Multi-Photon Excitation of Fluorescence and Light Quenching by Stimulated Emission

Time-resolved fluorescence is widely used to study the structure and dynamics of biological macromolecules. The time-resolved decays are typically initiated by one-photon excitation. In this lecture, we describe the emerging topic of multi-photon excitation, in which the fluorophores simultaneously absorb two or more long wavelenths photons to reach the first excited state. Multi-photon excitation provides a more highly oriented population then does single-photon excitation. Additionally, the excited molecules are spatially localized.

This lecture will also describe the use of long wavelength pulses to modify the excited state population, a phenomena we call light quenching. Light quenching can be used to delete fluorophores based on their emission wavelength, orientation or lifetime. Light quenching provides a new approch to time-resolved fluorescence based on the use of multiple light pulses to modify the excited state population.