Finalist EPFL doctorate Award 2014 - Stefan Geissbühler
© 2014 EPFL
Special distinction from the selection committee to Stefan Geissbühler for his thesis "Structural and Functional Stochastic Super-Resolution Microscopy". Thesis n° 5826 (2013). Thesis directors : Prof. T. Lasser, M. Leutenegger
Super-resolution fluorescence microscopy visualizes biological structures with about an order of magnitude below the classical resolution limit and thus promises to strengthen our understanding of living processes. However, its successful application is challenging, requiring particular care in sample preparation and relying on tedious alignment procedures and/or complicated data analysis software.
This Ph.D. thesis focused on the development and characterization of novel concepts for super-resolution microscopy based on stochastic fluorescence fluctuations in order to extend and/or simplify existing techniques. Stochastic super-resolution microscopy overcomes the diffraction barrier by processing movies of randomly blinking fluorophores either with single-molecule localization (PALM/STORM/GSDIM) or with a statistical analysis based on higher-order cumulants (SOFI). While localization microscopy can pro¬vide higher resolution improvements under appropriate conditions, SOFI scores in the compatibility with weakly emitting fluorophores and a wide range of blinking conditions. With balanced SOFI we extended the original SOFI analysis by combining several cumulant orders to map molecular statistics and linearize the bright-ness and blinking response of higher-order cumulants. Using cross-cumulation between multiple depth and/or color channels, we demonstrated whole-cell 3D super-resolution microscopy without mechanical scanning as well as spectral unmixing of multiple fluorophores with overlapping emission spectra.
Figure 1 : Live-cell multi-plane SOFI of a Hela cell expressing Vimentin-Dreiklang