Efficient simultaneous fluorescence orientation, spectrum, and lifetime detection for single molecule dynamics

We report on the simultaneous detection of the fluorescence lifetime, spectrum, and three-dimensional dipole orientation determination of single perylene diimide molecules deposited on a silica surface as a model system for studying fluorophore internal and orientational dynamics. We employ a multi-parameter detection scheme to demonstrate how jumps in the orientation of the molecule can be disentangled from spectral jumps, both leading to changes of the detected total fluorescence intensity. The fluorescence lifetime determined simultaneously from the same photons is also sensitive to the orientation of the dipole with respect to the interface between media with different refractive indices. The correlated changes of the lifetime and orientation we observe are in good agreement with theory. (C) 2012 American Institute of Physics. [http://dx.doi.org/10.1063/1.4759108]