W. Richard Bowen and Nidal Hilal 4

214 7. MICRO/NANOENgINEERINg ANd AFM FOR CELLULAR SENSINg
and appropriate physiological conditions (medium temperature and pH).
Although advances in cell immobilisation and AFM imaging modes now
permit soft, live cells to be imaged, the process remains far from trivial [74].
For living cell imaging, an environmental control chamber with suitable
medium is essential to keep cells alive and viable. When imaging
in contact mode, cantilevers with the lowest spring constants (0.003–
0.06 N m �1 ) operating with a small loading force (�1 nN) are preferred.
Following these basic considerations, it is possible to successfully image
live cells. Figure 7.9(a)–(f) show height and error images of live 3T3 cells
on a fibronectin-coated glass slide and a fibronectin-coated PDMS microgrooved
substrate. A well-spread cell morphology with fibrous cytoskeleton
structures can be seen clearly in Figure 7.9(a) in comparison to the
restrained morphology of the cell on a flat PDMS substrate (Figure 7.9(c)
and (d)). This can be further demonstrated by quantitative analysis of the
cell height and its projecting area. It should be noted that cells tend to
detach from the PDMS substrates during imaging. All of these observations
demonstrate that 3T3 fibroblasts have much weaker adhesion to
soft PDMS substrates. On microstructured PDMS structures, cells tend
to position their nucleus in the trough of the channel, close to one edge
and mostly attach filopodia to the upper surfaces (Figure 7.9(e) and (f)).
This gives rise to a preferred cell alignment along the channel length, as
discussed earlier. Thus, AFM imaging provides a quantitative and high
resolution approach to studying the behaviour of living cells.
7.3.2 Elasticity Measurement of living Cells
On a soft sample surface, an approaching tip with controlled force will
cause an indentation. Although this is often problematic for AFM imaging
of living cells, it is a way to measure and map the elastic properties
of the cell structure. For a force–distance measurement, the deflection of
the cantilever is plotted as function of the z-separation of the probe and
the sample. On a stiff sample (e.g. glass), the deflection is proportional
to the probe sample separation. However, on the soft sample, the movement
of the tip will be less than that of the sample, and the difference is
the indention of sample (Figure 7.10(a)).
In order to be able to extract material properties from an AFM force–
distance curve, we must interpret the deflection using an appropriate
model. As a starting point, it is reasonable to consider an AFM force–
distance curve as arising from a conical tip indenting a planar surface.
This was first considered by Hertz in 1882 [75] (equation (7.1)) and later
generalised by Sneddon [76].
F � �
2
2 E
⋅ ⋅
π
2
1 � v
( )
⋅ ( �)
tan
(7.1)