Biology Reference
In-Depth Information
forward-detected CARS (F-CARS) signals are collected using an air condenser
(N.A.
0.55). External photomultiplier tube (PMT) (H7422-40 and R3896,
Hamamatsu, Japan) detectors are used to receive the E-CARS and F-CARS
signals. The same intensity scale should be used for each set of experiment.
¼
9.1.2.1.3
Results
Figure 9.2
shows E-CARS images of the upper jejunum (section 2) from three dif-
ferent chow-fed, female mice. The left panel is from a mouse that has been fasted for
6 h prior to euthanasia. The middle panel is from a mouse that has been fasted for 6 h
and then gavaged with 200
l olive oil 2 h prior to euthanasia. The right panel is from
a mouse that has been fasted for 6 h and then gavaged with 200
m
l olive oil 4 h prior
to euthanasia. The middle panel has the strongest signal from the CLDs as observed
by the white signals present in the villi. These images show the dynamic nature of
CLDs in the small intestine—appearance and disappearance of lipids from dietary
sources.
m
9.1.2.2
Oil Red O
The mechanism of Oil Red O staining is based on the physical properties of the dye
itself, in which it is more soluble in neutral lipids rather than in the solvent that it is
dissolved in. For Oil Red O staining, fixed tissue is required. One limitation of this
method is that fixing the tissue can lead to inadvertent extraction of lipid from the
tissue. Alternate dyes such as Sudan Black B or fluorescent dye may also be used for
visualizing TAG in tissues.
9.1.2.2.1
Materials
1.
10% formalin
2.
10% sucrose in PBS
3.
Tissue-Tek O.C.T. compound
4.
Mold or aluminum foil
FIGURE 9.2
CARS images of the jejunum in mice. (A) Fasted for 6 h prior to euthanasia. (B) Fasted for 6 h
and then gavaged with 200
l olive oil. Mice were euthanized 2 h post gavage. (C) Fasted
for 6 h and then gavaged with 200 ml olive oil 4 h post gavage.
m
