期刊
INVESTIGATIVE OPHTHALMOLOGY & VISUAL SCIENCE
卷 54, 期 5, 页码 3094-3101出版社
ASSOC RESEARCH VISION OPHTHALMOLOGY INC
DOI: 10.1167/iovs.13-11642
关键词
multiphoton fluorescence microscopy; coherent anti-Stokes Raman scattering (CARS); retina; photoreceptors; retinal pigment epithelium (RPE)
资金
- National Institutes of Health (NIH)/National Institute of Diabetes and Digestive and Kidney Diseases Grant [K25DK095232]
- NIH/National Center for Research Resources (NCRR) [S10RR025447]
- NIH/NCRR/Colorado County Technical Services, Inc. [UL1 RR025780]
PURPOSE. To image the cellular and noncellular structures of the retina in an intact mouse eye without the application of exogenous fluorescent labels using noninvasive, nondestructive techniques. METHODS. Freshly enucleated mouse eyes were imaged using two nonlinear optical techniques: coherent anti-Stokes Raman scattering (CARS) and two-photon autofluorescence (TPAF). Cross sectional transverse sections and sequential flat (en face) sagittal sections were collected from a region of sclera approximately midway between the limbus and optic nerve. Imaging proceeded from the surface of the sclera to a depth of similar to 60 mu m. RESULTS. The fluorescent signal from collagen fibers within the sclera was evident in the TPAF channel; the scleral collagen fibers showed no organization and appeared randomly packed. The sclera contained regions lacking TPAF and CARS fluorescence of similar to 3 to 15 mu m in diameter that could represent small vessels or scleral fibroblasts. Intense punctate CARS signals from the retinal pigment epithelial layer were of a size and shape of retinyl storage esters. Rod outer segments could be identified by the CARS signal from their lipid-rich plasma membranes. CONCLUSIONS. CARS microscopy can be used to image the outer regions of the mammalian retina without the use of a fluorescent dye or exogenously expressed recombinant protein. With technical advancements, CARS/TPAF may represent a new avenue for noninvasively imaging the retina and might complement modalities currently used in clinical practice.
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