Carl Zeiss Microscopy Gmbh

Joerg Siebenmorgen

Thomas Kalkbrenner

Helmut Lippert

Ralf Netz

Ingo Kleppe

Ralf Wolleschensky

Thomas Ohrt

Tobias Kaufhold

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

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 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

A method for optically examining a plurality of microscopic samples. The samples are channeled one after the other by means of a flow into at least one flow channel in which the samples advance along a flow direction. The samples are illuminated, and light emitted from the samples is detected and analyzed. A device for carrying out the method in which samples are illuminated in that at least one light sheet with a light sheet plane is directed onto the at least one flow channel, wherein the light sheet is oriented so as to intersect the at least one flow channel in an intersection region, and the normal of the light sheet plane forms an angle differing from null together with the flow direction in the intersection region. The light emitted from the sample is registered by an imaging optical detection unit, and the focal plane of said optical detection unit lies in the intersection region.

Method and device for optically examining a plurality of microscopic samples

A light sheet microscope which includes an illumination apparatus generating coherent illumination light for several illumination wavelengths, a beam-shaping module generating a light sheet from illumination light, an illumination objective illuminating a specimen with the light sheet and a detection objective for imaging light which is emitted by the specimen onto a laminar detector, wherein the optical axes of the detection objective and of the illumination objective are not parallel to each other. In such a light sheet microscope, the beam-shaping module includes a phase-selective element with several selection areas separated from each other spatially, wherein in each case one selection area is assigned to one specific illumination wavelength, and wherein a phase distribution predefined for the respective illumination wavelength is impressed on each selection area. The beam-shaping module further includes means for the sequential or simultaneous selection of the selection areas in dependence on the respective illumination wavelength.

Light sheet microscope with a phase-selective element for illumination

A device and a method for imaging a sample arranged in an object plane. The device includes an optical relay system that images an area of the sample from the object plane into an intermediate image plane. The device may also include an optical imaging system with an objective having an optical axis that lies perpendicularly on the intermediate image plan, and which is focused on the intermediate image plane, with the result that the object plane can be imaged undistorted onto a detector. The device also can include an illumination apparatus for illuminating the sample with a light sheet, wherein the light sheet lies essentially in the object plane and defines an illumination direction, and wherein the normal of the object plane defines a detection direction.

Device for imaging a sample with detection of an asymmetrically distributed angular range

A method for examining a specimen via light sheet microscopy includes selecting several illumination wavelengths for the specimen. To structure the illumination light, a predefined phase distribution is impressed on the phase-selective element and a predefined aperture structure is impressed on an aperture in the aperture plane. The phase-selective element is then illuminated in an intermediate image plane in an illumination beam path with illumination light, which is structured by the phase-selective element. The structured illumination light is imaged into an aperture plane arranged downstream of the phase-selective element. The aperture structure is adapted such that the zero orders of the structured illumination light in the aperture plane are substantially removed. The specimen is illuminated with the structured light sheet in the light sheet plane. Light emitted by the specimen is detected in a detection direction which forms an angle different from zero with the light sheet plane.

Growing community of inventors

Jena, Germany

Joerg Siebenmorgen