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Applied Optics

Applied Optics


  • Vol. 43, Iss. 27 — Sep. 20, 2004
  • pp: 5173–5182

Simultaneous two-photon spectral and lifetime fluorescence microscopy

Damian K. Bird, Kevin W. Eliceiri, Ching-Hua Fan, and John G. White  »View Author Affiliations

Applied Optics, Vol. 43, Issue 27, pp. 5173-5182 (2004)

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When a fluorescence photon is emitted from a molecule within a living cell it carries a signature that can potentially identify the molecule and provide information on the microenvironment in which it resides, thereby providing insights into the physiology of the cell. To unambiguously identify fluorescent probes and monitor their physiological environment within living specimens by their fluorescent signatures, one must exploit as much of this information as possible. We describe the development and implementation of a combined two-photon spectral and lifetime microscope. Fluorescence lifetime images from 16 individual wavelength components of the emission spectrum can be acquired with 10-nm resolution on a pixel-by-pixel basis. The instrument provides a unique visualization of cellular structures and processes through spectrally and temporally resolved information and may ultimately find applications in live cell and tissue imaging.

© 2004 Optical Society of America

OCIS Codes
(170.3650) Medical optics and biotechnology : Lifetime-based sensing
(170.3890) Medical optics and biotechnology : Medical optics instrumentation
(180.0180) Microscopy : Microscopy
(180.2520) Microscopy : Fluorescence microscopy

Original Manuscript: March 2, 2004
Revised Manuscript: May 4, 2004
Manuscript Accepted: June 18, 2004
Published: September 20, 2004

Damian K. Bird, Kevin W. Eliceiri, Ching-Hua Fan, and John G. White, "Simultaneous two-photon spectral and lifetime fluorescence microscopy," Appl. Opt. 43, 5173-5182 (2004)

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