Biology Reference
In-Depth Information
and precise control of the force applied to the cantilever can be maintained.
This combination facilitates the evaluation of physical characteristics of
microbes. Third, rather than yielding the composite, statistical average of cell
populations, as is the case with many biochemical assays, the behaviour of
single cells can be monitored.
Despite the potential of AFM in microbiology, there are several limitations
that must be considered. For example, the time required to record an image
allows for the study of gross events such as cell division or membrane
degradation from an antibiotic but precludes the evaluation of biological
reactions and events that happen in just fractions of a second. Additionally, the
AFM is a topographical tool and is restricted to imaging surfaces. Therefore,
it cannot be used to look inside cells as with optical and transmission
electron microscopes. Other practical considerations are the limitation on
the maximum scan size (roughly 100
s
100 μm) and the restricted movement
of the cantilever in the
Z
(or height) direction. In most commercial AFMs,
the
range is restricted to roughly 10 μm such that the height of cells to
be imaged must be seriously considered. Nevertheless, AFM can provide
structural-functional information at nanometre resolution and do so in
physiologically relevant environments. Further, instrumentation for scanning
probe microscopy continues to advance. Systems for high-speed imaging are
becoming available, 1-3 and techniques for looking inside the cells are being
demonstrated.
Z
The ability to combine AFM with other imaging modalities is
likely to have an even greater impact on microbiological studies.
AFM studies of intact microbial cells started to appear in the literature in
the 1990s. For example, AFM studies of
4
examined
budding scars after cell division and detailed changes related to cell growth
processes. 5,6 Also, the irst AFM studies of bacterial bioilms appeared. 7 In
the late 1990s, AFM studies of intact fungal spores described clear changes
in spore surfaces upon germination, and studies of individual bacterial cells
were also described.
Saccharomyces cerevisiae
These early bacterial imaging studies examined
changes in bacterial morphology due to antimicrobial peptides exposure and
bacterial adhesion properties.
8-10
8,11
The majority of these early studies were carried out on dried samples
and took advantage of the resolving power of AFM. The lack of cell mounting
procedures presented an impediment for cell imaging studies. Subsequently,
several approaches to mounting microbial cells have been developed, and these
techniques are described later. Also highlighted are general considerations
for microbial imaging and a description of some of the various applications
of AFM to microbiology.
 
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