Fluctuation-based fluorescence microscopy

A microscope for fluctuation-based fluorescence microscopy of a sample, which, after excitation, emits fluorescence radiation in a blinking manner, the microscope comprising:

• an illumination beam path for exciting the sample to emit the fluorescence radiation,
• an imaging beam path for imaging the sample emitting the fluorescence radiation from a focal plane onto a spatially resolving detector, which comprises pixels, and for producing an image sequence of frames that differ in terms of a blinking state of the sample, and
• an evaluation device comprising a processor, the evaluation device configured for evaluating the image sequence and for producing a high-resolution sample image,
• wherein:
• the imaging beam path comprises an optical manipulator which optically influences the imaging of the sample emitting the fluorescence radiation in dependence on a parameter, wherein the parameter is a position with respect to the focal plane and/or a wavelength of the emitted fluorescence radiation, and the influencing causes point emitters in each frame to be imaged into imaged point emitter representations that in each case have at least two image lobes, the relative position of which depends on the parameter, and
• the evaluation device is configured for producing subframes for each frame on the basis of the influencing that is dependent on the parameter and caused by the optical manipulator and for specifying them with respect to the parameter, and for performing the evaluation of the image sequence of frames on the basis of the subframes.