Biomedical Engineering Reference
In-Depth Information
Figure 8.16
Principle of the atomic force microscope.
is kept at a fixed value and the sample is dynamically moved up and down to keep
this force constant during the scanning. This way, the two hurdles just mentioned
above vanish: on the one hand, the force applied on the sample is constant so the
perturbation to the sample is the same everywhere and, on the other hand, a con-
stant force means a constant position of the cantilever extremity with respect to
the laser. There is thus no more need for a calibration of the detector since the read
value remains constant. As the sample vertical position is controlled by a piezo-
electric actuator easily well calibrated in distance, the topography of the sample is
directly given by these vertical displacements. The lateral resolution is limited by
the exact shape of the tip and is hard to define in the classical way as the influence
of this parameter depends on the size and shape of the imaged objects. It is certainly
one of the strengths of the AFM to be able to image samples at high resolution in a
liquid and in particular in a buffer solution. On this respect, it is a major advantage
compared to other techniques such as electron microscopy. Because it is a simpler
technique to use, AFM is often used also on dried samples.
It is clear from Figure 8.17 that AFM imaging is resolutive enough to access
some protein structures. Although the resolution is somewhat poor compared to
traditional diffraction techniques and very partial, as only the surface can be im-
aged, there is no need for crystallization since single molecules or complex can be
imaged, an invaluable advantage in the case of membrane proteins. Furthermore,
these images are taken in buffer solutions, sometimes directly on cells, thus in con-
ditions very close to the functional environments of the proteins.
In biology, the samples one is interested in can have two characteristics that
make them difficult to image with an AFM: they can be very soft and they may
need to stay hydrated. Most of the time, it is both. The interaction of the stylus
