Carl Zeiss Microscopy Gmbh

Carl Zeiss Microimaging Gmbh

Lionix International Bv

Ralf Netz

Ingo Kleppe

Yauheni Novikau

Thomas Kalkbrenner

Christoph Nieten

Jörg Siebenmorgen

Gerhard Krampert

Joerg Siebenmorgen

Ralf Wolleschensky

Helmut Lippert

Tiemo Anhut

Wolfgang Bathe

Michael Goelles

Kai Wicker

Frank Hecht

Matthias Langhorst

Joerg-Michael Funk

Daniel Schwedt

Michael Kempe

Ingo Fahlbusch

Joerg Steinert

Ronald Dekker

Nils Langholz

Michael Kieweg

Werner Kleinschmidt

Edwin Jan Klein

Michael Brehm

Michael Gölles

Jörg Steinert

Wibke Hellmich

Gunther Lorenz

Stephan Wagner-Conrad

Klaus Weisshart

Douwe Harmen Geuzebroek

Marcus Heidkamp

An apparatus and method for structured illumination microscopy, having an illumination beam path for irradiating a sample with excitation light with a two-dimensional illumination pattern at angles greater than the angle for total internal reflection. The illumination beam path has an illumination objective used to irradiate the sample. A first separation device for separating the excitation light in a first linear coordinate direction in a pupil plane and a displacement device for laterally displacing the illumination pattern in a sample plane, having a detection beam path containing at least one microscope objective for guiding emission light to a camera. The emission light is emitted by the sample as a consequence of the irradiation by the excitation light. A camera for recording images of the sample, has a control unit for calculating microscopic images of the sample using partial images of the sample recorded for different positions of the illumination pattern in the sample plane. The apparatus has an adjustable second separation device for variably separating the excitation light in a second linear coordinate direction which is independent of the first linear coordinate direction, this variable separation being independent of the separation in the first linear coordinate direction in the pupil plane.

Apparatus for structured illumination microscopy, method for illuminating a sample and method of structured illumination microscopy

In a light sheet microscope, a detection device images a sample volume and an illumination device illuminates the sample with a light sheet. A light sheet plane is arranged substantially perpendicularly to the optical imaging axis and an illumination width extends perpendicular to the light sheet plane. A processing device is provided to control setting of the location of the light sheet plane in the sample volume, and recording a plurality of images of the sample volume with different locations of the light sheet plane. For the plurality of images recorded, a spacing, which is measured along the optical imaging axis, between adjacent locations of the light sheet plane is greater than or equal to half the illumination width, but not greater than the illumination width. Intermediate images for the stack of images in additional locations of the light sheet plane in the sample volume are computed.

Light sheet microscope and method for light sheet microscopy

An illumination apparatus for illuminating a sample plane and a sample that is optionally arranged therein along an illumination beam path includes: a first light source (for outputting light of at least one first wavelength (λ), a second light source for outputting light of at least one second wavelength (λ), and a diffraction grating in the illumination beam path between the first and second light sources and the sample plane. Light of the first wavelength (λ) is not diffracted by the diffraction grating, and light of the second wavelength (λ) is diffracted due to the effect of the diffraction grating. The illumination apparatus can be used in a microscope.

Method and device for illuminating a sample plane

A light beam shaping arrangement for a light microscope has a first and a second liquid crystal region or lifting micromirror region, each of which has a plurality of independently switchable liquid crystal elements or mirrors with which a phase of incident light is changeable in a settable manner, an input-/output-coupling polarization beam splitter, a polarization beam splitter arranged between the input-/output-coupling polarization beam splitter and the liquid crystal regions or lifting micromirror regions such that the polarization beam splitter separates the light coming from the input-/output-coupling polarization beam splitter in a polarization-dependent manner into a first partial beam. The first partial beam is directed to the first liquid crystal region or lifting micromirror region, and into a second partial beam, which is directed to the second liquid crystal region or lifting micromirror region, and the two partial beams returning from the liquid crystal regions and directs them together as an outgoing light beam.

Optical arrangement and method for light beam shaping for a light microscope

A method for performing SIM microscopy on a sample, includes: generating n raw images of the sample, in each case by illuminating the sample using the same SIM illumination pattern albeit with an individual positioning for each raw image, wherein p orders of diffraction are assigned to the SIM illumination pattern, and generating an image of the sample from the n raw images. An image reconstruction is carried out using the orders of diffraction, wherein t highest orders of diffraction are suppressed during the image reconstruction and n=p−t applies.

Method for SIM microscopy

An optics arrangement for flexible multi-color illumination for a light microscope includes an acousto-optical tunable filter ('AOTF'). The AOTF is set up to diffract two light components from incident illumination light into different order-of-diffraction directions. The two light components differ in their wavelengths and polarizations. Alternatively, an electro-optical modulator ('EOM') can be used, with which two temporally successive light components of different wavelengths are set to different polarization directions. A polarization beam splitter separates the two light components of different wavelengths and polarizations into reflection light, which is reflected at the polarization beam splitter, and transmission light, which is transmitted at the polarization beam splitter. A light structuring apparatus imprints different structures onto the transmission light and the reflection light. The structured transmission light and the structured reflection light are then recombined by the polarization beam splitter or a further polarization beam splitter onto a common beam path.

Optics arrangement for flexible multi-color illumination for a light microscope and method to this end

A single plane illumination microscope having an illumination optical system for illuminating a sample located on a sample carrier in a medium, and which is parallel to a planar reference surface. The sample is illuminated by a light sheet via an illumination light path. A detection optical system has a detection beam path. The optical axes of the illumination and detection optical systems each define an angle that is not equal to zero degrees along with the normal to the reference surface. A barrier layer system includes at least one layer of a given material having a given thickness and separates the medium from the illumination and detection optical systems. A base area of the barrier layer system is in contact with the region that is accessible for illumination and detection activities, said base area running parallel to the reference surface. At least one corrective element in the illumination beam path and/or the detection beam path allows those aberrations to be reduced which are created when light to be detected or light for illuminating the sample penetrates interfaces of the barrier layer system at an angle. The microscope has means, which are independent of the generation of the light sheet, for applying, via at least one manipulation beam path, light intensity to the sample in substantially point-shaped regions of the light sheet plane or in a given volume that at least temporarily encompasses the light sheet plane.

Single plane illumination microscope

A method for operating a light microscope with structured illumination includes: providing illumination patterns by means of a structuring device which splits impinging light into at least three coherent beam parts which correspond to a &#x2212;1., 0., and +1. diffraction orders of light; generating different phases of the illumination patterns by setting different phase values for the beam parts with phase shifters; and recording at least one microscope image for each of the illumination patterns and calculating a high resolution image from the microscope images. Phase shifters are provided not only for the beam parts of the &#x2212;1. and +1. diffraction orders but also at least one phase shifter for the beam part of the 0. diffraction order. At least two different phase values &#x3a6;are set with the at least one phase shifter for the 0. diffraction order to provide a plurality of illumination patterns with different phases.

Method for operating a light microscope with structured illumination and optic arrangement

In high-resolution scanning microscopy, a sample is excited by illumination radiation to emit fluorescence radiation in such a way that the illumination radiation is focused at a point in or on the sample to form a diffraction-limited illumination spot. The point is imaged in a diffraction-limited manner into a diffraction image on a spatially resolving surface detector, wherein the surface detector has a spatial resolution that resolves a structure of the diffraction image. The sample is scanned by means of different scanning positions with an increment of less than half the diameter of the illumination spot. An image of the sample is generated from the data of the surface detector and from the scanning positions assigned to said data, said image having a resolution that is increased beyond a resolution limit for imaging. For the purposes of distinguishing between at least two predetermined wavelength regions in the fluorescence radiation from the sample, a corresponding number of diffraction structures are generated on the surface detector for the at least two predetermined wavelength ranges, said diffraction structures differing but having a common center of symmetry. The diffraction structures are evaluated when generating the image of the sample.

High-resolution scanning microscopy with discrimination between at least two wave-length ranges

A method for illuminating samples in microscopic imaging methods, wherein a number m of different wavelengths &#x3bb;, with m&#x3e;I and i=I, . . . , m, is selected for the illumination. For each of the wavelengths &#x3bb;a target phase function &#x394;&#x3c6;(x, y, &#x3bb;) is predefined, wherein x and y denote spatial coordinates in a plane perpendicular to an optical axis z and each target phase function &#x394;&#x3c6;(x, y, &#x3bb;) is effective only for the corresponding wavelength &#x3bb;. The target phase functions &#x394;&#x3c6;are predefined depending on the structure of the sample and/or the beam shape and/or illumination light structure to be impressed on the light used for illumination. A total phase mask is then produced which realises all target phase functions &#x394;&#x3c6;(x, y, &#x3bb;). This total phase mask is then illuminated simultaneously or successively with coherent light of wavelengths &#x3bb;such that the predefined structure of the illumination light is generated in the region of the sample.

Method for illuminating samples in microscopic imaging methods

An optical arrangement for light beam shaping in a light microscope has a first and a second liquid crystal region, each of which has a plurality of independently switchable liquid crystal elements with which a phase of incident light is changeable in a settable manner. A first polarization beam splitter is arranged in such a way that incident light is split in a polarization-dependent manner into reflection light, which is reflected in the direction of the first liquid crystal region, and transmission light, which is transmitted in the direction of the second liquid crystal region. The first or a second polarization beam splitter is arranged such that the reflection light and transmission light are combined onto a common beam path after phase modulation by means of the liquid crystal regions.

An arrangement for increasing resolution of a laser scanning microscope has a simplified adjustment and lower susceptibility to errors. The pupil beam from the laser scanning microscope is coupled into a shortened common path interferometer, to make wavefronts of a pupil image mirrored at at least one axis and wavefronts of an unchanged pupil image interfere. The area of a pupil from the pupil beam is split into two complementary portions P and Q producing two partial beams separately supplied to at least one beam deflection means by total-internal reflection along the common path interferometer. The light of the interferometer branches from transmitted light of the one interferometer branch and reflected light of the other interferometer branch is made to interfere at a partly transmissive beam splitter layer to cause constructive interference C and destructive interference D of the wavefronts from the two different portions P and Q of the pupil.

Assembly for increasing the resolution of a laser scanning microscope

A light microscope comprises: a structuring optical unit comprising a waveguide chip for providing a structured illumination; an input selection device for variably directing light to one of several inputs of the waveguide chip; the waveguide chip further comprising a light guide path following each of the inputs; each light guide path divides into several path divisions; and each path division leads to one output of the wave-guide chip. The outputs of the waveguide chip can be arranged at a pupil plane of the light microscope, and an exit direction of light from the outputs is transverse to a plane defined by the waveguide chip. A method for providing structured illumination light using the light microscope is also described.

Light microscope and method for providing structured illumination light

A microscope and method for high resolution scanning microscopy of a sample, having an illumination device, an imaging device for the purpose of scanning at least one point or linear spot across the sample and of imaging the point or linear spot into a diffraction-limited, static single image below a reproduction scale in a detection plane. A detector device is used for detecting the single image in the detection plane for various scan positions, with a location accuracy which, taking into account the reproduction scale in at least one dimension/measurement, is at least twice as high as a full width at half maximum of the diffraction-limited single image.

High-resolution scanning microscopy

An arrangement for TIRF microscopy, having an illumination optical unit with an illumination objective for illuminating a specimen on a specimen carrier in a specimen plane via an illumination beam path. An optical axis of the illumination objective includes an illumination angle that differs from zero with the normal of the specimen plane. A detection optical unit with a detection objective in a detection beam path includes a detection angle that differs from zero between an optical axis thereof and the normal of the specimen plane. A transition element between the specimen carrier and both objectives is arranged both in the illumination beam path and in the detection beam path. The transition element corrects aberrations that arise on account of the passage through media with different refractive indices of radiation to be detected and/or radiation for illuminating the specimen.

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Jena, Germany

Ralf Netz