Calcium-dependent fluorescence lifetimes of Indo-1 for one- and two-photon excitation of fluorescence.
We characterized the fluorescence intensity decays of Indo-1, which is commonly used as an emission wavelength-ratiometric calcium probe. The apparent lifetime of the long-wavelength side of the emission of Indo-1 is dependent on Ca2+. This long-wavelength emission displays the characteristics of an excited-state reaction, that is, a negative preexponential component in the multiexponential analysis. The emission spectra and lifetime of Indo-1 appear to be identical for one-photon and two-photon excitation at 351 and 702 mn, respectively, suggesting that the relative one- and two-photon cross sections are similar for the calcium-free and calcium-bound forms of Indo-1. Also, the two-photon cross section of Indo-1 is relatively high, about 4 x 10(-49) cm4 s/photon molecule at 690 nm for both the calcium-free and calcium-bound forms. Hence, Indo-1 can be used for calcium imaging based on one- or two-photon excitation, using either emission wavelength ratios or lifetime imaging methods.
Szmacinski
H
H
Department of Biological Chemistry, University of Maryland at Baltimore 21201.
Gryczynski
I
I
Lakowicz
J R
JR
D002118:Calcium / Q000737:chemistry*; D005453:Fluorescence; D005456:Fluorescent Dyes / Q000737:chemistry*; D007211:Indoles / Q000737:chemistry*; D017785:Photons
D005456:Fluorescent Dyes; D007211:Indoles; C048960:indo-1; D002118:Calcium