Luminescence microscope for imaging a sample or for localizing or tracking emitters in a sample

A method for imaging a sample or for localizing emitters in a sample or tracking emitters in a sample by obtaining position estimates of an emitter moving in the sample at time intervals to obtain a trajectory of the emitter, the method being performed by a luminescence microscope comprising a first light source, at least one light modulator comprising a first active surface and a second active surface, an objective lens and a first beam displacement element, wherein an input light beam is generated by the first light source, wherein a first output light beam and/or a second output light beam is generated from the input light beam in a polarization-dependent manner by the first beam displacement element, such that if the input light beam has only one linearly polarized component that is parallel to a preferred direction of the first beam displacement element, only the first output light beam is formed, if the input light beam has only one linearly polarized component that is orthogonal to the preferred direction of the first beam displacement element, only the second output light beam is formed, and if the input light beam has components that are parallel and components that are orthogonal to the preferred direction of the at least one beam displacement element, the input light beam is split into the first output light beam and the second output light beam, wherein the first output light beam and the second output light beam form an angle of less than 90° with respect to each other when exiting the first beam displacement element, and wherein the first output light beam impinges on the first active surface of the at least one light modulator, wherein the first active surface modulates the first output light beam in its phase and/or in its amplitude, and/or wherein the second output light beam impinges on the second active surface of the at least one light modulator, wherein the second active surface modulates the second output light beam in its phase and/or in its amplitude, and wherein the first output light beam modulated in its phase and/or in its amplitude and/or the second output light beam modulated in its phase and/or in its amplitude are focused into the sample by the objective lens so that at least one light intensity distribution is formed in the sample.

A luminescence microscope for imaging a sample or for localizing emitters in a sample or tracking emitters in a sample, the tracking being performed by obtaining position estimates of an emitter moving in the sample at time intervals to obtain a trajectory of the emitter, the luminescence microscope comprising

• a first light source configured to generate an input light beam,
• at least one light modulator comprising a first active surface and a second active surface, the first active surface and the second active surface being configured to modulate the phase and/or the amplitude of light incident on the respective active surface,
• an objective lens configured to focus the light modulated by the light modulator in its phase and/or amplitude into the sample so that at least one light intensity distribution is formed in the sample,
• wherein the luminescence microscope comprises a first beam displacement element which is configured to generate, in a polarization-dependent manner, a first output light beam and/or a second output light beam from the input light beam, such that if the input light beam has only one linearly polarized component that is parallel to a preferred direction of the first beam displacement element, only the first output light beam is formed, if the input light beam has only one linearly polarized component that is orthogonal to the preferred direction of the first beam displacement element, only the second output light beam is formed, and if the input light beam has components that are parallel and components that are orthogonal to the preferred direction of the at least one beam displacement element, the input light beam is split into the first output light beam and the second output light beam, the first beam displacement element being arranged in such a way that the first output light beam impinges on the first active surface and the second output light beam impinges on the second active surface, wherein the first beam displacement element is configured such that the first output light beam and the second output light beam form an angle of less than 90° with respect to each other when exiting the first beam displacement element, wherein the first active surface and the second active surface modulate light incident on the respective active surface in its phase and/or in its amplitude in a polarization-dependent manner, such that the phase and/or the amplitude of light that is linearly polarized in a preferred direction of the at least one light modulator is modulated and the phase and/or the amplitude of light that is polarized orthogonally to the preferred direction is not modulated, wherein the first light source is configured to provide the input light beam as a combination of a first linearly polarized component and a second linearly polarized component, wherein the polarization direction of the first component is orthogonal to the polarization direction of the second component, wherein the luminescence microscope is configured so that light of the first component generated by the first light source, modulated in its phase and/or its amplitude by the at least one light modulator and focused by the objective lens and light of the second component modulated in its phase and/or its amplitude by the at least one light modulator and focused by the objective lens form a combined light intensity distribution in the sample.

A luminescence microscope for imaging a sample or for localizing emitters in a sample or tracking emitters in a sample, the tracking being performed by obtaining position estimates of an emitter moving in the sample at time intervals to obtain a trajectory of the emitter, the luminescence microscope comprising

• a first light source configured to generate an input light beam,
• at least one light modulator comprising a first active surface and a second active surface, the first active surface and the second active surface being configured to modulate the phase and/or the amplitude of light incident on the respective active surface,
• an objective lens configured to focus the light modulated by the light modulator in its phase and/or amplitude into the sample so that at least one light intensity distribution is formed in the sample,
• wherein the luminescence microscope comprises a first beam displacement element which is configured to generate, in a polarization-dependent manner, a first output light beam and/or a second output light beam from the input light beam, such that if the input light beam has only one linearly polarized component that is parallel to a preferred direction of the first beam displacement element, only the first output light beam is formed, if the input light beam has only one linearly polarized component that is orthogonal to the preferred direction of the first beam displacement element, only the second output light beam is formed, and if the input light beam has components that are parallel and components that are orthogonal to the preferred direction of the at least one beam displacement element, the input light beam is split into the first output light beam and the second output light beam, the first beam displacement element being arranged in such a way that the first output light beam impinges on the first active surface and the second output light beam impinges on the second active surface, wherein the first beam displacement element is configured such that the first output light beam and the second output light beam form an angle of less than 90° with respect to each other when exiting the first beam displacement element, wherein the first active surface and the second active surface modulate light incident on the respective active surface in its phase and/or in its amplitude in a polarization-dependent manner, such that the phase and/or the amplitude of light that is linearly polarized in a preferred direction of the at least one light modulator is modulated and the phase and/or the amplitude of light that is polarized orthogonally to the preferred direction is not modulated, wherein the luminescence microscope comprises an output beam combination unit which is configured to combine the first output light beam and the second output light beam into a combined output light beam, wherein the output beam combination unit is the first beam displacement element or a further second beam displacement element, wherein the luminescence microscope comprises a fourth polarization rotator arranged between the at least one light modulator and the output beam combination unit, the fourth polarization rotator being configured to rotate the polarization direction of the first output light beam or the second output light beam, so that the first output light beam and the second output light beam are combined by the output beam combination unit into the combined output light beam.

A luminescence microscope for imaging a sample or for localizing emitters in a sample or tracking emitters in a sample, the tracking being performed by obtaining position estimates of an emitter moving in the sample at time intervals to obtain a trajectory of the emitter, the luminescence microscope comprising

• a first light source configured to generate an input light beam,
• at least one light modulator comprising a first active surface and a second active surface, the first active surface and the second active surface being configured to modulate the phase and/or the amplitude of light incident on the respective active surface,
• an objective lens configured to focus the light modulated by the light modulator in its phase and/or amplitude into the sample so that at least one light intensity distribution is formed in the sample,
• wherein the luminescence microscope comprises a first beam displacement element which is configured to generate, in a polarization-dependent manner, a first output light beam and/or a second output light beam from the input light beam, such that if the input light beam has only one linearly polarized component that is parallel to a preferred direction of the first beam displacement element, only the first output light beam is formed, if the input light beam has only one linearly polarized component that is orthogonal to the preferred direction of the first beam displacement element, only the second output light beam is formed, and if the input light beam has components that are parallel and components that are orthogonal to the preferred direction of the at least one beam displacement element, the input light beam is split into the first output light beam and the second output light beam, the first beam displacement element being arranged in such a way that the first output light beam impinges on the first active surface and the second output light beam impinges on the second active surface, wherein the first beam displacement element is configured such that the first output light beam and the second output light beam form an angle of less than 90° with respect to each other when exiting the first beam displacement element, wherein the first active surface and the second active surface modulate light incident on the respective active surface in its phase and/or in its amplitude in a polarization-dependent manner, such that the phase and/or the amplitude of light that is linearly polarized in a preferred direction of the at least one light modulator is modulated and the phase and/or the amplitude of light that is polarized orthogonally to the preferred direction is not modulated, wherein the first light source is configured to generate illumination light which affects light emissions from emitters in the sample, a control unit of the luminescence microscope being configured to perform an illumination sequence with a plurality of illumination steps, wherein the sample is respectively illuminated with a light intensity distribution of the illumination light with a local minimum in the illumination steps, such that illumination positions in the sample are illuminated with different light intensities of the illumination light in the illumination steps, wherein the local minimum of the light intensity distribution is positioned in a region around a presumed position of an emitter in the sample in the illumination steps, and wherein the luminescence microscope comprises a detector and a computing unit, wherein the detector is configured to detect light emissions of the emitter for the respective illumination steps, and wherein the computing unit is configured to determine the position of the emitter in the sample from the light emissions detected for the respective illumination steps.