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James R. Beattie, Sorcha Finnegan, Ross W. Hamilton, Manir Ali, Christopher F. Inglehearn, Alan W. Stitt, John J. McGarvey, Paul M. Hocking, William J. Curry; Profiling Retinal Biochemistry in the MPDZ Mutant Retinal Dysplasia and Degeneration Chick: A Model of Human RP and LCA. Invest. Ophthalmol. Vis. Sci. 2012;53(1):413-420. doi: https://doi.org/10.1167/iovs.11-8591.
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Raman microscopy, a rapid nondestructive technique that profiles the composition of biological samples, was used to characterize retinal biochemistry in the retinal dysplasia and degeneration (rdd) and wild-type (wt) chick retina during retinogenesis and at hatching.
Embryonic day (E)13 and posthatch day (P)1 rdd and wt retinal cross-sections (n = 3 of each line at each age) were profiled using 633 helium–neon laser excitation. The biochemical composition was determined using computational analysis of the Raman spectra. In parallel histology, TUNEL and glial fibrillary acidic protein (GFAP) immunostaining were used to visualize retinal dysfunction.
Principal component (PC) analysis of the Raman spectra identified 50 major biochemical profiles, but only PCs that made significant contributions to variation within rdd and wt retina were mapped. These significant PCs were shown to arise from DNA, various fatty acids, melanin, and a number of proteins. Distinct patterns of GFAP immunostaining and a larger population of TUNEL-positive nuclei were observed in the rdd versus wt retina.
This study has demonstrated that Raman microscopy can discriminate between major retinal biomolecules, thus providing an unbiased account of how their composition varies due to the impact of the MPDZ null mutation in the rdd chick relative to expression in the normal wt retina.
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