In conclusion, the FITC-Phalloidin/DAPI/Alexa Fluor
568 goat anti-rabbit combined stain allowed us to differentiate between cell
components including actin microfilaments such as stress fibres, lamellipodia
and filopodia, and also focal adhesions, whilst allowing the visualisation of
the nucleus. The fluorescence imaging of cells is important to cell biology as
it allows us to study them in greater detail to obtain a better understanding
of their structure and function, and their molecular interactions. In future,
we may consider using live cell imaging techniques to observe cellular
processes in real time, using time-lapse photography, which may provide more
insight into cellular dynamics.
During our experiment,
it was possible that fluorescence may have been lost due to photobleaching
which may have affected our results but to prevent this, we used a mounting
media with anti-fade protection.
Figure 2A shows the location of the focal adhesions
stained in red by Alexa Fluor 568 in relation to the nucleus, which is stained
in blue by DAPI. Upon merging this stain with FITC-phalloidin, the filamentous
actin network is visualised in green shown in figure 2B; It shows
co-localization of actin filaments and focal adhesion sites, and it becomes
apparent that there is a close relationship between the two. It is evident that
the actin filaments are anchored by, and terminate in, focal adhesion sites;
they link the actin filaments to the extracellular matrix. Therefore, the actin
filaments are crosslinked and bundled together to form stress fibres and, as
expected, there is a strong accumulation of focal adhesions associated with the
ends of the stress fibres. Stress fibres
can also be distinctly identified in fig. 3. The coordinated regulation of the
focal adhesion sites and the actin cytoskeleton is essential for cell
migration. Spike-like protrusions of the plasma membrane, called filopodia, can
also be observed in figure 2C (a magnified image of 1B). They appear in green
as they are actin-rich structures.
displaying the fixed HeLa cells we stained with the FITC-Phalloidin/DAPI/ Alexa
Fluor 568 goat anti-rabbit combined stain, allow us to observe various
components of the cytoskeleton
including actin microfilaments as illustrated in green by FITC-phalloidin in
fig.1. The actin filaments are clearly shown to be highly concentrated at the
periphery of the cell, as they underlie the plasma membrane. This association
of the actin cytoskeleton with the plasma membrane provides mechanical support
for the cell, contributing to its structure and function. Other components are
the lamellipodium which consists of a dense, ‘sheet-like’
network of peripheral actin filaments which can be also observed in figure 1.
Extending beyond this, filopodia are visible, indicated by the white arrow.
They contain tight parallel bundles of actin filaments and are seen to be
protruding beyond the leading edge of the cell, which are used in movement and
sensing a cell’s environment.