Super-resolution microscopy by TIRF-SIM
| Background and Problem: Super-resolution microscopy techniques require significantly more illumination photons than conventional micros-copies. This leads to either longer acquisition times and/or enhanced fluorescence bleaching making them difficult to apply to dynamic processes in biology. Approach: Using a combination of structured illumination microscopy (SIM) and total internal reflection fluorescence (TIRF) we are able to achieve nearly 100 nm resolution inside living cells at a frame rate of nearly 10 Hz and more than 70 acquisitions without significant bleaching. Thereby we were able to investigate and understand the multi-motor transport of the cytoskeleton filament MreB inside living bacteria (B. Subtilis). |
Figures
Yeast cell with the fluorescently labeled membrane protein Sag1. (Left) Normal TIRF image. (Right) TIRF-SIM image of the same cell showing higer resolution and contrast. Scale bar is 1 µm. (Collaboration with the lab of R. Wedlich-Söldner) | MreB dynamics in the bacteria Bacillus Subtilis. (Left) Brightfield image of a B. subtilis cell. (Right) TIRF-SIM time series of GFP-labeled MreB in the same cell showing high dynamics. Scale bar is 1 µm. (Collaboration with the lab of P. Graumann) | |
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| Time-lapse image comparison of a lifeAct-mCherry stained HeLa cell. Left: TIRF-SIM, right: TIRF. | ||
Publications
- Huber M, Schreiber A, Olshausen P, Varga B, Kretz O, Barnert S, Schubert R, Eimer S, Kele P, Schiller S
Designer amphiphilic proteins as building blocks for the intracellular formation of organelle-like compartments
2014 Nat Mater - Olshausen P, Soufo H, Wicker K, Heintzmann R, Graumann P, Rohrbach A
Superresolution Imaging of Dynamic MreB Filaments in B. subtilis - A Multiple-Motor-Driven Transport?
2013 Biophys J, Band: 105, Nummer: 5, Seiten: 1171 - 1181 - Spira F, Mueller N S, Beck G, Olshausen P, Beig J, Wedlich-Söldner R
Patchwork organization of the yeast plasma membrane into numerous coexisting domains
2012 Nat Cell Biol, Band: 14, Seiten: 640 - 648 - Rohrbach, A.
Observing secretory granules with a multi-angle evanescent wave microscope
2000 Biophysical Journal, Band: 78, Seiten: 2641 - 2654