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Proceedings Paper

Nanosecond fluorescence microscopy of single cells
Author(s): Susan M. Keating; Theodore G. Wensel
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Paper Abstract

A microscope based time-correlated single photon counting instrument has been used to measure nanosecond fluorescence decays from single cells. The excitation source for the instrument is a frequency doubled train of picosecond pulses from the cavity dumped output of a synchronously pumped dye laser. The dye laser is pumped by a mode-locked argon ion laser. In the microscope, the sample is excited and the emission collected using epi-illumination optics before being transmitted through an adjustable diaphragm, which can be closed to 10 μm in diameter. A Hamamatsu R928 photomultiplier is used to collect the fluorescence which is then analyzed using a non-linear least squares procedure. The microscope has been used to measure the intensity decays of model probes to determine the instrument performance and sensitivity. In addition, intensity and anisotropy decays collected from fura-2 loaded into single adherent rat basophilic leukemia cells were measured to demonstrate that the nanosecond fluorescence microscope can be used to obtain information about the environment and mobility of fluorescent probes in single cells.

Paper Details

Date Published: 1 May 1990
PDF: 7 pages
Proc. SPIE 1204, Time-Resolved Laser Spectroscopy in Biochemistry II, (1 May 1990); doi: 10.1117/12.17685
Show Author Affiliations
Susan M. Keating, Stanford Univ. School of Medicine (United States)
Theodore G. Wensel, Baylor College of Medicine (United States)

Published in SPIE Proceedings Vol. 1204:
Time-Resolved Laser Spectroscopy in Biochemistry II
Joseph R. Lakowicz, Editor(s)

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