Picosecond fluorescence lifetime imaging microscope for imaging of living glioma cells

Qiyin Fang; Jingjing Wang; Yinghua Sun; Thomas Vernier; Thanassis Papaioannou; Javier Jo; Mya Mya Thu; Martin A. Gundersen; Laura Marcu

doi:10.1117/12.592474

29 March 2005 Picosecond fluorescence lifetime imaging microscope for imaging of living glioma cells

Qiyin Fang, Jingjing Wang, Yinghua Sun, Thomas Vernier, Thanassis Papaioannou, Javier Jo, Mya Mya Thu, Martin A. Gundersen, Laura Marcu

Author Affiliations +

Proceedings Volume 5699, Imaging, Manipulation, and Analysis of Biomolecules and Cells: Fundamentals and Applications III; (2005) https://doi.org/10.1117/12.592474
Event: SPIE BiOS, 2005, San Jose, CA, United States

Abstract

In this communication, we report the imaging of living glioma cells using fluorescence lifetime imaging (FLIM) technique. The growing interests in developing novel techniques for diagnosis and minimally invasive therapy of brain tumor have led to microscopic studies of subcellular structures and intracellular processes in glioma cells. Fluorescence microscopy has been used with a number of exogenous molecular probes specific for certain intracellular structures such as mitochondria, peripheral benzodiazepine receptor (PBR), and calcium concentration. When probes with overlapping emission spectra being used, separate samples are required to image each probe individually under conventional fluorescence microscopy. We have developed a wide-field FLIM microscope that uses fluorescence lifetime as an additional contrast for resolving multiple markers in the same essay. The FLIM microscope consists of a violet diode laser and a nitrogen-pumped dye laser to provide tunable sub-nanosecond excitation from UV to NIR. The detection system is based on a time-gated ICCD camera with minimum 80 ps gate width. The performance of the system was evaluated using fluorescence dyes with reported lifetime values. Living rat glioma C6 cells were stained with JC-1 and Rhodamine 123. FLIM images were acquired and their lifetimes in living cells were found in good agreements with values measured in solutions by a time-domain fluorescence spectrometer. These results indicate that imaging of glioma cells using FLIM can resolve multiple spectrally-overlapping probes and provide quantitative functional information about the intracellular environment.

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Qiyin Fang, Jingjing Wang, Yinghua Sun, Thomas Vernier, Thanassis Papaioannou, Javier Jo, Mya Mya Thu, Martin A. Gundersen, and Laura Marcu "Picosecond fluorescence lifetime imaging microscope for imaging of living glioma cells", Proc. SPIE 5699, Imaging, Manipulation, and Analysis of Biomolecules and Cells: Fundamentals and Applications III, (29 March 2005); https://doi.org/10.1117/12.592474

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KEYWORDS

Luminescence

Fluorescence lifetime imaging

Microscopes

Picosecond phenomena

Semiconductor lasers

Microscopy

Imaging systems

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