Single Molecule Wide-Field Microscope with Light Sheet Illumination

• Subject Area: Medicine
• Term: Funded in 2020
• Project identifier: Deutsche Forschungsgemeinschaft (DFG) - Project number 437835311

The dynamic visualization of subcellular structures provides an essential tool to explain fundamental, biological processes, such as the cell cycle, protein interactions, compartmentalization or transport. Recent methodological advances enabled optical resolutions below the limit of light diffraction, in a range of 20 – 120 nm. While pointillism and STED resolve structures down to 20 nm, these methods only allow live cell imaging in specialized contexts, i.e. with prokaryotic cells and using specific dyes and custom made setups. SIM however is a wide-field method achieving super-resolution (120 nm) by structured illumination patterns with fast camera-based acquisition ideal for live cell applications. As such, this method is compatible with conventional fluorophores. By changing the structured illumination from a grid to a lattice pattern, the number of raw images and thereby the amount of phototoxicity is reduced, which represents a critical requirement for live cell imaging of more photosensitive mammalian cells. Further, Lattice SIM offers a higher penetration depth than other approaches, enabling volumetric live cell imaging of larger specimen, such as organoids. The setup is the only commercially available microscope implementing the proprietary Lattice SIM with SMLM (single molecule localization microscopy) offering versatile, straight forward and flexible super-resolution microscopy for different experimental conditions. Such imaging platform shall enable the high resolution spatiotemporal characterization of fundamental biological processes such as for example nuclear actin or single molecule dynamics, which were previously difficult to tackle by conventional microscopy.

• DFG Programme: Major Research Instrumentation
• Major Instrumentation: Single Molecule Weit-Feld-Fluoreszenzmikroskop mit Light Sheet Illumination
• Instrumentation Group: 5040 Spezielle Mikroskope (außer 500-503)
• Applicant Institution: Albert-Ludwigs-Universität Freiburg
• Leader: Professor Dr. Robert Grosse