Abstract: A method for operating a point laser-scanning microscope includes scanning a sample with a focused illumination laser beam; recording a plurality of images by detecting elements being 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 an individual position n, or to a time-resolved mode, in which the recorded images are time-resolved images gi,jt(n, t), the collected photons being discriminated based on their arrival times to individual detecting elements; calculating a fingerprint image a by summing the plurality of intensity images gi,j(n) over all positions n; estimating shift matrices sx and sy from the intensity images gi,j(n); reconstructing at least one of a time-resolved object function ft and 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˜.

Abstract: Method for increasing the optical resolution of a stimulated emission depletion microscope, or STED microscope (Stimulated Emission Depletion), based on the modulation of the intensity of a STED beam on an arbitrary time scale during the acquisition of an image and the analysis of the induced dynamics, without increasing the intensity of the STED beam and in a simple and economic manner.

Abstract: A method for operating a point laser-scanning microscope comprises scanning a sample with a focused illumination laser beam; recording a plurality of images by detecting elements being 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 laser beam on an individual position n, or to a time-resolved mode, in which the recorded images are time-resolved images gi,jt(n,t), the collected photons being discriminated based on their arrival times to individual detecting elements; calculating a fingerprint image a by summing the plurality of intensity images gi,j(n) over all positions n; estimating shift matrices sx and sy from the intensity images gi,j(n); reconstructing at least one of a time-resolved object function ft and 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˜.

Abstract: Method for increasing the optical resolution of a stimulated emission depletion microscope, or STED microscope (Stimulated Emission Depletion), based on the modulation of the intensity of a STED beam on an arbitrary time scale during the acquisition of an image and the analysis of the induced dynamics, without increasing the intensity of the STED beam and in a simple and economic manner.

Abstract: Method of optical microscopy by scanning a sample containing an excitable species, the method comprising: directing a first and a second light beam onto respective, partially overlapped areas of the sample, wherein the first light beam is provided for exciting members of the excitable species, and the second light beam is provided for reducing the number of excited members; detecting an optical signal coming from the sample, comprising a main component and a spurious component, during consecutive first and second time gates, the first time gate being provided for detecting the optical signal for a time interval during which the main component and the spurious component are both present, and the second time gate being provided for detecting the optical signal for a time interval during which the main component tends to or is zero; processing the detected optical signal to separate its main component.

Abstract: In a STED fluorescence light microscope pulses of excitation light (3) are applied to a sample, which excite fluorescent entities contained in the sample for fluorescence, and which are focused on at least one focal area. Further, de-excitation light (12) is applied to the sample, which de-excites the excited fluorescent entities and which comprises an intensity zero point in the at least one focal area, as a continuous wave. Fluorescence light emitted by the excited fluorescent entities in the sample is registered after each pulse of the excitation light (3) and overlapping with applying the de-excitation light (13) with high temporal resolution between consecutive pulses of the excitation light (3).

Abstract: Method of optical microscopy by scanning a sample containing an excitable species, the method comprising: directing a first and a second light beam onto respective, partially overlapped areas of the sample, wherein the first light beam is provided for exciting members of the excitable species, and the second light beam is provided for reducing the number of excited members; detecting an optical signal coming from the sample, comprising a main component and a spurious component, during consecutive first and second time gates, the first time gate being provided for detecting the optical signal for a time interval during which the main component and the spurious component are both present, and the second time gate being provided for detecting the optical signal for a time interval during which the main component tends to or is zero; processing the detected optical signal to separate its main component.

Abstract: In a STED fluorescence light microscope pulses of excitation light (3) are applied to a sample, which excite fluorescent entities contained in the sample for fluorescence, and which are focused on at least one focal area. Further, de-excitation light (12) is applied to the sample, which de-excites the excited fluorescent entities and which comprises an intensity zero point in the at least one focal area, as a continuous wave. Fluorescence light emitted by the excited fluorescent entities in the sample is registered after each pulse of the excitation light (3) and overlapping with applying the de-excitation light (13) with high temporal resolution between consecutive pulses of the excitation light (3).