Regensburg 2016 – wissenschaftliches Programm

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BP: Fachverband Biologische Physik

BP 48: Posters - Bioimaging and Spectroscopy

BP 48.16: Poster

Mittwoch, 9. März 2016, 17:00–19:00, Poster C

Time Modulated Stimulated Emission Depletion (STED) Based Fluorescence Correlation Spectroscopy (FCS) — •Benedikt Prunsche¹, Peng Gao^1,2, Karin Nienhaus¹, and Gerd Ulrich Nienhaus^1,3 — ¹Institute of Applied Physics, Karlsruhe Institute of Technology, Wolfgang-Gaede-Str. 1, 76131 Karlsruhe, Germany — ²Institute of Nanotechnology, Karlsruhe Institute of Technology, Hermann-von-Helmholtz-Platz 1, 76344 Eggenstein-Leopoldshafen, Germany — ³Department of Physics, University of Illinois at Urbana-Champaign, 1110 West Green Street, Urbana, Illinois 61801, USA

Fluorescence correlation spectroscopy (FCS) is a powerful tool to study bio-molecular dynamics, such as protein diffusion or receptor-ligand interactions inside living cells. It is based on the correlation analysis of fluorescence intensity fluctuations in a small observation volume. These fluctuations arise due to Brownian motion of fluorescent particles in and out of the volume. Because of the diffraction-limited size of the focus volume, conventional FCS is only sensitive to fluorescence fluctuations induced by fluorophores at nanomolar concentrations, which is typically not realized in biological samples. To overcome this limitation, we have reduced the focal volume in all three dimensions by stimulated emission depletion. Background noise, which is inherent in conventional STED based FCS, has been reduced by time-gated detection, and further compensated by using an auxiliary Gaussian depletion beam. As a result, the dynamics of biomolecules at ~10-fold higher concentrations can be quantified with 3D STED based FCS.