TY  - JOUR
A1  - Wolter, Steve
A1  - Endesfelder, Ulrike
A1  - Linde, Sebastian van de
A1  - Heilemann, Mike
A1  - Sauer, Markus
T1  - Measuring localization performance of super-resolution algorithms on very active samples
JF  - Optics Express
N2  - Super-resolution fluorescence imaging based on  inglemolecule localization relies critically on the availability of efficient processing algorithms to distinguish, identify, and localize emissions of single fluorophores. In multiple current applications, such as threedimensional, time-resolved or cluster imaging, high densities of fluorophore emissions are common. Here, we provide an analytic tool to test the performance and quality of localization microscopy algorithms and demonstrate that common algorithms encounter difficulties for samples with high fluorophore density. We demonstrate that, for typical single-molecule localization microscopy methods such as dSTORM and the commonly used rapidSTORM scheme, computational precision limits the acceptable density of concurrently active fluorophores to 0.6 per square micrometer and that the number of successfully localized fluorophores per frame is limited to 0.2 per square micrometer.
Y1  - 2011
U6  - http://nbn-resolving.de/urn/resolver.pl?urn:nbn:de:bvb:20-opus-85936
ER  - 
TY  - JOUR
A1  - Vogel, Benjamin
A1  - Löschberger, Anna
A1  - Sauer, Markus
A1  - Hock, Robert
T1  - Cross-linking of DNA through HMGA1 suggests a DNA scaffold
N2  - Binding of proteins to DNA is usually considered 1D with one protein bound to one DNA molecule. In principle, proteins with multiple DNA binding domains could also bind to and thereby cross-link different DNA molecules. We have investigated this possibility using high-mobility group A1 (HMGA1) proteins, which are architectural elements of chromatin and are involved in the regulation of multiple DNA-dependent processes. Using direct stochastic optical reconstruction microscopy (dSTORM), we could show that overexpression of HMGA1a-eGFP in Cos-7 cells leads to chromatin aggregation. To investigate if HMGA1a is directly responsible for this chromatin compaction we developed a DNA cross-linking assay. We were able to show for the first time that HMGA1a can cross-link DNA directly. Detailed analysis using point mutated proteins revealed a novel DNA cross-linking domain. Electron microscopy indicates that HMGA1 proteins are able to create DNA loops and supercoils in linearized DNA confirming the cross-linking ability of HMGA1a. This capacity has profound implications for the spatial organization of DNA in the cell nucleus and suggests cross-linking activities for additional nuclear proteins.
KW  - DNA
Y1  - 2011
U6  - http://nbn-resolving.de/urn/resolver.pl?urn:nbn:de:bvb:20-opus-68865
ER  -