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H.–Y. Tan, S.–J. Lin, W. Lo, S.–W. Teng, R.–J. Wu, W.–C. Lin, C.–H. Hsiao, D.–K. Ma, S.–M. Huang, C.–Y. Dong; Multiphoton Autofluorescence and Second Harmonic Generation Microscopy for Imaging Corneal Pathologies . Invest. Ophthalmol. Vis. Sci. 2006;47(13):2953.
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© ARVO (1962-2015); The Authors (2016-present)
To demonstrate the application of multiphoton autofluorescence and second harmonic generation microscopy for imaging corneal pathologies.
Human corneal specimens with representative corneal pathologies with different etiologies were obtained during penetrating keratoplasty. A home–built multiphoton microscope was used for imaging the cellular and collagenous structural alterations. Cellular components can be visualized with cytoplasmic autofluorescence, and the collagen fibers can be demonstrated with the non–linear optical effect of second harmonic generation.
Autofluorescent activated keratocytes and inflammatory cells can be visualized within diseased cornea, which is different from the queiscent autofluorescence–free keratocytes in normal cornea. Destruction and alteration of stroma collagen can also be highlighted with the malalignment or abscence of second harmonic generation signals.
We demonstrate that multiphoton autofluorescence and second harmonic generation microscopy can provide subcellular resolution for noninvasively imaging corneal pathologies, which may be a powerful tool for corneal physiological and pathological researches, with the potential to be developed into an in vivo imaging modality.
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