The present invention relates to systems and methods for tissue processing and imaging including a counterintuitive inverse relationship between protein dye concentration and tissue sample protein content. Varying dye concentration in such a manner affords higher quality fluorescent imaging at depth in tissue when using optical sectioning microscopy or second harmonic generation (SHG) analysis. Methods of the invention thereby provide more usable histopathology images while reducing waste and reagent costs and are of particular utility in combination assays that include simultaneous protein and nuclear staining and SHG analysis.

A super class of polarized transverse vector vortex photon beams patterns are mathematically represented here, which are Majorana-like among them are the radial and azimuthal Laguerre-Gaussian, hybrid π-vector beams, and Airy beams. These optical beams are consider spin-orbit coupled beams based on OAM and SAM parts of light. A Majorana photon is a photon that is identical to its anti-photon. It has within itself both chirality, right and left-handed twist in polarization (SAM) and wavefront (OAM). Applications using Majorana photons improve optical deeper imaging, higher resolution imaging, Nonlinear Optics effects (SHG, SRS, SC), optical communication in free space and fibers, quantum computer as basic qubit, and entanglement for security.

A system for multimodal nonlinear optical imaging is provided. Each mode uses a high NA objective to cause total internal reflection excitation at a sample-substrate interface. The system has a femtosecond oscillator to generate pulses used for two beams. The objective receives at least one beam, redirects the received at least one beam through a dielectric substrate to cause the TIR and produces corresponding evanescent waves in a portion of the sample adjacent to the sample-substrate interface, and collects a backward-propagating beam of pulses of responsive light. The portion of the sample illuminated by the evanescent waves emits responsive light. Different modes or combinations of the distinct modalities may be selected to access complementary chemical and structural information for various chemical species near the sample-substrate interface. Each mode may have mode-specific control such as selective beam blocking, power ratios and filtering.

Embodiments disclosed include methods and apparatus for Fluorescent Enhanced Photothermal Infrared (FE-PTIR) spectroscopy and chemical imaging, which enables high sensitivity and high spatial resolution measurements of IR absorption with simultaneous confocal fluorescence imaging. In various embodiments, the FE-PTIR technique utilizes combined/simultaneous OPTIR and fluorescence imaging that provides significant improvements and benefits compared to previous work by simultaneous detection of both IR absorption and confocal fluorescence using the same optical detector at the same time.

Surface sensing methods for imaging a scanned surface of a sample via sum-frequency vibrational spectroscopy are disclosed herein. The methods include exposing a sampled location of the scanned surface to a visible light beam and exposing the sampled location to a tunable infrared beam such that the tunable infrared beam is at least partially coincident with the visible light beam. The methods also include varying a frequency of the tunable infrared beam an inducing optical resonance within an imaged structure that extends at least partially within the sampled location. The methods further include receiving at least a portion of an emitted light beam from the sampled location and scanning the visible light beam and the runnable infrared beam across the scanned portion of the scanned surface. The methods also include generating an image of the scanned portion of the scanned surface based upon the receiving and the scanning.

A method of analysing an aqueous fluid comprising obtaining a 2D-IR spectrum of a sample of the aqueous fluid using a 2D-IR spectrometer configured to apply a sequence of IR pulses to the sample, wherein the sequence comprises a pump process followed by a probe pulse, where the pump process is a single pump pulse or a sequence of a first pump pulse and a second pump pulse, and a waiting time T.sub.w between applying the single pump pulse or the second pump pulse and applying the probe pulse is from 150 to 350 fs.

A method for imaging a sample, wherein the sample changes a polarization state of light as a function of position, wherein the method includes changing a polarization state of a purely polarized light of an incident light striking a micro-retarder array, thereby inducing a changed polarization state of the polarization state. The micro-retarder array is placed in a rear conjugate focal plane of a microscope. The method additionally includes projecting the changed polarization state of the polarization state into an object plane of the microscope containing the sample.

Disclosed herein is a super resolution imaging method and system for obtaining an image in a crystal material and/or device.

The invention relates to a method and to an assembly (200) for localizing an optical coupling point (11) and to a method for producing a microstructure (100) at the optical coupling point (11). The method for localizing an optical coupling point (11) comprises the following steps: a) providing an optical component (10), which comprises an optical coupling point (11), the optical coupling point having an interaction region (15) lying outside of a volume encompassed by the optical component (10); b) producing optical radiation in a production region (120), the production region (120) overlapping at least partly with the interaction region (15) of the optical coupling point (11), light being applied to a medium (19) located in the production region (120), which light is modified by the medium (19) in such a way that the optical radiation is thereby produced; c) sensing at least part of the produced optical radiation in a sensing region (130), the sensing region (130) overlapping at least partly with the interaction region (15) of the optical coupling point (11), and determining a spatially resolved distribution of the sensed part of the produced optical radiation; and d) determining the localization of the optical coupling point (11) from the determined spatially resolved distribution of the sensed part of the produced optical radiation, the optical radiation being produced or at least the part of the produced optical radiation being sensed through the optical coupling point (11). The optical coupling point (11) can thereby be precisely localized with a relative positioning tolerance of better than 1 μm. Thus, low coupling losses of an optical connection to the optical component (10) can be achieved and microstructures (100) can be precisely placed at the optical coupling point (11).

Systems and methods are provided for analyzing a biomechanical property of a medium using stimulated Brillouin scattering microscopy. The method can include a first step of applying a probe beam and pulsed pump beam to a target section of the medium, wherein the pump beam interacts with the probe beam to generate at least one acoustic wave in the medium and at least one Brillouin signal is produced as a result of the generated acoustic wave. The method can also include a second step of receiving the produced Brillouin signal and a third step of determining, using a processor and the