Markus Sauer
Markus Sauer, Department
of Biotechnology and Biophysics, University of Wuerzburg, Germany
Title
Super-Resolution Expansion Microscopy
Abstract
In the last decade, super-resolution
microscopy has evolved as a very powerful method for sub-diffraction resolution
fluorescence imaging of cells and structural investigations of cellular
organelles. Super-resolution microscopy
methods can now provide a spatial resolution that is well below the diffraction
limit of light microscopy, enabling invaluable insights into the
spatial organization of proteins in biological samples. However, current super-resolution measurements become
error-prone below 25 nm. In addition, refined single-molecule
localization microscopy methods achieved localization precisions of only a few
nanometers, but here too translation of such high localization precisions into
sub-10 nm spatial resolution in biological samples remains challenging. In my
contribution I will discuss two possibilities to bypass these limitations. One
is based on physical expansion of the cellular
structure by linking a protein of interest into a dense, cross-linked
network of a swellable polyelectrolyte hydrogel. Since its first introduction
by Boyden and co-workers in 2015, expansion microscopy (ExM) has shown
impressive results including the magnified visualization of pre- or
post-expansion labeled proteins and RNAs with fluorescent proteins, antibodies,
and oligonucleotides, respectively, in cells, tissues, and human clinical
specimen. By combining ExM with super-resolution microscopy it is potentially
possible to enable multicolor molecular resolution fluorescence imaging. The
other approach uses resonance energy transfer
between fluorophores separated by less than 10 nm and reveals interfluorophore
distance information from time-resolved fluorescence detection in combination
with photoswitching fingerprint analysis. We will show how the method can be
used advantageously to determine the number and distance even of
spatially unresolvable fluorophores in the sub-10 nm range. In combination with
genetic code expansion (GCE) with unnatural amino acids and bioorthogonal
click-labeling with small fluorophores photoswitching fingerprint analysis
should thus enable sub-10 nm resolution fluorescence imaging in cells.
Biosketch
Markus Sauer studied Chemistry at the University Heidelberg where he received his Diploma in 1991 and finished his PhD in 1995 in Physical Chemistry. 1998 he has been awarded the BioFuture Prize for Detection, Analysis and Handling of Single Molecules, which allowed him to establish his own group for single-molecule fluorescence detection and single-molecule DNA sequencing. Since 2009 he is Professor and Chair of the Department of Biotechnology and Biophysics at the Julius Maximilian University Würzburg. His research interests are single-molecule fluorescence spectroscopy and imaging with a particular focus on super-resolution fluorescence imaging by direct stochastic optical reconstruction microscopy (dSTORM) and its applications in neurobiology and immunology. 2021 he has been awarded a ERC Synergy Grant together with Silvio Rizzoli and Ed Boyden to work on Ultraresolution Imaging. He has published more than 270 journal papers and coordinates several super-resolution microscopy projects.
Website: www.super-resolution.de