1 July 2004 Confocal and two-photon imaging in cartilage: expression patterns of Filamin A and B
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Abstract
Optical imaging in cartilage is challenging due to the high levels of intra- and inter-cellular autofluorescence. We report here on high-resolution confocal and two-photon imaging of endogenous fluorescence of cartilage and of exogenous fluorescence of filamin A and B protein markers. Confocal laser scanning microscopy offers the advantage of quasi-theoretical spatial resolution and minimizes the autofluorescence contribution by eliminating the out-of-focus light. In non-labeled cartilage, we observe mostly intracellular autofluorescence that, due to the uniform distribution within the cell, can be further effectively minimized by careful choice of experimental parameters. The fluorescence of the exogenous markers AlexaFluor 488 and AlexaFluor 568 labeling Filamins A and B, respectively, could also be detected and quantitated using this procedure, revealing topologically different expression levels of filamin A and B proteins in the cartilage growth plate. Two-photon excited fluorescence imaging yielded further resolution improvements and structural and functional information.
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Sebastian Wachsmann-Hogiu, Deborah Krakow, Eiman T. Sebald, Cristina Bertolotto, Dora Acuna, Daniel L. Farkas, "Confocal and two-photon imaging in cartilage: expression patterns of Filamin A and B", Proc. SPIE 5322, Imaging, Manipulation, and Analysis of Biomolecules, Cells, and Tissues II, (1 July 2004); doi: 10.1117/12.561883; http://dx.doi.org/10.1117/12.561883
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KEYWORDS
Confocal microscopy

Cartilage

Tissues

Luminescence

Proteins

Two photon imaging

Laser scanners

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