Single-molecule recognition imaging microscopy

Abstract

Atomic force microscopy is a powerful and widely used imaging technique that can visualize single molecules and follow processes at the single-molecule level both in air and in solution. For maximum usefulness in biological applications, atomic force microscopy needs to be able to identify specific types of molecules in an image, much as fluorescent tags do for optical microscopy. The results presented here demonstrate that the highly specific antibodyantigen interaction can be used to generate single-molecule maps of specific types of molecules in a compositionally complex sample while simultaneously carrying out high-resolution topographic imaging. Because it can identify specific components, the technique can be used to map composition over an image and to detect compositional changes occurring during a process.

A nanomechanical interface to rapid single-molecule interactions

“…It enhances binding probability by providing conformational flexibility for the ligand to reach the receptor and orient itself for binding. The flexible linker also provides a temporal delay between the tip-substrate interaction and the unbinding event 5,20 , which encodes the unbinding information onto the tip-sample force waveform. This information can be recovered by a computer that monitors the twisting motion of the T-shaped cantilever.…”

“…P recise recognition mechanisms of biological molecules offer a tremendous potential for the assembly [1][2][3] , imaging 4,5 and analysis of complex biological materials 6 . Single-molecule force spectroscopy (SMFS) experiments with the atomic force microscope (AFM) 7 have demonstrated the potential of intermolecular forces to be used for chemically specific nanoscale imaging 8 , manipulation 9 , directed assembly 10 , and biophysical studies of deformation and failure of biomolecules 11 .…”

“…Evidence for this is provided by the molecular recognition images obtained in the topography and recognition imaging mode (TREC) 5 . In this mode, molecular binding events produce detectable patterns in the oscillation amplitude.…”

Localization of the ergtoxin-1 receptors on the voltage sensing domain of hERG K+ channel by AFM recognition imaging

“…under/or near physiological conditions and without the need for rigorous sample preparation or labeling [46]. With the recent development of TREC (topography and recognition imaging), it becomes possible to quickly and easily obtain maps of binding sites with the lateral accuracy of several nanometers across variety surfaces, as it has been demonstrated on model receptor-ligand pairs [37,39] and remodeled chromatin structures [38]. In addition, TREC represents an exquisite method to quickly obtain the local submicron distribution of receptors on cellular surface with an unprecedented lateral resolution of 5 nm [40].…”

“…Combination of high-resolution atomic force microscope (AFM) topography imaging with single molecule force spectroscopy provides a unique possibility for the detection of specific molecular recognition events. With the recent development of simultaneous topography and recognition imaging (TREC) [37][38][39] it becomes possible to quickly obtain the local distribution of receptors on cell surface with unprecedented lateral resolution of 5 nm [40]. At present, no other microscopic techniques are able to provide directly both structural information of a biological sample and related functional information at such high spatial resolution.…”