Biomedical Engineering Reference
In-Depth Information
(a)
(b)
(c)
1.0
Uncleared
1.0
75% Glycerol
25% Glycerol
25% Glycerol
0.8
50% Glycerol
75% Glycerol
0.8
0.6
0.6
0.4
0.4
0.2
0.2
0.0
0.0
0
100
200
300
400
0
50
100
150
200
250
300
350
400
Depth (µm)
Depth (µm)
FIgurE 8.3
3-D SHG imaging of cleared striated muscle tissue. (a)
x
-
z
projections of the SHG image stacks
for 25% (left) and 75% (right) glycerol treatments. Scale = 50 μm. (b) The normalized axial attenuation of the
forward SHG intensity for the control and three glycerol treatments. All the glycerol treatments result in greater
imaging depth relative to the control. (c) Comparison of the simulations and experimental data for the 75% and
25% glycerol treatment, considering a relative SHG creation efficiency of twofold greater for the former treatment.
(Reproduced from LaComb, R. et al. 2008.
J. Biomed. Opt
. 13:021108.)
in swelling of the muscle cells, where the higher concentrations result in thicker tissues for the same
exposure time. This is due to the hyperosmotic effect of the glycerol, which replaces the lower refrac-
tive index cytoplasm. Thus, swelling must be taken into account when directly comparing the data
because the physical thickness affects the subsequent measured SHG signal attenuation and direction-
ality. Owing to decreased scattering and absorption losses, the 50% and 75% glycerol treatments result
in similar axial attenuation profiles, suggesting that 50% glycerol already provides sufficient clearing
effect. The measured bulk optical parameters for the muscle are listed in Table 8.1, where the data for
SHG wavelength are approximated to the closest available argon laser line of 457 nm. The values for
µ
s
′
and
µ
a
were measured by a dual integrating sphere setup. We took the
g
values (0.95) from the literature
as reliable values are difficult to obtain by goniometry when
g
is >0.9 and the light is essentially forward
scattered, as is the case for highly organized tissues such as the muscle and tendon. At both wavelengths,
scattering is at least an order of magnitude stronger than absorption. More notably, 25% glycerol reduces
scattering by a factor of 2 over the control tissue whereas 50% and 75% glycerol solution deliver an order
