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  Two-photon-excited fluorescence imaging of human RPE cells with a femtosecond Ti:Sapphire laser

Bindewald-Wittich, A., Han, M., Schmitz-Valckenberg, S., Snyder, S. R., Giese, G., Bille, J. F., et al. (2006). Two-photon-excited fluorescence imaging of human RPE cells with a femtosecond Ti:Sapphire laser. Investigative Ophthalmology and Visual Science, 47(10), 4553-4557. doi:10.1167/iovs.05-1562.

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Item Permalink: http://hdl.handle.net/11858/00-001M-0000-002B-ABC4-5 Version Permalink: http://hdl.handle.net/11858/00-001M-0000-002B-ABC5-3
Genre: Journal Article
Alternative Title : Two-photon-excited fluorescence imaging of human RPE cells with a femtosecond Ti:Sapphire laser

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InvestOphthalmolVisSci_47_ 2006_4553.pdf (Any fulltext), 383KB
 
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https://dx.doi.org/10.1167/iovs.05-1562 (Any fulltext)
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 Creators:
Bindewald-Wittich, Almut, Author
Han, Meng, Author
Schmitz-Valckenberg, Steffen, Author
Snyder, Sarah Rebecca1, Author              
Giese, Günter1, Author              
Bille, Josef F., Author
Holz, Frank G., Author
Affiliations:
1Department of Biomedical Optics, Max Planck Institute for Medical Research, Max Planck Society, ou_1497699              

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 Abstract: PURPOSE: To record the distribution and spectrum of human retinal pigment epithelial cell lipofuscin (LF) by two-photon-excited fluorescence (TPEF) and confocal laser scanning microscopy. METHODS: Ex vivo TPEF imaging of the human retinal pigment epithelium (RPE) of human donor eyes was conducted with a multiphoton laser scanning microscope that employs a femtosecond Ti:sapphire laser as an excitation laser source. The spectrum of autofluorescence of LF granules was analyzed with a confocal laser scanning microscope coupled to a UV argon laser. RESULTS: TPEF examination allowed for imaging of RPE cell morphology and intracellular distribution of LF granules with high-contrast and submicrometer resolution. Variations in cell size and shape as well as in autofluorescence spectra of individual LF granules were recorded. The typical diameter of LF granules was found to be below 1 mum, with some RPE cells possessing larger granules. Remarkably, enhanced blue-green autofluorescence was observed from these larger LF granules. CONCLUSIONS: TPEF imaging represents a novel tool for the investigation of morphologic and spectral characteristics of human RPE cells. Spectral variations of individual LF granules may indicate differences in the complex molecular composition. Compared to conventional single-photon excited autofluorescence, TPEF with a tunable laser source allows for reduced photo damage and deeper sensing depth. It may help to elucidate further the pathophysiological role of LF accumulation as a common downstream pathogenetic pathway in retinal diseases. With the proof of principle from this ex vivo study, further work is now planned to evaluate the safety of TPEF RPE imaging in RPE cultures and animal models.

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Language(s): eng - English
 Dates: 2005-12-082006-08-142006-10-01
 Publication Status: Published in print
 Pages: 5
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 Rev. Method: Peer
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Title: Investigative Ophthalmology and Visual Science
Source Genre: Journal
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Publ. Info: Hagerstown, MD, etc. : Association for Research in Vision and Ophthalmology, etc.
Pages: - Volume / Issue: 47 (10) Sequence Number: - Start / End Page: 4553 - 4557 Identifier: ISSN: 0146-0404
CoNE: https://pure.mpg.de/cone/journals/resource/110978984074949