Biology Reference
In-Depth Information
11.2.1 The AFM
Being originally designed as an imaging tool to characterize the topography of
electrically nonconductive materials, the AFM was rapidly applied to measure
surface forces. Its application to the characterization of biological samples
was accomplished by improvement of the method to detect the cantilever
delection, which allowed the operation under luid conditions. The AFM was
thereafter extensively used to characterize nonspeciic forces in air and liquid
as well as the mechanical properties of samples.
A schematic diagram of an AFM is shown in
Fig. 11.2a
. The most important
part of the AFM is the cantilever and its tip, the latter being responsible
of making contact with the sample, and the former of applying pushing
and pulling forces. The cantilever tip is moved relative to the sample with
subnanometre resolution by means of piezoelectric elements. An optical
system, composed of a light source and a segmented photodiode, allows us
to monitor the delection, by focusing the laser beam on the backside of the
cantilever and detecting the relected light with the photodiode. For small
delections (
d
), the cantilever responds like a spring of constant
k
, and the
force (
. When working on living
cells, it is very helpful to visualize the cells using optical microscopy to enable
us to position the AFM tip on the precise location of the cell. The coupling
F
) is obtained following Hooke's law,
F
=
kd
(a)
(b)
Figure 11.2.
(a) Schematic diagram showing the major components of an atomic
force microscope coupled to an inverted optical microscope to allow observation of
biological samples. (b) Representative example of a force-distance curve (approach
in gray) showing a single-molecule rupture event during retraction (black line) of the
interaction between an integrin expressed on the surface of a monocytic cell and a
ligand immobilized on the tip. The diagrams represent the position of the cantilever
and substrate during the force curve cycle. The optical micrograph shows a THP-1 cell
immobilized on a poly-L-lysine-coated dish and an AFM cantilever (Biolever, Olympus)
during the force measurements (the cantilever width is ~30 μm).
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