Measuring localization performance of super-resolution algorithms on very active samples

Steve Wolter, Ulrike Endesfelder, Sebastian van de Linde, Mike Heilemann, Markus Sauer

Research output: Contribution to journalArticlepeer-review

58 Citations (Scopus)


Super-resolution fluorescence imaging based on singlemolecule 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.

Original languageEnglish
Pages (from-to)7020-7033
Number of pages14
JournalOptics Express
Issue number8
Publication statusPublished - 29 Mar 2011


  • localization
  • performance
  • super-resolution
  • algorithms
  • very active samples
  • fluorescence imaging
  • single-molecule localization
  • microscopy

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