12th Carl Zeiss sponsored workshop on ... - Institut Fresnel

Inverse- Fluorescence Correlation Spectroscopy
Stefan Wennmalm, Lei Xu, Per Thyberg and Jerker Widengren
Royal Institute of Technology, Applied Physics,
Experimental Biomolecular Physics Group
106 91 Stockholm, Sweden
Contact: stewen@kth.se
Using Fluorescence Correlation Spectroscopy (FCS) for analysis of biomolecules requires
labelling. An alternative approach that circumvents the need for labelling is inverse-FCS
(iFCS), where the signal from a medium surrounding the particles of interest is analyzed, as
opposed to a signal from the particles themselves. As unlabeled particles traverse the
detection volume, part of the medium will be displaced whereby fluorescence fluctuations are
generated. This allows for analysis of non-labelled particles or potentially biomolecules. We
have tested iFCS by measuring on unlabeled polystyrene beads of 800, 400, 200 and 100 nm
diameter, in a surrounding medium of highly concentrated alexa 488-fluorophores. Our aim is
now to enhance the sensitivity of iFCS and thereby allow for analysis of unlabelled
biomolecules of ≤ 10 nm diameter.
A
B
Fig. 1. Cartoon showing the principle of inverse-FCS: A) An unperturbed, high, fluorescence
signal is measured when the detection volume is absent from particles. B) The signal is
reduced upon the entrance of a particle. C) The fluorescence signal is restored as the particle
leaves the detection volume.
12 th <strong>Zeiss</strong> <strong>sponsored</strong> FCS <strong>workshop</strong> – Cargèse, Corsica, France – <strong>12th</strong> -16th October 2009
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