Abstract: For forming and shifting a light intensity distribution in a focal area of an objective lens, portions of coherent input light are one by one directed into non-identical two-dimensional pupil areas of a pupil of the objective lens. Each of the portions of coherent input light is collimated in the pupil. The pupil areas include a pair of two pupil areas which are axially symmetrically arranged on opposite sides of an optical axis of the objective lens. At least one of the two discrete portions of coherent input light that are directed into the pair of pupil areas is separately modulated with regard to its phase by means of an electro optical modulator such as to form the light intensity distribution in the focal area with a local intensity minimum delimited by intensity maxima and to shift the local intensity minimum laterally with regard to the optical axis.

Abstract: Upstream a microscope objective lens, a polarization direction of a light beam is tilted with a first electro-optical deflector between a first polarization direction with which the light beam is deflected by a first polarization beam splitter by a first angle and a second polarization direction with which it is deflected by a second angle. With a second electro-optical deflector, the polarization direction of the light beam is tilted between a third polarization direction with which the light beam is deflected by a second polarization beam splitter by a third angle and a fourth polarization direction with which it is deflected by a fourth angle. By rotating the polarization direction of the light beam by means of the first and second electro-optical deflectors in a coordinated way the light beam is tilted about a fixed point in a pupil of the objective lens.

Abstract: A segmented birefringent chromatic beam shaping device comprises at least three birefringent chromatic segments arranged side by side in a pupil of the beam shaping device. The birefringent chromatic segments are essentially n? waveplates at a first design wavelength. At a second design wavelength, the birefringent chromatic segments are essentially (m+1/2)? waveplates. Each of the birefringent chromatic segments comprises a stack of birefringent elements including at least three chromatic birefringent elements. Orientations of fast axes of each pair of directly consecutive chromatic birefringent elements of each of the birefringent chromatic segments differ by at least 5 deg. The at least three birefringent chromatic segments comprise same sequences of materials, thicknesses and orientations of their birefringent elements so that they only differ is their orientations in the pupil.

Abstract: In a method of detecting changes in direction of a collimated coherent light beam, the light beam is split into partial light beams which are superimposed on a camera to form an interference pattern displaying light intensity minima and maxima alternatingly following to one another in a transverse direction oriented transversely to an average propagation direction of the partial beams. The light beam is focused into at least one focus located in front of the camera. Pictures of the interference pattern including a plurality of the light intensity maxima are registered with the camera. An average shift of the plurality of light intensity maxima with regard to the camera in the at least one transverse direction is determined from the pictures. A change in angular orientation of the collimated coherent light beam in the at least one transvers direction is deduced from the average shift.

Abstract: For forming and shifting a light intensity distribution in a focal area of an objective lens, portions of coherent input light are one by one directed into non-identical two-dimensional pupil areas of a pupil of the objective lens. Each of the portions of coherent input light is collimated in the pupil. The pupil areas include a pair of two pupil areas which are axially symmetrically arranged on opposite sides of an optical axis of the objective lens. At least one of the two discrete portions of coherent input light that are directed into the pair of pupil areas is separately modulated with regard to its phase by means of an electro optical modulator such as to form the light intensity distribution in the focal area with a local intensity minimum delimited by intensity maxima and to shift the local intensity minimum laterally with regard to the optical axis.

Abstract: A fluorescence microscope instrument includes a light source; an objective focusing light from the light source into a sample and collecting fluorescence light emitted out of the sample; a detector detecting the fluorescence light; and a beam path separator arranged in a first beam path between the light source and the objective and in a second beam path between the objective and the detector. Wavelengths of the light to be directed to the sample and of the fluorescence light to be detected by the detector fall into an extended range of wavelengths. The beam path separator separates the two beam paths in that it is transferable between being transparent for any wavelength of the range of wavelengths and coming along the first beam path, and a second state in which it is transparent for any wavelength of the range of wavelengths and coming along the second beam path.

Abstract: Upstream a microscope objective lens, a polarization direction of a light beam is tilted with a first electro-optical deflector between a first polarization direction with which the light beam is deflected by a first polarization beam splitter by a first angle and a second polarization direction with which it is deflected by a second angle. With a second electro-optical deflector, the polarization direction of the light beam is tilted between a third polarization direction with which the light beam is deflected by a second polarization beam splitter by a third angle and a fourth polarization direction with which it is deflected by a fourth angle. By rotating the polarization direction of the light beam by means of the first and second electro-optical deflectors in a coordinated way the light beam is tilted about a fixed point in a pupil of the objective lens.

Abstract: A segmented chromatic phase plate includes at least three stacks. The three stacks are arranged around and stacked along a main axis, and have same overall heights. Each stack includes at least two plane-parallel optical flats of different materials. The materials and the thicknesses of the optical flats are selected such that optical path lengths of light of a first wavelength passing through the different stacks along the main axis differ by integer multiples of the first wavelength, whereas optical path lengths of light of a second wavelength differ by integer multiples plus defined fractions of the second wavelength. The materials of the at least two optical flats have different refractive indices for both the first and second wavelengths; and derivatives of the differences between the optical path lengths with respect to wavelength are zero at at least one of the first and second wavelengths.

Abstract: A segmented birefringent chromatic beam shaping device comprises at least three birefringent chromatic segments arranged side by side in a pupil of the beam shaping device. The birefringent chromatic segments are essentially n? waveplates at a first design wavelength. At a second design wavelength, the birefringent chromatic segments are essentially (m+½)? waveplates. Each of the birefringent chromatic segments comprises a stack of birefringent elements including at least three chromatic birefringent elements. Orientations of fast axes of each pair of directly consecutive chromatic birefringent elements of each of the birefringent chromatic segments differ by at least 5 deg. The at least three birefringent chromatic segments comprise same sequences of materials, thicknesses and orientations of their birefringent elements so that they only differ is their orientations in the pupil.

Abstract: In a method of monitoring a relative position of a microscope objective with regard to a sample a test beam of light is directed onto at least one at least partially reflective surface connected to the sample, and light of the test beam reflected at the at least one at least partially reflective surface is registered and evaluated. Additionally, the test beam is directed onto a reflective surface of the microscope objective facing the sample, and light of the test beam reflected at the reflective surface of the microscope objective is also registered and evaluated.

Abstract: In a method of monitoring a relative position of a microscope objective with regard to a sample a test beam of light is directed onto at least one at least partially reflective surface connected to the sample, and light of the test beam reflected at the at least one at least partially reflective surface is registered and evaluated. Additionally, the test beam is directed onto a reflective surface of the microscope objective facing the sample, and light of the test beam reflected at the reflective surface of the microscope objective is also registered and evaluated.

Abstract: A segmented chromatic phase plate comprises at least three stacks. The three stacks are arranged around and stacked along a main axis, and have same overall heights. Each stack comprises at least two plane-parallel optical flats of different materials. The materials and the thicknesses of the optical flats are selected such that optical path lengths of light of a first wavelength passing through the different stacks along the main axis differ by integer multiples of the first wavelength, whereas optical path lengths of light of a second wavelength differ by integer multiples plus defined fractions of the second wavelength. The materials of the at least two optical flats have different refractive indices for both the first and second wavelengths; and derivatives of the differences between the optical path lengths with respect to wavelength are zero at at least one of the first and second wavelengths.

Abstract: A fluorescence microscope instrument includes a light source; an objective focusing light from the light source into a sample and collecting fluorescence light emitted out of the sample; a detector detecting the fluorescence light; and a beam path separator arranged in a first beam path between the light source and the objective and in a second beam path between the objective and the detector. Wavelengths of the light to be directed to the sample and of the fluorescence light to be detected by the detector fall into an extended range of wavelengths. The beam path separator separates the two beam paths in that it is transferable between being transparent for any wavelength of the range of wavelengths and coming along the first beam path, and a second state in which it is transparent for any wavelength of the range of wavelengths and coming along the second beam path.

Abstract: An apparatus forming an area of minimum light intensity enclosed by high light intensity includes two objectives focusing light of opposite directions into a common focal area. Light intensities of a first pair of light beams extinguish each other in a first partial area of the focal area. Beam paths of the first pair of beams pass through the objectives in a first pair of partial areas of the pupils of the objectives. Light intensities of a second pair of light beams extinguish each other in a second partial area of the focal area. Beam paths of the second pair of light beams pass through the objectives in a second pair of partial areas which are offset with regard to the first pair of partial areas of the pupils; and the light of the second pair does not interfere with the light of the first pair of light beams.

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: An apparatus forming an area of minimum light intensity enclosed by high light intensity includes two objectives focusing light of opposite directions into a common focal area. Light intensities of a first pair of light beams extinguish each other in a first partial area of the focal area. Beam paths of the first pair of beams pass through the objectives in a first pair of partial areas of the pupils of the objectives. Light intensities of a second pair of light beams extinguish each other in a second partial area of the focal area. Beam paths of the second pair of light beams pass through the objectives in a second pair of partial areas which are offset with regard to the first pair of partial areas of the pupils; and the light of the second pair does not interfere with the light of the first pair of light beams.

Abstract: In a method for imaging a structure (33) marked with a fluorescent dye in a sample, the sample is repeatedly scanned in a scanning range (28) with a light intensity distribution localised around a focal point (29) of a focused fluorescence excitation light beam. The light intensity distribution further comprises a focused fluorescence inhibiting light beam (7) whose wave fronts are modulated so that a fluorescence inhibiting light intensity distribution comprises a minimum at the focal point (29) of the fluorescence excitation light beam (4). The scanning conditions are coordinated in such a way that the fluorescence light is emitted out of the scanning range (28) as individually detectable photons. When these photons are detected, the location (32) of the focal point (28) at the respective point in time is allocated to them. An image of the structure (33) is composed of the locations (32) to which the detected photons have been allocated during several repetitions of scanning the scanning range (28).

Abstract: A fluorescence light scanning microscope (2) comprises a light source providing excitation light (8) for exciting a fluorophore in a sample to be imaged for spontaneous emission of fluorescence light, and suppression light (7) for suppressing spontaneous emission of fluorescence light by the fluorophore on a common optical axis (4), the suppression wavelength differing from the excitation wavelength; an objective (19) focusing both the excitation (8) and the suppression (7) light to a focus point; a detector (21) detecting fluorescence light (11) spontaneously emitted by the fluorophore; and a chromatic beam shaping device (1) arranged on the common optical axis (4), and including a birefringent chromatic optical element (3) adapted to shape a polarization distribution of the suppression light (7) such as to produce an intensity zero at the focus point, and to leave the excitation light such as to produce a maximum at the focus point.

Abstract: To the end of three-dimensionally localizing light emitting marker entities of unknown orientation and unknown position in a sample, the light emitted by each single marker entity is imaged in at least two different ways onto at least one detection plane which corresponds to a focal plane (13) in the sample resulting in at least two images of the marker entity. Virtual x- and y-positions of the marker entity in parallel to the focal plane (13) are separately determined from the emitted light intensity distribution over each image of the marker entity. Further, the z-position of the marker entity normal to the focal plane is determined from the emitted light intensity distributions over the images of the marker entity. The real x- and y-positions of the marker entity in parallel to the focal plane (13) are determined based on its virtual x- and y-positions and on its z-position.

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).