Biology Reference
In-Depth Information
et al.
63
summarized the use of AFM-based technology in
combination with others in the study of tissue remodelling especially that
of elasticity mapping and identiication of proteolytic activity. Dazzi
Ludwig
et al.
64
proposed the use of an infrared spectro-microscopy method based on a
photo-thermal effect, which is able to localize single viruses, including when
they are located inside the bacteria they have infected. Kim
et al.
65
probed
the distribution of olfactory marker protein (OMP) on a tissue section
of vomeronasal organ using a glass bead that was coated with anti-OMP
antibodies. Francius
et al.
66
reported the result of pulling polysaccharides
on live cells in a similar manner to Gad
et al.
36
Verbelen
et al.
67
mapped
lipoarabinomannans on
et al.
68
measured the change
in the nanomechanical and topographical change in the form of mapping on
live bacterial cells,
Mycobacteria.
Kumar
Brevibacterium
, under stress from environmental heavy
metal ions. Xu
et al.
69
used higher resonance frequencies of the cantilever to
probe a wider range of mechanical and dynamical properties and identiied
the different components in biological materials. Plomp
et al.
70
used
immunolabelling technique on the cell surface and later used an AFM to
identify protein compositions. Carberry
et al.
71
used a high speed scanning
AFM to map a wide area with a high lateral resolution suitable for imaging
as well as component mapping.
12.7 MAPPING OF INTRACELLULAR mRNA
Besides mapping surface proteins, AFM can also probe intracellular molecules.
An interesting example is the localization of intracellular mRNA, as recently
performed by Osada
developed a method to
retrieve a small portion of intracellular mRNAs without compromising the
viability of the targeted cell.
et al
.
72
and Uehara
et al
.
73,74
They
Their AFM-based method was to push a bare
AFM probe into a live cell by applying a compressive force of 10-100 nN and,
after keeping it inside of the cell for a short time, pull it out for subsequent
ampliication and quantitation of retrieved mRNAs into cDNA through
quantitative RT-PCR and PCR procedures. The ampliied DNAs were analyzed
for their identiication by gel electrophoresis. When analyzed for mRNA of
the housekeeping protein of
72-74
B
-actin, the successful detection rate was about
97% (
170). This method can be used for the mapping of (temporary)
β-actin mRNA was high near the nucleus in the inactive state of the cell and,
after activation, mRNA concentration increased in a more peripheral region
in the direction of cell movement. A correlation between the AFM-based
method and the conventional
n
z
hybridization method using luorescently
labelled complementary DNA was established,
in situ
74,75
as shown in
Fig. 12.6
.
Search WWH ::
Custom Search