Reactant concentrations from fluorescence correlation spectroscopy with tailored fluorescent probes. An example of local calibration-free pH measurement.
The present account is concerned with the measurement of local reactant concentrations by observing specific fluorescent probes in fluorescence correlation spectroscopy (FCS). The Theoretical Analysis section revisits the photophysical, thermodynamic, and kinetic information that is contained in the corresponding FCS correlation curves. In particular, we examine the conditions under which FCS is revealed as a superior tool to measure concentrations of reactive species. Careful molecular engineering of the specific fluorescent probes that simultaneously integrates photophysical, thermodynamic, and kinetic constraints will be required to benefit most from FCS. We illustrate the FCS titration approach with a series of fluorescent probes that we tailored to measure pH at around 4-6 by FCS after two-photon excitation. We show that an optimal design allows one to access pH without any preliminary calibrations such as the determination of the protonation constant or the photophysical properties of the fluorescent probe.
Charier
Sandrine
S
Département de Chimie (CNRS UMR 8640), Ecole Normale Supérieure, 24 rue Lhomond, 75231 Paris Cedex 05, France.
Meglio
Adrien
A
Alcor
Damien
D
Cogné-Laage
Emmanuelle
E
Allemand
Jean-François
JF
Jullien
Ludovic
L
Lemarchand
Annie
A
D005456:Fluorescent Dyes; D006863:Hydrogen-Ion Concentration; D013050:Spectrometry, Fluorescence; D017186:Titrimetry
D005456:Fluorescent Dyes