Fluorescence cross-correlation spectroscopy (FCCS) was introduced by Eigen and Rigler in 1994 and experimentally realized by Schwille in 1997. It extends the fluorescence correlation spectroscopy (FCS) procedure by introducing high sensitivity for distinguishing fluorescent particles which have a similar diffusion coefficient. FCCS uses two species which are independently labelled with two spectrally separated fluorescent probes. These fluorescent probes are excited and detected by two different laser light sources and detectors commonly known as green and red respectively. Both laser light beams are focused into the sample and tuned so that they overlap to form a superimposed confocal observation volume.
The normalized cross-correlation function is defined for two fluorescent species and which are independent green, G and red, R channels as follows:
where differential fluorescent signals at a specific time, and at a delay time, later is correlated with each other.
Read more about Fluorescence Cross-correlation Spectroscopy: Modeling