Biomedical Engineering Reference
In-Depth Information
to integrate with optical techniques which are very important for many experiments in the
life sciences, such as epifluorescence microscopy or confocal microscopy. Combining
these advantages with the possibilities that AFM offers to carry out other experiments,
such as mechanical probing, molecular interaction measurement by force spectroscopy,
etc. means that AFM is an extremely important tool in many biological areas.
For these reasons, the number of AFM applications in enormous. The sections below
include only a few selected areas where AFM has proven particularly useful, but more
applications in the life sciences have been covered elsewhere [577-579].
7.3.1 Biomolecule imaging
Biomolecules form the basis of life, and understanding the structure, function and inter-
actions of biomolecules has been the key to the incredible progress in the life sciences, and
medicine in particular, over the last 50 years. As a technique with sub-molecular resolution
and the ability to image soft samples in water, AFM is very appropriate for the study of the
huge range of natural biomolecules. The four major classes of biomolecules are carbo-
hydrates, proteins, nucleic acids and lipids.
Of these, probably the least-well studied by AFM is the class of carbohydrates, although
even here, a number of different systems have been studied. These include self-assembled
monolayers of glycoconjugates [9, 580], glycosylated particles [280, 581], polysacchar-
ides [582, 583] and force spectroscopy of carbohydrate interactions [144, 584].
On the other hand, proteins have been extensively studied by AFM, not just by imaging
but also by other measurements, such as electrical and mechanical measurements [320,
567, 585-587]. Due to their importance in disease and biological processes, proteins are
one of the most widely-studied classes of molecules, and literature searching reveals
thousands of studies of proteins using AFM. A few representative examples will be
given here.
AFM is a particularly suitable technique for protein studies, due to the coupling of high
resolution with the ability to study samples under physiological conditions, which is
necessary because protein structure can be highly sensitive to the nature of the proteins'
environment. However, despite the incredible resolution achievable on flat atomically
well-defined surfaces, AFM of single proteins in physiological-mimicking conditions
often gives rather low resolution, only revealing sub-molecular features for very large
proteins or multi-domain protein complexes. This is due to the soft, yet tightly packed and
globular nature of most protein structures, meaning an AFM image of the outside
topography shows few structural details. Indeed, it is extremely challenging to obtain
angstrom-level resolution of native proteins under such conditions by
any
technique
because of the fact that the molecules are under constant movement.
However one area where AFM has been used to provide great details is in protein
complexes. Such assemblies are often studied by TEM or crystallographic techniques,
which suffer in that they study the complexes under of non-realistic conditions, giving
AFM an obvious advantage in the fidelity of the data to biological systems [301]. One
such protein complex that has been quite widely studied by AFM is the GroEL/GroES
chaperonin complex. This complex is of interest because of its role in assisting the
protein folding process [588, 589], and has been quite widely studied by both contact and
oscillating AFM modes in buffer solutions [99, 590-592]. Sub-molecular resolutions,
