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Journal of Biomedical Optics

Confocal fluorescence spectroscopy of subcutaneous cartilage expressing green fluorescent protein versus cutaneous collagen autofluorescence
Author(s): Daniel S. Gareau; Paulo R. Bargo; William A. Horton; Steven L. Jacques
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Paper Abstract

Optically monitoring the expression of green fluorescent protein (GFP) in the cartilage underlying the skin of a mouse allows tracking the expression of the chondrocyte phenotype. This paper considers how confocal microscopy with spectral detection can sense GFP fluorescence in the cartilage despite light scattering and collagen autofluorescence from the overlying skin. An in vivo experiment tested the abilities of a topical optical fiber measurement and a confocal microscope measurement to detect GFP in cartilage under the skin versus the collagen autofluorescence. An ex vivo experiment tested the ability of a confocal microscope without and with its pinhole to detect a fluorescent microsphere underneath an ex vivo skin layer versus the collagen autofluorescence. In both systems, spectroscopic detection followed by linear analysis allowed spectral discrimination of collagen autofluorescence (MC) and the subdermal green fluorescence (MG) due to either GFP or the microsphere. Contrast was defined as MG/(MG + MC). The in vivo contrast for GFP using optical fiber and confocal measurements was 0.16 and 0.92, respectively. The ex vivo contrast for a fluorescent microsphere using a confocal system without and with a pinhole was 0.13 and 0.48, respectively. The study demonstrates that a topical optical fiber measurement is affected by collagen autofluorescence, while a confocal microscope can detect subdermal fluorescence while rejecting collagen autofluorescence.

Paper Details

Date Published: 1 March 2004
PDF: 5 pages
J. Biomed. Opt. 9(2) doi: 10.1117/1.1645798
Published in: Journal of Biomedical Optics Volume 9, Issue 2
Show Author Affiliations
Daniel S. Gareau, Oregon Health and Science Univ. (United States)
Paulo R. Bargo, Providence St. Vincent Hospital (United States)
William A. Horton, Oregon Health and Science Unvi. (United States)
Steven L. Jacques, Oregon Health and Science Univ. (United States)

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