@article{ProppertWolterHolmetal.2014, author = {Proppert, Sven and Wolter, Steve and Holm, Thorge and Klein, Theresa and van de Linde, Sebastian and Sauer, Markus}, title = {Cubic B-spline calibration for 3D super-resolution measurements using astigmatic imaging}, series = {Optics Express}, volume = {22}, journal = {Optics Express}, number = {9}, issn = {1094-4087}, doi = {10.1364/OE.22.010304}, url = {http://nbn-resolving.de/urn:nbn:de:bvb:20-opus-119730}, pages = {10304-16}, year = {2014}, abstract = {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.}, language = {en} } @article{WolterEndesfelderLindeetal.2011, author = {Wolter, Steve and Endesfelder, Ulrike and Linde, Sebastian van de and Heilemann, Mike and Sauer, Markus}, title = {Measuring localization performance of super-resolution algorithms on very active samples}, series = {Optics Express}, journal = {Optics Express}, url = {http://nbn-resolving.de/urn:nbn:de:bvb:20-opus-85936}, year = {2011}, abstract = {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.}, language = {en} }