Maximizing far-field optical microscopy resolution through selected fluorophore transitions [Elektronische Ressource] / put forward by Eva Rittweger

Dissertationsubmitted to theCombined Faculties for the Natural Sciences and for Mathematicsof the Ruperto-Carola University of Heidelberg, Germanyfor the degree ofDoctor of Natural SciencesPut forward byDiplom-Physikerin Eva Rittwegerborn in ColognethOral examination: 16 December 2009Maximizing far-field opticalmicroscopy resolution throughselected fluorophore transitions0Referees: Prof. Dr. Stefan W. HellProf. Dr. Ju¨rgen WolfrumAbstract:Stimulated emission depletion (STED) microscopy and related nanoscopy tech-niques, which utilize a saturable optical transition between a bright and a darkstate, overcome the diffraction barrier by confining one of the states to an areasmaller than the Airy disk. Scanning this area across the specimen yields sub-diffraction images by registering inseparable fluorescent markers sequentially intime. Despite the progress made in nanoscopy so far, maximizing the resolu-tion has been hampered by the efficiency of the utilized optical transition andthe photostability of the fluorophores. Here, the optical transition responsiblefor breaking the barrier was studied in order to maximize its efficiency. For arange of fluorophores (dyes, proteins, quantum dots, color centers) the nature ofthe responsible process could be clarified. It was also investigated whether heatcould serve as an imaging contrast to provide an alternative tofluorescence.