Biomedical Engineering Reference
In-Depth Information
Fig. 5
Use of the vital dye calcein and fl uorescence to track neuronal death and arborization
caused by addition of AbP to the buffer solution (
left
). Images taken at time 0 (control) (
A
,
C
,
E
,
G
)
and 45 min after addition (
B
,
D
,
F
,
H
). Simultaneously, AFM can be used to image the cytoskeletal
structure of cells to track the effects of AbP on the cells and to test various blocking agents for their
performance in blocking toxicity effects. The
right panels
show that AßP-induced changes in cel-
lular morphology were blocked by Zn
2+
and by the removal of extracellular calcium ions, but not by
tachykinin (physalaemin) and antioxidants. Images (
a
,
c
,
e
,
g
) are controls at time 0, images (
b
,
d
,
f
,
h
)
are taken 30 min after adding reagents. For details, see Lin et al. (
2001
) and Bhatia et al. (
2000
)
Furthermore, since the AFM operates under biological liquids, online additions
of reagents or exchange of buffer conditions can be performed during AFM imag-
ing. In this way, the blocking capability of, for instance, zinc ions and extracellular
calcium ion levels to prevent such cellular morphology changes can be shown in
real time (Fig.
5
, right panels). Amyloid-induced morphology changes in endothe-
lial cells were inhibited by zinc ions and by the removal of calcium ions from the
medium, but not by adding tachykinin (physalaemin), antioxidants, and cadmium
ions. Figure
5
(right) shows images at time 0 and 30 min after addition of the respec-
tive reagents. Another application of AFM imaging of whole live cells is the uptake
of small vesicles by endocytosis into the cell. For instance, vesicles loaded with
cisplatin anticancer agent were observed by both AFM and combined fl uorescence
to be taken up into the cell, after which live/dead assays showed the functional
activity of the drug (Ramachandran et al.
2006
). A multimodal AFM combined with
fl uorescence can give valuable additional information and conformation of pro-
cesses observed with the AFM. For example, the vital dye calcein can be used to
