Fluorescence lifetime spectroscopy for pH sensing in scattering media.

Fluorescence lifetime spectroscopy in the presence of tissuelike scattering is demonstrated from measurements of phase and modulation ratio as a function of modulation frequency using a pH-sensitive dye, Carboxy Seminaphthofluorescein-1 (C-SNAFL-1). From the optical diffusion equation describing the propagation and generation of fluorescence within solutions of 0.5 microM C-SNAFL-1 containing 2.0% (by volume) of Intralipid as a scatterer, the values of the average lifetime of C-SNAFL-1 were determined as the solution pH varied between 5 and 9. Average lifetime values were found to match those measured using traditional phase-modulation measurement in nonscattering media. Furthermore, the robustness of the spectroscopic technique was demonstrated by conducting lifetime measurements at varying scatterer concentrations (1.5-3.0 vol % Intralipid). These results confirm the approach for analytical sensing in scattering media via fluorescence lifetime kinetics in order to track changes in analyte concentrations.