Elson_04_Time-domain FLIM applied to biological tissue_PPS_Submitted.doc

Elson_04_Time-domain FLIM applied to biological tissue_PPS_Submitted.doc

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Photochem Photobiol Sci 2004 3 795-801 TIME-DOMAIN FLUORESCENCE LIFETIME IMAGING APPLIED TO BIOLOGICAL TISSUE Dan Elson, Jose Requejo-Isidro, Ian Munro, Fred Reavell, Jan Siegel, Klaus Suhling, Paul Tadrous, Richard Benninger, Peter Lanigan, James McGinty, Clifford Talbot, Bebhinn Treanor, Stephen Webb, Ann Sandison, Andrew Wallace, Dan Davis, John Lever, Mark Neil, David Phillips, Gordon Stamp, and Paul French Photonics Group, Physics Department, Imperial College London, London SW7 2AZ, U.K. [email protected] Abstract: Fluorescence lifetime imaging (FLIM) is a functional imaging methodology that can provide information,   not   only   concerning   the   localisation   of   specific   fluorophores,   but   also   about   the   local fluorophore environment.  It may be implemented in scanning confocal or multi-photon microscopes, or in wide-field microscopes and endoscopes.   When applied to tissue autofluorescence, it reveals intrinsic excellent contrast between different types and states of tissue.   This article aims to review our recent progress in developing time-domain FLIM technology for microscopy and endoscopy and applying it to biological tissue.  OCIS codes: (170.2520) Fluorescence microscopy; (170.3650) Lifetime based sensing; (170.6920) Time resolved imaging Introduction Fluorescence lifetime imaging (FLIM) is a functional imaging methodology that is currently experiencing a rapid increase in uptake due to its excellent specificity and its relative insensitivity to intensity artefacts. FLIM is applicable to imaging intracellular function, e.g. using the Förster resonance energy transfer (FRET) technique and can provide information, not only concerning the localisation of a specific fluorophore, but also about the local fluorophore environment (e.g. oxygen, [Ca 2+ ], pH etc [ 1 ]). It may be implemented in scanning confocal or multi-photon microscopes, as well as in wide-field microscopes and endoscopes. When applied to tissue autofluorescence, it reveals intrinsic contrast between different types and states of tissue, potentially providing new non-invasive functional/diagnostic imaging modalities [ 2 ]. This paper aims to describe our approach to time- domain FLIM and to review our recent work, as presented at the ESP 2003 conference [ 3 ] in Vienna, with particular emphasis on its application to biological tissue. It is not intended as a general discussion of FLIM – for this we could suggest the papers by Cubeddu et al. [ 4 ], Wouters et al. [ 5 ] and Tadrous [ 6 ]. For a discussion of FLIM applied to FRET, we suggest the paper by Bastiaens and Squire [ 7 ]. 1
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Photochem Photobiol Sci 2004 3 795-801 Fluorescence imaging is particularly powerful since the use of fluorescent labels can yield high specificity, while quantitative analysis of the fluorescence signal can provide information about the local environment of the fluorophore molecules as well as their localisation. The fluorescence quantum efficiency,
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