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Proceedings Paper

Method for intracellular imaging of ion concentrations using confocal microscopy and fluorophore lifetimes
Author(s): Kjell Carlsson; Anders Liljeborg; Ronnie M. Andersson; Hjalmar Brismar
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Paper Abstract

There exist a number of fluorescent probes whose lifetimes change in response to ion concentrations (for example H+ and Ca2+) in the surrounding medium. We describe a technique for utilizing this property in a confocal scanning laser microscope. The technique is based on intensity-modulated laser illumination of the specimen, and phase-sensitive lock-in detection of the fluorescent light. In this way we get a lifetime-dependent output signal which, after calibration, can provide information concerning ion concentrations. In the current study we have used a pH sensitive fluorophore, SNAFL-2, to study the performance of this technique. We find that the sensitivity is such that a pH difference of 0.1 units can easily be detected in an 8- bit digital image. Noise measurements show that under realistic conditions we can expect a pixel-to-pixel standard deviation of approximately one to two pH units.

Paper Details

Date Published: 2 May 2000
PDF: 8 pages
Proc. SPIE 3919, Three-Dimensional and Multidimensional Microscopy: Image Acquisition Processing VII, (2 May 2000); doi: 10.1117/12.384198
Show Author Affiliations
Kjell Carlsson, Royal Institute of Technology (Sweden)
Anders Liljeborg, Royal Institute of Technology (Sweden)
Ronnie M. Andersson, Astrid Lindgren Children's Hospital (Sweden)
Hjalmar Brismar, Astrid Lindgren Children's Hospital (Sweden)

Published in SPIE Proceedings Vol. 3919:
Three-Dimensional and Multidimensional Microscopy: Image Acquisition Processing VII
Jose-Angel Conchello; Carol J. Cogswell; Andrew G. Tescher; Tony Wilson, Editor(s)

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