Apparent Dependence of Rupture Force on Loading Rate in Single‐Molecule Force Spectroscopy

Abstract

Force spectroscopy is becoming an important tool in physicochemical research and allows the extraction of kinetic parameters of dissociation that are not available to other techniques. The theory of single-molecule force spectroscopy has received considerable attention over the last several years and continues to be developed. Atomic force microscopy (AFM) in the force measurement mode is one of the most common methods to perform force spectroscopy measurements. In these experiments, rupture forces between interacting molecules are measured as a function of the AFM probe velocity. The resulting dependence of most probable rupture force on loading rate is then used to extract kinetic information. Although this technique is becoming widespread, we find that some important experimental aspects require more thorough consideration. One of these seemingly simple aspects is a methodology of extracting rupture force from a force curve. Herein, we discuss how this methodology might have a significant impact on the interpretation of force spectroscopy measurements. Consider the force-separation curve with a rupture event shown in Figure 1. A part of the force curve corresponding to the stretching of the polymeric tether prior to the rupture without added noise is shown with a solid line. With noise this dependence might show a considerable amount of scatter. Therefore, if the rupture force is obtained as a difference between the most outstanding data points in the vicinity of the rupture event, then the root-mean-square (RMS) amplitude of the noise will be added to the rupture force, as illustrated in the figure. The RMS amplitude of noise that is added might depend on the pulling velocity and therefore affect kinetic parameters obtained in force spectroscopy measurements. The dependence of the noise amplitude Fn the on probe velocity v can be written as [Eq. (1)]: