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Atomic force microscopy

Introduction


The development of microscopes is important for a better understanding of biological processes. Visualization techniques are mainly based on optical imaging utilizing electromagnetic waves. The atomic force microscope (AFM), or scanning force microscope (SFM) was invented in 1986 by Binnig, Quate and Gerber. The AFM utilises a sharp probe moving over the surface of a sample in a raster scan.

In the case of the AFM, the probe is a tip on the end of a cantilever (length of about 200 Ám) with a low spring constant (of the order of 1 Newton/m) which bends in response to the force between the tip and the sample. Most AFMs employ an optical lever technique: as the cantilever flexes, light from a laser is reflected onto a split photo-diode. By measuring the difference signal, changes in the bending of the cantilever can be measured. The movement of the tip or sample is performed by a precise positioning device made from piezo-electric ceramics. Thus, in contrast to optical microscopes, AFMs measure surfaces in all three dimensions, with x- and y-resolutions in a typical range of 20 ┼ (under special conditions it could be better than 1 ┼) and a z-resolution in a typical range of better than 1 ┼.

Two AFM-modes can be distinguished: In the "constant force mode" (also called "height mode") the tip is in contact with the probe and measures the three-dimensional surface. The user could modify (consciously or unconsciously) the structure of the probe. The second mode is called "dynamic mode". Approaching the sample with an oscillating cantilever results in tapping of the sample by the tip. When the tip encounters elevation changes of the surface, the amplitude changes in consequence of interactions on the atomic scale (e.g. van der Waals Forces). In this case, the feedback system responds to keep the amplitude of the cantilever on a constant level.
In the past years AFM has been increasingly used to study biological samples, e.g. cell-surface morphology, proteins, and DNA.
Another possible application is the detection of ultralow forces. This has been used to observe receptor-ligand interactions on single molecule level: the ligand is bound to an AFM tip and receptors to a substrate or vice versa. Tip and substrate have to be brought into contact and the force required for receptor-ligand dissociation is measured. This application can be used for e.g. antibody-antigen recognition.