Biomedical Engineering Reference
In-Depth Information
A comprehensive confocal Raman microscopy study was carried out by
Glenn et al. [29] to identify and quantify AGEs in Bruch's membrane in a non-
destructive, analytical fashion. The authors dissected Bruch's membrane and
the innermost retinal layers of the underlying choroid from 56 postmortem
eye-cups, placed the samples onto microscope slides, and recorded Raman
spectra with a confocal microscope. In parallel, they analyzed homogenized
tissue samples with standard chemical analytical techniques including reverse-
phase high-performance liquid chromatography (HPLC), mass spectrometry,
and immunohistochemistry. One of their main Raman results for a spectral
analysis of Bruch's membrane is shown in Fig. 12.7. Using a range of statistical
approaches, the authors were able to deconvolute the Raman spectra into
component spectra characteristic for heme, collagen, and AGE contributions,
all shown, respectively, in Fig. 12.7a-c. Importantly, three AGE-specific Stokes
lines could be identified at 880, 980, and 1080-1090 cm
−
1
[29].
The Raman data set for the AGEs provided quantitative data that com-
pared well with the conventional analytical results, and as an advantage they
offered the possibility to investigate AGEs with high spatial resolution. Using
the Raman spectral fingerprint of AGEs, the authors were able to predict
chronological age of the tissue samples and to confirm a gender-specific differ-
ence in AGE concentration, with higher levels found in females. The ability to
accurately predict chronological age in Bruch's membrane from Raman spec-
tra is a significant finding, showing that AGEs in long-lived proteins correlate
Fig. 12.7.
Analysis of Raman spectra obtained in vitro for Bruch's membrane,
showing component spectra
(a)
and
(b)
for heme and collagen, respectively, and
a residual spectrum
(c)
assigned to Raman responses of age-related glycation end
products and proteins. Adapted from [29]
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