Total Internal Reflection Fluorescence for Adsorbed Probe Molecule Studies of Liquid/Solid Interfacial Environments

Total internal reflection fluorescence (TIRF) is developed as a surface selective method to allow the environment of a liquid/solid interface to be probed by fluorescent molecules which are adsorbed from solution. The method has been used to detect pyrene sorbed to an octadecylsilane-derivatized fused-silica plate and resolve its spectral emission so that vibronic intensity ratios can be calculated and the surface environment characterized. Adsorption equilibria of the fluorescent probe to the surface and the depth of penetration of the evanescent excitation beam provide the basis for predicting interference from probe molecules in the solution phase. These predictions were validated by replacing the solution overlaying the alkylated silica interface with saturated vapor and comparing the apparent surface environments.