TIME-RESOLVED IMAGING METHOD WITH HIGH SPATIAL RESOLUTION

Method for operating a point laser- scanning microscope, comprising scanning a sample with a focused illumination laser beam (EB); recording, by means of an array of detecting elements (23), a plurality of images of the sample over a scan by the laser beam, wherein the detecting elements (23) are configurable to an intensity mode, in which the recorded images are intensity images gi,j(n) related to photons collected during an entire dwell time of the illumination beam on the individual position n, or in a time-resolved mode, in which the recorded images are time -resolved images gt ij(n, t), in which the collected photons are discriminated based on their times of arrival to the individual detecting elements (23); calculating a fingerprint image a by summing the plurality of intensity images gi,j(n) over all positions n of the illumination beam (EB) on the sample, the fingerprint image depending simultaneously on an illumination point-spread function, hereinafter illumination PSF, hexc, and a detection point-spread function, hereinafter detection PSF, hdet, estimating shift matrices sx and sy from the intensity images gi,j(n), reconstructing at least one of: i) a time-resolved object function ft, and ii) an intensity object function f, and, visualizing at least one of a high-resolution time-resolved image ft~ and a high- resolution intensity image f~, based on said time -resolved object function and intensity object function.