W. Richard Bowen and Nidal Hilal 4

for in situ monitoring of changes in intracellular signalling in combination
with cellular physical variations as a result of the presence of drugs. These
are most readily revealed by optical fluorescence assays. Consequently, in
the last few years, instrument manufacturers have developed a number
of combined AFM-optical microscope platforms that are now starting to
benefit studies in the fields of bioengineering, cell engineering and basic
cell biology. In particular, configurations involving confocal microscopy
[66] and total internal reflection fluorescence microscopy [67] have already
demonstrated their power in in situ living cell studies.
Since the invention of the AFM in 1986, we have witnessed significant
growth of the employment of AFM to probe biological systems. In the
next section we focus on recent developments associated with intact cell
measurement.
7.3 AFM In CEll MEASurEMEnt
The unique advantages of AFM in cell measurement lie in its capability
of being able to simultaneously (1) image cell topology under nearphysiological
conditions, (2) measure mechanical properties of living
cells, and (3) monitor functional cellular components and intracellular
processes in conjunction with optical microscopy. This section will demonstrate
the great potential of AFM for investigating the interaction of
cells with their environment.
7.3.1 AFM Imaging of Cells
7.3 AFM IN CELL MEASUREMENT 211
In the early days of AFM imaging of cells, the main restriction was
the limited scan size of the instruments [68], due to cells being relatively
large structures. As soon as the first instruments with scan ranges of several
micrometres were developed, they were deployed to image cells [69].
Today, modern bio-AFM instruments integrate an AFM platform onto the
stage of a conventional inverted optical microscope, thus enabling easy
positioning of AFM tip over a particular region of cells (Figure 7.7(a)).
This development has been very useful in identifying regions of interest
in cell morphology and where cells react to the microstructured substrate
(Figure 7.7(b)). In addition, the optical imaging also permits simultaneous
monitoring of the lateral morphology of cells and/or intracellular
signalling during the investigation by AFM [70]. Through a ‘direct overlay’
technique, it is possible to integrate AFM with optical images and
use the optical image to guide AFM operation. This enables correlation of
the biophysical and biochemical functionalities of a cell. In reality, the different
operating principles of AFM and optical microscopy can result in