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Advanced Optical Microscopy Centre

Equipment & expertise

Advanced Optical Microscopy Centre

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Fluorescence fluctuation spectroscopy (FFS)

A family of methods that allow quantifying molecular properties such as concentrations, mobility, stoichiometry, interaction affinities of diffusing molecules at equilibrium. The simplest implementation is fluorescence correlation spectroscopy (FCS), where fluorescence signals from freely diffusing molecules is recorded on a confocal microscope (without image scanning). The resulting time trace is analyzed via autocorrelation, which then informs on diffusion rates and concentration.


 Illustration of FCS. Adapted from Schrimpf et al., 2018.

Illustration of FCS. Adapted from Schrimpf et al., 2018.

Image correlation spectroscopy extends FFS analysis to imaging data. With Raster ICS (RICS), for example, mobility and concentration information is obtained from confocal laser scanning microscopy images.  


Illustration of RICS. Adapted from Schrimpf et al., 2018.

Arbitrary-region ICS (ARICS) even allows mapping molecular properties in e.g. a biological structure, providing detailed insights in the workings of complex biological machineries.

Illustration of ARICS. Adapted from Hendrix et al., 2016.

In our research, we develop continuously refine existing FFS methods and develop novel FFS modalities to extract maximal and quantitative (molecular) information from confocal fluorescence data.