Raman Imaging and Raman Mapping - Emerging Raman Applications and Techniques in Biomedical and Pharmaceutical Fields - page 103

progress of cell division can be followed by imaging the beta-sheet amide I

signature at 1 , 686 cm − 1 . Cytochrome c 753 cm − 1 is also concentrated near
the nascent boundary of the two daughter cells and lipids 2 , 852 cm − 1 at
the cell walls. A short movie of the process can be viewed at the following
web site. http://dx.doi.org/10.1117/1.2952192.1.
Incorporation and distribution of amino acids into cells can be followed
by Raman mapping using deuterium-labeled amino acids in the growth me-
dia [30]. The advantage of this stable isotope technique is that it puts C-D
stretches of the amino acids into the 2 , 100-2 , 300 cm − 1 region, where there
is no interference from other Raman bands. Distribution of incorporated
phenylalanine-d 5 into a HeLa cell is shown in Fig. 5.4. As expected, structures
such as nucleoli in the nucleus are regions of concentrated accumulation. The
cell takes up either phenylalanine or phenylalanine-d 5 randomly, as shown by
the uniform distribution of phenyl C-D/phenyl C-H stretch intensity across
the cell. Because point-by-point mapping (dwell time 1 s) was used acquisi-
tion of the 900 pixel images (30 × 30 0 . 47 μ m steps, 15 × 15 μ m area) required

15 min, plus microscope stage and camera readout time.

5.3.2 Lung Tissue

Congenital cystic adenomatoid malformations (CCCMs) have been examined
in four specimens taken from the lungs of infants who had undergone lung
surgery [31]. There are several classes of CCCMs, all of which prevent for-
mation of normal alveolar growth. From the specimens both FTIR images
(64

64 pixels) and point-by-point Raman maps with definitions ranging be-
tween 42

×

80 pixels were obtained with spatial resolution similar
to that for the FTIR images. Coarse resolution was deliberately used to allow
examination of relatively large regions. Unsupervised cluster analysis was used
to separate the specimens into groups. The combination of FTIR and Raman
provided better results than either alone. FTIR is more sensitive to mucus
(i.e., to polysaccharides) and lipids than Raman spectroscopy. However, iden-

× 46 and 80 ×

tification of lipids by low wavenumber < 1 , 000 cm − 1 is better performed by
Raman spectroscopy. The combination of the two modalities provided better
results than either alone. Of course, as discussed above, the time required for
Raman mapping could be reduced from many hours to a few minutes with
line scanning or the other modes discussed in earlier sections.

5.3.3 Cell and Small Animal Imaging with Surface-Enhanced
Raman Spectroscopic (SERS) and Other Nanoparticle Tags

SERS is discussed in greater detail in Chap. 8. We discuss only two recent
imaging applications. An advantage of SERS labels is that several different
labels, and therefore several cell or tissue components, can be tracked si-
multaneously. In theory, SERS labeling should allow dozens or perhaps even
hundreds of distinguishable labels to be used. SERS-active metallic particles