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Phys Rev Lett. 2005 May 6;94(17):178104. Epub 2005 May 4.

Fluorescence fluctuation spectroscopy in subdiffraction focal volumes.

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Department of NanoBiophotonics, Max Planck Institute for Biophysical Chemistry, 37077 Göttingen, Germany.


We establish fluorescence fluctuation spectroscopy (FFS) with nanoscale detection volumes generated by stimulated emission depletion. Our method applies fluorescence correlation spectroscopy and fluorescence intensity distribution analysis to extract molecular information about mobilities and fluorescence emission in solution. The combination of correlation analysis with that of photon intensity distributions reveals a fivefold squeezing of the detection volume over current diffraction-limited systems, which is in full agreement with the simultaneously demonstrated 25-fold reduction in (axial) focal transit time. Our method significantly extends the potential of far-field FFS, including for the noninvasive investigation of molecular reactions at higher concentrations.

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