A confocal optical system includes a light source and a spinning polarizer disposed in the optical pathway such the light emitted from the light source passes through the spinning polarizer. A first objective lens is disposed in the optical pathway to allow passage of light that passes through the spinning polarizer. A microlens array member is disposed adjacent the first objective lens to receive light. The microlens array member includes a plate having a plurality of holes arranged in an array pattern. A second objective lens is disposed in the optical pathway to receive and allow passage of light to a sample. The optical pathway is arranged such that, after reaching the sample, the light is directed back through the second objective lens, the microlens or microlens with filter array, and the first objective lens and a fluorescent filter cube as an emission beam to reach at least one camera which provides an image of the sample.

A modular microscope can quickly be modified for specific scanning applications. The microscope includes a microscope main body which has slots into which long pass filter modules, dichroic mirror modules, notch filter modules, and LED modules can be selectively placed, removed, and changed out. In some applications, the interchangeable components permit quickly changing between Photoluminescence (PL) and Raman spectroscopy (microscope) systems.

A confocal microscope device for scanning a two-dimensional array of illumination beams over a target surface and scanning a corresponding two-dimensional array of emission beams stimulated by the array of illumination beams on to a sensor of an imaging device. The device comprises first scanning optics operable to scan the array of illumination beams over the target surface along a first axis and scan the array of emission beams over the sensor along the first axis. The device further comprises second scanning optics operable to deflect, on a second axis, the array of illumination beams as they are scanned over the target surface along the first axis, such that uneven stimulation of the target surface by the array of illumination beams due to interference of the illumination beams is reduced, and deflect, on the second axis, the array of emission beams as they are scanned over the sensor of the imaging device along the first axis such that uneven stimulation of the sensor by the array of emission beams due to interference of the emission beams is reduced.

Laser emission based microscope devices and methods of using such devices for detecting laser emissions from a tissue sample are provided. The scanning microscope has first and second reflection surfaces and a scanning cavity holding a stationary tissue sample with at least one fluorophore/lasing energy responsive species. At least a portion of the scanning cavity corresponds to a high quality factor (Q) Fabry-Pérot resonator cavity. A lasing pump source directs energy at the scanning cavity while a detector receives and detects emissions generated by the fluorophore(s) or lasing energy responsive species. The second reflection surface and/or the lasing pump source are translatable with respect to the stationary tissue sample for generating a two-dimensional scan of the tissue sample. Methods for detecting multiplexed emissions or quantifying one or more biomarkers in a histological tissue sample, for example for detection and diagnosis of cancer, or other disorders/diseases are provided.

This disclosure includes an imaging system that is configured to image in parallel multiple focal planes in a sample uniquely onto its corresponding detector while simultaneously reducing blur on adjacent image planes. For example, the focal planes can be staggered such that fluorescence detected by a detector for one of the focal planes is not detected, or is detected with significantly reduced intensity, by a detector for another focal plane. This enables the imaging system to increase the volumetric image acquisition rate without requiring a stronger fluorescence signal. Additionally or alternatively, the imaging system may be operated at a slower volumetric image acquisition rate (e.g., that of a conventional microscope) while providing longer exposure times with lower excitation power. This may reduce or delay photo-bleaching (e.g., a photochemical alteration of the dye that causes it to no longer be able to fluoresce), thereby extending the useful life of the sample.

The invention relates to an optical arrangement, particularly for the detection beam path of a multi-spot scanning microscope, comprising a detection plane, in which a detector is positionable, comprising a dispersive device for spectrally splitting detection light. According to the invention, the optical arrangement is characterized in that a distorting optical unit is present for guiding the detection light into the detection plane, said distorting optical unit being arranged downstream of the dispersive device and upstream of a detection plane, and in that a rotating device is present for the relative rotation of a luminous field of the spectrally separated detection light and the distorting optical unit. The invention additionally relates to a multi-spot scanning microscope and a method for operating a microscope.

A re-scan microscope for forming an image of a sample is disclosed. The system comprises an illumination optical system for directing, and optionally focusing, illumination light at the sample herewith providing an illumination light spot at the sample. The illumination light spot causes emission light from the sample. The microscope system further comprises a detection optical system for focusing at least part of the emission light onto an imaging plane of an imaging system herewith causing an emission light spot on the imaging plane. The microscope system also comprises a rotatable element for, when rotating, moving the illumination light spot over and/or through the sample and simultaneously moving the emission light spot over said imaging plane of the imaging system. The rotatable element comprises at least two reflective surfaces.

A method of contrast enhancement of a three-dimensional light sheet microscopy image formed from N individual images each corresponding to a light sheet plane and spaced apart from each other in the z-direction by at least a distance d, the x/y-plane being the light sheet plane and the x-direction being the propagation direction of the light sheet of the light sheet plane, comprising:

Deconvolution of the three-dimensional image in the z-direction, comprising: For each intensity vector of N intensities (I.sub.x,y,1, . . . ,I.sub.x,y,N) having the same x/y value, performing a multiplication with a tridiagonal N×N deconvolution matrix, which assigns to each voxel (x, y, n) a correction parameter f1, with which, by the multiplication of the deconvolution matrix with the intensity vector, for a component I(x, y, n) of the intensity vector the intensities Ix, y,n+1 and Ix, y,n−1 of the corresponding voxels of the neighboring image plane are multiplied, before they are subtracted from the intensity value I(x, y,n).

A photon peak event detection system accepts an analog output from a photon sensor, directly digitizes the analogy output and includes a graphics processing unit (GPU) programmed to conduct a photon peak event detection in real-time via a photon count program that analyzes the digitized photon sensor output in sampling periods each having at least three consecutive data points to determine a local maximum among the consecutive data points and compare the local maximum to one or more predetermined thresholds to determine whether or not a photon was received in each sampling period, the algorithm providing photon counts to a phasor analysis program in the GPU. The phasor analysis program calculates pixelwise fluorescence lifetime phasor data in real-time and sends the data to a central processing unit.

Provided is a two-photon spectroscopy including a light source configured to generate first laser light having a pulse, a pulse width correction device configured to receive the first laser light to output a second laser light, an optical system through which the second laser light passes, a first two-photon sensor configured to measure a first pulse width of the first laser light generated from the light source, and a second two-photon sensor configured to measure a second pulse width of the second laser light passing through the optical system, wherein the pulse width correction device corrects a difference between the first pulse width and the second pulse width.