Raman Spectroscopy of Ocular Tissue - Emerging Raman Applications and Techniques in Biomedical and Pharmaceutical Fields - page 281

concentrations, and to track changes occurring with developing tissue patholo-
gies. The tissue components could include internal structural compounds such
as proteins and lipids, molecules such as glucose, external compounds such as
drugs, and also molecular biomarkers for health or disease, such as macular
pigment or glycation products. Also, the Raman method is capable, in princi-
ple, of generating all spectral information from very small tissue volumes. It is
therefore useable for spatially resolved detection of bio-medically interesting
tissue compounds as well as for their spatially integrated detection. Spatially
resolved detection could be achieved in high-resolution imaging configurations.
Since Raman scattering is a relatively weak optical effect, high excitation
light levels and/or exposure times are usually required to obtain spectra with
good signal-to-noise ratios. This poses a strong challenge for applications to
the living human eye, since the retina is highly light sensitive, and therefore
can be easily and irreversibly damaged. Raman measurements of anterior oc-
ular media can employ light excitation and detection geometries that avoid
excessive retinal exposure, such as excitation light paths that are perpendic-
ular to the optical axis. Probing of inner ocular structures such as the lens,
however, necessitates some degree of a collinear detection scheme, and expo-
sure of the retina with high light levels can be expected to be problematic.
In this chapter, we describe selected examples of Raman work carried out
in ocular tissue research, with emphasis on recent in vivo applications where
possible. For earlier, more comprehensive reviews, including Raman studies
of visual pigments, we refer the reader to the literature [1, 2].
12.2 Protein Structure Investigations of the Lens
The lens is positioned between the aqueous humor and the vitreous humor
and provides about 30% of the eye's refractive power. It consists of a highly
viscous, gel-like crystalline protein structure and is able to change its cur-
vature and, correspondingly, its refractive power. Combined with the fixed
refractive power of the cornea, precise focusing of object rays is achieved in
healthy eyes onto the photoreceptor cells of the retina. The lens contains
about 34% protein and 66% water, with the protein content being the high-
est among human tissues. Protein content is highest in the central regions of
the lens (nucleus) and decreases slightly toward outer regions (cortex). Lens
proteins are synthesized by a monolayer of epithelial cells on the anterior
surface of the lens. In this process, the epithelial cells at the equator of the
lens elongate to form fiber cells that lose their intracellular organelles. The
youngest fiber cells, the cortical fibers, occupy the outer two-thirds of the
lens. With age, the cortical fibers are compressed toward the central nuclear
region.
Since there is virtually no turnover of proteins in the lens with aging, it
can be expected that aging and environmental stress factors lead to changes
in protein structure and water content of the lens, and that these changes