Fluorescence lifetime imaging of intracellular calcium in COS cells using Quin-2.
We describe the first fluorescence lifetime images of cells. To demonstrate this new capability we measured intracellular images of Ca2+ in COS cells based on the Ca(2+)-dependent fluorescence lifetime of Quin-2. Apparent fluorescence lifetimes were measured by the phase-modulation method using a gain-modulated image intensifier and a slow-scan CCD camera. We describe methods to correct the images for photobleaching during acquisition of the data, and to correct for the position-dependent response of the image intensifier. The phase angle Quin-2 images were found to yield lower than expected Ca2+ concentrations, which appears to be the result of the formation of fluorescent photoproducts by Quin-2. Fluorescence lifetime imaging (FLIM) does not require wavelength-radiometric probes and appears to provide new opportunities for chemical imaging of cells.
Lakowicz
J R
JR
Department of Biological Chemistry, University of Maryland School of Medicine, Baltimore.
Szmacinski
H
H
Nowaczyk
K
K
Lederer
W J
WJ
Kirby
M S
MS
Johnson
M L
ML
D000818:Animals; D002118:Calcium / Q000032:analysis*; D002460:Cell Line; D002522:Chlorocebus aethiops; D007668:Kidney / Q000737:chemistry* / Q000166:cytology; D008856:Microscopy, Fluorescence / Q000295:instrumentation / Q000379:methods*
D002118:Calcium