Supercritical angle localization microscopy

• Applicant: Professor Dr. Jonas Ries
• Subject Area: Biophysics
• Term: from 2013 to 2017
• Project identifier: Deutsche Forschungsgemeinschaft (DFG) - Project number 240272737

Single-molecule localization-based superresolution microscopy improves the resolution in optical microscopy by one order of magnitude through stochastic activation and subsequent localization of individual fluorophores. The aim of this project is to develop a robust and simple 3D detection scheme for localization microscopy with nanometer thee-dimensional resolution, based on the principle of surface-generated fluorescence. Surface-generated fluorescence occurs when a fluorophore is in the vicinity of a water-glass interface. It is emitted into large angles above the critical angle and its intensity strongly depends on the distance of a fluorophore from the interface. By splitting high and low emission angles and imaging them simultaneously we can determine the precise axial position of single molecules from their relative intensities in the two channels.As a first application of this technique we will investigate the complex organization of proteins involved in endocytosis with an unprecedented three-dimensional resolution. As a second application we will determine the precise three-dimensional positions of several components of the nuclear pore complex. This will allow us to match these proteins to the 3D electron tomography data of the whole nuclear pore and to determine how they are organized in the nuclear pore complex.

• DFG Programme: Research Grants