Time-resolved fluorescence correlation spectroscopy

A new method of performing fluorescence correlation spectroscopy (FCS) measurements for mixtures of several fluorescent molecular species is introduced. It uses time-resolved fluorescence detection for separating the different FCS-contributions from the different species. This allows simultaneous and independent monitoring of the diffusion of several molecular species in one sample, or performing multi-label cross-correlation measurements. In this way, the proposed method is equivalent to dual- or multi-color FCS. However, it is simpler to implement experimentally, because it requires only single wavelength excitation and detection. This Letter outlines the theoretical basis and presents experimental results of the method.