Patent Document ID: 10139611
Application ID: 16021674
Patent Status: 1

Claim One:
1. A method for high-resolution 3D-localization microscopy of a sample comprising fluorescence emitters, in which the fluorescence emitters in the sample are excited to emit fluorescent radiation and the sample is displayed with spatial resolution in wide-field microscopy, said excitation being caused such that in reference to the spatial resolution at least some fluorescence emitters are isolated, and in a localization analysis a three-dimensional localization is determined, which includes in a depth direction of a display a z-coordinate and a x-coordinate as well as a y-coordinate orthogonal in reference thereto for each isolated fluorescence emitter, showing a precision exceeding the spatial resolution, comprising: determining localization imprecision, which states an imprecision of the localization, namely at least regarding its z-coordinate as a function of the z-coordinate and a number of photons collected during imaging in the wide-field microscopy, and determining said localization imprecision for each localized fluorescence emitter by accessing table of localization imprecision for the localization determined during the localization analysis, with the localization imprecision in said table being determined by way of simulation comprising the following steps: a) simulating an image for an ideal, dot-shaped fluorescence emitter for the predetermined number of photons emitted by the fluorescence emitters for the image, for an average number of photons, b) modifying said simulated image with a pixel confusion function to form an expanded image, with the pixel confusion function dependent on a predetermined localization of the fluorescence emitter, c) modifying the expanded image in consideration of statistic noise influences to form a noisy, expanded image, with the statistic noise influences being dependent on the number of photons, d) subjecting the expanded image to a localization analysis, in order to obtain a simulated localization from the noisy, expanded image, and e) repeating steps b) through d) several times for various values of the statistic noise influences, a spread of the several simulated localizations obtained here, or a spread of differences between these several simulated localizations and a predetermined localization, is determined, and from said spread the imprecision of the predetermined localization is determined.