The disclosed technology brings histopathology into the operating theatre, to enable real-time intra-operative digital pathology. The disclosed technology utilizes confocal imaging devices image, in the operating theatre, “optical slices” of fresh tissue—without the need to physically slice and otherwise process the resected tissue as required by frozen section analysis (FSA). The disclosed technology, in certain embodiments, includes a simple, operating-table-side digital histology scanner, with the capability of rapidly scanning all outer margins of a tissue sample (e.g., resection lump, removed tissue mass). Using point-scanning microscopy technology, the disclosed technology, in certain embodiments, precisely scans a thin “optical section” of the resected tissue, and sends the digital image to a pathologist rather than the real tissue, thereby providing the pathologist with the opportunity to analyze the tissue intra-operatively. Thus, the disclosed technology provides digital images with similar information content as FSA, but faster and without destroying the tissue sample itself.

An optical microscope according to one aspect of the present disclosure includes: a light source; a first scanner to scan a spot position of a light beam on a sample; an objective lens to focus the light beam deflected by the first scanner and cause the light beam to be made incident on the sample; a spectroscope including a slit on an incident side which an outgoing light emitted from an area on the sample onto which the light beam has been illuminated enters; a detector configured to detect an outgoing light from the spectroscope; and a first relay optical system including a first off-axis parabolic mirror that is arranged in an optical path from the first scanner to the objective lens and reflects the light beam deflected by the first scanner and a second off-axis parabolic mirror that reflects the light beam reflected in the first off-axis parabolic mirror.

According to one method, a sample with cells contains a phototagging agent. The sample is imaged to identify at least one target cell to be isolated. The identified target cell in the sample is selectively irradiated with photo-activating light for selectively activating the phototagging agent in the target cell to change its fluorescence response. The irradiated target cell is isolated from other cells in the sample based on a difference in its fluorescence response compared to non-activated phototagging agent in the other cells. Further aspects are directed to a corresponding microscope system and chemical compound for use as the phototagging agent.

An optical microscope device (10) according to the present disclosure includes: a first illumination optical system (13A) including a light source (101) that emits illumination light for illuminating a specimen including biomaterials changing their functions in response to light, an LCOS spatial light modulation element (113) that controls a polarization state of the illumination light, a first illumination optical member that uniformly illuminates the LCOS spatial light modulation element, and a polarization optical element that controls a transmission state of the illumination light directed to the specimen from the LCOS spatial light modulation element in response to the polarization state of the illumination light; a second illumination optical system (13B) including a second illumination optical member that images a light flux from the first illumination optical system (13A) on a specimen surface; and an imaging optical system (13C) for imaging the specimen surface, the imaging optical including an imaging optical member and an imaging element (127).

A Multi-Z confocal microscopy system can simultaneously record from multiple Z-sections, and thus performs high speed volumetric imaging. An illumination line can be formed by under-filling the illumination beam in the aperture of the microscope objective. The illumination line extends in the Z dimension into the target sample to be imaged and an X-Y scanning mechanism can be used to scan the illumination line over the sample. The detection signal emanating from the scanned sample can be collected through the full numerical aperture of the microscope objective and directed to a detector subsystem. The detector subsystem includes an array of reflecting pinhole detectors and each reflecting pinhole detector is configured to image a volume at a different depth in the sample. This configuration enables reflecting pinhole detector array to image more than one depth volume at the same time.

The disclosure relates to a method for scanning microscopy wherein a specimen is scanned simultaneously with a plurality of illumination spots of an excitation light. The light emitted by one specimen location irradiated with one illumination spot is detected independently of the light emitted by another specimen location illuminated with another illumination spot. A microscopic image of the specimen can be compiled from the emitted light detected for the different specimen locations. The method provides that the intensities of the different illumination spots are set independently of one another, and in that the illumination spots are guided over the specimen one after another in a scan line. The disclosure additionally relates to a scanning microscope.

A spectroscopic microscope according to the present embodiment includes a light source that generates laser light that enters a sample, a multi-slit part having a plurality of slits through which signal light branched by the edge filter passes, the slits being arranged in the slit width direction, and a spectrometer that disperses the signal light having passed through the slits in the dispersion direction intersecting the slit length direction and detects the signal light with a two-dimensional array photodetector.

A system for the simultaneous videographic or photographic acquisition of images, in particular of samples in a plurality of sample chambers of a sample plate, preferably a microtiter plate, includes an array of microscopes having mutually parallel optical axes, wherein each microscope includes an imaging chip and an objective. The imaging chips are attached to a carrier board as an array of columns and rows. An electronics unit for processing image data for all the microscopes is associated with the carrier board.

The invention relates to a method for scanning microscopy wherein a specimen is scanned simultaneously with a plurality of illumination spots of an excitation light. The light emitted by one specimen location irradiated with one illumination spot is detected independently of the light emitted by another specimen location illuminated with another illumination spot. A microscopic image of the specimen can be compiled from the emitted light detected for the different specimen locations. The method provides that the intensities of the different illumination spots are set independently of one another, and in that the illumination spots are guided over the specimen one after another in a scan line. The invention additionally relates to a scanning microscope.

Scanning fluorescence microscopes with an observation beam path from a measurement volume to an image plane. A beam combiner is provided for coupling an illumination system and a diaphragm arranged in the image plane for slow composition of the image because of the sequential scanning and subject the sample to loading as a result of inefficient use of the excitation light. The microscope simultaneously detects fluorescence from different focal planes in each case quasi-confocally. The observation beam path between the beam combiner and the image plane has a first diffractive optics for splitting light beams into beam bundles along different orders of diffraction, imparting to the light beams a spherical phase that is different from the other orders of diffraction. A second diffractive optics is provided for the compensation of chromatic aberrations of the split beam bundles, and a collecting optics is provided for focusing split beam bundles into the image plane.