TY  - JOUR
A1  - Kunz, Tobias C.
A1  - Götz, Ralph
A1  - Sauer, Markus
A1  - Rudel, Thomas
T1  - Detection of chlamydia developmental forms and secreted effectors by expansion microscopy
JF  - Frontiers in Cellular and Infection Microbiology
N2  - Expansion microscopy (ExM) is a novel tool to improve the resolution of fluorescence-based microscopy that has not yet been used to visualize intracellular pathogens. Here we show the expansion of the intracellular pathogen Chlamydia trachomatis, enabling to differentiate its two distinct forms, catabolic active reticulate bodies (RB) and infectious elementary bodies (EB), on a conventional confocal microscope. We show that ExM enables the possibility to precisely locate chlamydial effector proteins, such as CPAF or Cdu1, within and outside of the chlamydial inclusion. Thus, we claim that ExM offers the possibility to address a broad range of questions and may be useful for further research on various intracellular pathogens.
KW  - expansion microscopy
KW  - chlamydia
KW  - secreted effectors
KW  - developmental forms
KW  - superresolution
KW  - imaging
Y1  - 2019
U6  - http://nbn-resolving.de/urn/resolver.pl?urn:nbn:de:bvb:20-opus-195716
SN  - 2235-2988
VL  - 9
IS  - 276
ER  - 
TY  - JOUR
A1  - Lando, David
A1  - Endesfelder, Ulrike
A1  - Berger, Harald
A1  - Subramanian, Lakxmi
A1  - Dunne, Paul D.
A1  - McColl, James
A1  - Klenerman, David
A1  - Carr, Antony M.
A1  - Sauer, Markus
A1  - Allshire, Robin C.
A1  - Heilemann, Mike
A1  - Laue, Ernest D.
T1  - Quantitative single-molecule microscopy reveals that CENP-A\(^{Cnp1}\) deposition occurs during G2 in fission yeast
JF  - Open Biology
N2  - The inheritance of the histone H3 variant CENP-A in nucleosomes at centromeres following DNA replication is mediated by an epigenetic mechanism. To understand the process of epigenetic inheritance, or propagation of histones and histone variants, as nucleosomes are disassembled and reassembled in living eukaryotic cells, we have explored the feasibility of exploiting photo-activated localization microscopy (PALM). PALM of single molecules in living cells has the potential to reveal new concepts in cell biology, providing insights into stochastic variation in cellular states. However, thus far, its use has been limited to studies in bacteria or to processes occurring near the surface of eukaryotic cells. With PALM, one literally observes and 'counts' individual molecules in cells one-by-one and this allows the recording of images with a resolution higher than that determined by the diffraction of light (the so-called super-resolution microscopy). Here, we investigate the use of different fluorophores and develop procedures to count the centromere-specific histone H3 variant CENP-A\(^{Cnp1}\) with single-molecule sensitivity in fission yeast (Schizosaccharomyces pombe). The results obtained are validated by and compared with ChIP-seq analyses. Using this approach, CENP-A\(^{Cnp1}\) levels at fission yeast (S. pombe) centromeres were followed as they change during the cell cycle. Our measurements show that CENP-A(Cnp1) is deposited solely during the G2 phase of the cell cycle.
KW  - nucleosome
KW  - fission yeast
KW  - identification
KW  - propagation
KW  - CSE4, CENP-A
KW  - CENP-A
KW  - schizosaccaromyces-pombe
KW  - fluorescent protein
KW  - centomeres
KW  - superresolution
KW  - chromatin
KW  - centromere
KW  - ingle-molecule microscopy
Y1  - 2012
U6  - http://nbn-resolving.de/urn/resolver.pl?urn:nbn:de:bvb:20-opus-134682
VL  - 2
IS  - 120078
ER  - 
TY  - JOUR
A1  - Proppert, Sven
A1  - Wolter, Steve
A1  - Holm, Thorge
A1  - Klein, Theresa
A1  - van de Linde, Sebastian
A1  - Sauer, Markus
T1  - Cubic B-spline calibration for 3D super-resolution measurements using astigmatic imaging
JF  - Optics Express
N2  - In recent years three-dimensional (3D) super-resolution fluorescence imaging by single-molecule localization (localization microscopy) has gained considerable interest because of its simple implementation and high optical resolution. Astigmatic and biplane imaging are experimentally simple methods to engineer a 3D-specific point spread function (PSF), but existing evaluation methods have proven problematic in practical application. Here we introduce the use of cubic B-splines to model the relationship of axial position and PSF width in the above mentioned approaches and compare the performance with existing methods. We show that cubic B-splines are the first method that can combine precision, accuracy and simplicity.
KW  - three-dimensional microscopy
KW  - fluorescence microscopy
KW  - medical and biological imaging
KW  - superresolution
Y1  - 2014
U6  - http://nbn-resolving.de/urn/resolver.pl?urn:nbn:de:bvb:20-opus-119730
SN  - 1094-4087
VL  - 22
IS  - 9
ER  -