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M. Han, G. Giese, S. Schmitz-Valckenberg, A. Bindewald-Wittich, F. G. Holz, M. H. Niemz, J. F. Bille; Two-photon Excited Autofluorescence Imaging of the Human Retina-Choroid Complex. Invest. Ophthalmol. Vis. Sci. 2007;48(13):4245.
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The development of age-related macular degeneration (AMD) is thought to result from an accentuation of the aging process. A minimally invasive, high resolution imaging modality is vital to delineate the age-related structural abnormalities in the retina-choroid complex prior to apparent pathological manifestations of AMD. Based on the endogenous chromophores in the human retina-choroid complex, we employed two-photon excited autofuorescence (TPEF) imaging to characterize the morphological and spectral properties of the human photoreceptors, RPE and choriocapillaries ex vivo.
The human retinas were obtained from four human Caucasian postmortem donor eyes (19, 55, 57 and 64 years old, with normal vision). TPEF imaging was performed with a customized multiphoton laser scanning microscope (Zeiss LSM 510) equipped with a femtosecond Ti:Sapphire laser (720-980 nm). The autofluorescence spectrum of RPE cells was measured with a confocal laser scanning microscope (Leica TCS SP2) coupled to a UV argon laser (364 nm).
As age increases, the regularity of the parafoveal photoreceptor mosaic appears to be preserved (Fig.a,b). Enlarged lipofuscin granules in the aged RPE demonstrate significantly blue-shifted autofluorescence (Fig.d), which coincides with the depletion of melanin pigments. Prominent fibrillar structures in elderly choriocapillaries may represent age-related structure and permeability alterations in the choroid (Fig.f).
TPEF imaging is an elegant and highly efficient tool to delineate the thick, fragile and opaque retina-choroid complex. Requiring neither slicing nor labeling, the most vital components (photoreceptors, RPE, choriocapillaries, Bruch's membrane) in the human retina-choroid complex can be examined with high contrast and subcellular resolution, which may provide fresh insights into the pathogenesis of AMD.
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