Atomic force microscopy with carbon nanotube probe resolves the subunit organization of protein complexes

Abstract

Among many scanning probe microscopies, atomic force microscopy (AFM) is a useful technique to analyse the structure of biological materials because of its applicability to non-conductors in physiological conditions with high resolution. However, the resolution has been limited to an inherent property of the technique; tip effect associated with a large radius of the scanning probe. To overcome this problem, we developed a carbon nanotube probe by attaching a carbon nanotube to a conventional scanning probe under a well-controlled process. Because of the constant and small radius of the tip (2.5-10 nm) and the high aspect ratio (1:100) of the carbon nanotube, the lateral resolution has been much improved judging from the apparent widths of DNA and nucleosomes. The carbon nanotube probes also possessed a higher durability than the conventional probes. We further evaluated the quality of carbon nanotube probes by three parameters to find out the best condition for AFM imaging: the angle to the tip axis; the length; and the tight fixation to the conventional tip. These carbon nanotube probes, with high vertical resolution, enabled us to clearly visualize the subunit organization of multi-subunit proteins and to propose structural models for proliferating cell nuclear antigen and replication factor C. This success in the application of carbon nanotube probes provides the current AFM technology with an additional power for the analyses of the detailed structure of biological materials and the relationship between the structure and function of proteins.

Tip artifact in atomic force microscopy observations of InAs quantum dots grown in Stranski–Krastanow mode

“…In AFM observations, it is widely known that the tip artifact prevents the determination of actual sizes 4,5,6,7,8,9,10,11,12,13,14,15,16,17 ( Fig. 1).…”

The substructure of immunoglobulin G resolved to 25kDa using amplitude modulation AFM in air

“…Such attractive force regime imaging has been successful with commercially available silicon cantilevers and integral tips, under ambient conditions at room temperature. Other attempts at sub-molecular resolution of individual globular proteins have been achieved by building a cryo-AFM using contact-mode imaging [10] and by optimising the tip shape and diameter using singlewalled carbon nanotubes [11].…”

Nuclear architecture and chromatin dynamics revealed by atomic force microscopy in combination with biochemistry and cell biology

“…The instrumentation and application of AFM have been enormously fruitful due to the development of sharp cantilevers for molecular imaging [36][37][38], cantilever modification techniques for single-molecule force measurement [39][40][41][42][43], and fast-scanning devices for realtime imaging [44][45][46][47][48].…”