title: Fluorescence lifetime imaging of calcium using Quin-2.
abstract: We describe the use of a new imaging technology, fluorescence lifetime imaging (FLIM), for the imaging of the calcium concentrations based on the fluorescence lifetime of a calcium indicator. The fluorescence lifetime of Quin-2 is shown to be highly sensitive to [Ca2+]. We create two-dimensional lifetime images using the phase shift and modulation of the Quin-2 in response to intensity-modulated light. The two-dimensional phase and modulation values are obtained using a gain-modulated image intensifier and a slow-scan CCD camera. The lifetime values in the 2D image were verified using standard frequency-domain measurements. Importantly, the FLIM method does not require the probe to display shifts in the excitation or emission spectra, which may allow Ca2+ imaging using other Ca2+ probes not in current widespread use due to the lack of spectral shifts. Fluorescence lifetime imaging can be superior to stationary (steady-state) imaging because lifetimes are independent of the local probe concentration and/or intensity, and should thus be widely applicable to chemical imaging using fluorescence microscopy.
authors.affiliation: Department of Biological Chemistry, University of Maryland, Baltimore.
authors.forename: J R
authors.initials: JR
authors.lastname: Lakowicz
authors.forename: H
authors.initials: H
authors.lastname: Szmacinski
authors.forename: K
authors.initials: K
authors.lastname: Nowaczyk
authors.forename: M L
authors.initials: ML
authors.lastname: Johnson
mesh_terms: D000634:Aminoquinolines; D002118:Calcium / Q000378:metabolism*; D005456:Fluorescent Dyes; D008027:Light; D008962:Models, Theoretical; D013050:Spectrometry, Fluorescence / Q000379:methods; D013223:Statistics as Topic
chemical_list: D000634:Aminoquinolines; D005456:Fluorescent Dyes; C035739:Quin2; D002118:Calcium