Fluorescence Depletion by Stimulated Emission in Single-Molecule Spectroscopy

This thesis presents the first application of the stimulated emission depletion (STED) technique to the field of single-molecule fluorescence spectroscopy. It is demonstrated that fluorescence quenching induced by STED is reversible and can be repeated a large number of cycles on a single molecule. Being ideal point-like probes, single molecules can therefore be used to characterize the resolution of STED microscopes. In a spectroscopic study, two simplifed models of the photophysical processes involved in STED are analyzed and applied to the experimental determination of the stimulated emission cross sections on a single-molecule level. In addition, the STED concept as applied in subdiffraction-resolution microscopy is transferred to fluorescence fluctuation spectroscopy. A successful implementation promises to expand the possibilities particularly of fluorescence correlation spectroscopy (FCS) which is already the most widely used fluctuation technique today but is restricted to concentrations on the nanomolar scale. Reducing the detection volume beyond the diffraction limit could render the micromolar range accessible and could thus open up new applications in the life sciences.

Level thesis:degree_level

thesis.doctoral

Grantor dc:publisher

Universität Heidelberg

Year

2004

Author dc:creator

• Kastrup, Lars

Contributors dc:contributor

• Wolfrum, Jürgen

Repository record source_url

http://www.ub.uni-heidelberg.de/archiv/5033

OAI identifier oai:identifier

oai:archiv.ub.uni-heidelberg.de:5033