CryoSTED microscopy [Elektronische Ressource] : a new spectroscopic approach for improving the resolution of STED microscopy using low temperature / presented by Arnold Giske

Dissertationsubmitted to theCombined Faculties for the Natural Sciences and for Mathematicsof the Ruperto-Carola University of Heidelberg, Germanyfor the degree ofDoctor of Natural Sciencespresented by Diplom-Physiker Arnold Giskeborn in Almaty, Kasachstanoral examination: 07. November 2007CryoSTED microscopyA new spectroscopic approach for improving the resolution ofSTED microscopy using low temperatureReferees: Prof. Dr. Stefan HellProf. Dr. Christoph CremerAbstractThis work presents a new approach for the further improving of the resolutionin the sub-diffraction fluorescence STED microscopy. The reduction of the sampletemperature down to 76K eliminates the disadvantages of anti-Stokes fluorescenceexcitation and allows the use of wavelengths with larger STED efficiencies. On thestudied dyes, this approach enhances the resolving power by a factor of 1.6 comparedtoroomtemperatureandsuggestsfurthertechnicalimprovementsforsuchresolutionincrease. Besides resolution, the present work solves the issue of the high tripletpopulationinfluorescenceimagingatliquidnitrogentemperature. Atechniqueforthetriplet depopulation has been developed. It provides a manifold brightness increase inthe fluorescence imaging at low temperatures. The utilization of the low temperaturealso increases the photo-stability of the fluorescent markers.