

摘要

Single molecule force spectroscopy has recently given access to an unprecedented level of information regarding the stress response of a host of biopolymers at the single chain level. However, particularly for polysaccharides, where the intrinsic chemistry of the biopolymer complicates the design of chain-end-specific handles, the majority of studies are still carried out with biopolymers physisorbed to the substrates between which they are stretched. This means that the nature of recorded force-curve data is somewhat unpredictable from one scan to the next, making measurements tedious and time-consuming. Herein a methodology is described for use with single polymer force-spectroscopy that ameliorates this experimental inefficiency. The study describes the set-up of both an electronics bench and computational routines that enable accurate and automated measurements to be made despite low-level analog voltages and noisy signals. The length, quality and maximum force of any stretches detected are monitored in real-time and compared with user-defined thresholds, thus filtering the data set on-the-fly. This methodology for the automatic detection of single molecule stretching can be used both as an important time-saving tool in analysis, where many thousands of stretch attempts may be made in a single experiment, and eliminates measurement bias related to subjective choices made by the user. In addition, this tool enables the simple generation of an ensemble of measurements of the properties of studied molecular chains.