Brillouin microscopy is a label-free and noncontact optical method to measure fundamental viscoelastic properties of biological systems. One of the main challenges in Brillouin microscopy is given by the strong Rayleigh background light overcoming the Brillouin peaks that are spectrally located at a few GHz from the laser frequency. Here, we demonstrate a common-path and broadband notch filter providing an unprecedented 65 dB extinction ratio. We demonstrate the filter by acquiring Brillouin spectra of highly turbid samples, such as pure milk and unprocessed vertebra bone tissues.
Standard imaging systems provide a spatial resolution that is ultimately dictated by the numerical aperture. In biological tissues, the resolution degraded by scattering which limits the imaging at a depth. Here, we exploit the properties of speckle patterns embedded into a strongly scattering matrix to illuminate the sample at high spatial frequency content. Combining adaptive optics with a custom deconvolution algorithm, we obtain a resolution improvement of a factor < 2.5. Our Scattering Assisted Imaging (SAI, M. Leonetti et al., Sci. Rep. 9:4591 (2019)) provides an effective solution to increase the resolution when long working distance optics are needed.
Standard imaging systems provide a spatial resolution that is ultimately dictated by the numerical aperture (NA) of the illumination and collection optics. In biological tissues, the resolution is strongly affected by scattering, which limits the penetration depth to a few tenths of microns. Here we exploit the properties of speckle patterns embedded into a strongly scattering matrix to illuminate the sample at high spatial frequency content. Combining illumination performed through a Digital Micromirror Device(DMD) and a custom deconvolution algorithm, we obtain an increase in the transverse spatial resolution by a factor of 2.5 with respect to the natural diffraction limit.
Altered biomechanics and phase transitions are implicated as key photogenic triggers in neurodegenerative diseases. However, standard methods to measure them require invasive contact with the sample or provide low spatial resolution. Here, we demonstrate Brillouin microscopy as a potential tool to investigate liquid-to-solid phase transitions in intracellular compartments in response to expression of ALS-linked proteins. In particular, we show how intracellular stress granules exhibit altered biomechanics in response to recruitment of RNA-binding proteins, such as ELAVL4 and FUS. Results pave the way to a better understanding of the dynamics that lead to formation of solid aggregates during neurodegeneration.
We demonstrate a compact on-chip gas sensor based on a SiN photonic integrated circuit providing an estimated detection limit of 0.1ppm for a variety of gas vapors, including IPA, ethanol and acetone. The sensor consists of an integrated Mach-Zehnder interferometer having one arm coated with an absorptive mesoporous layer to enhance the interaction of the gas vapors with the waveguide evanescent field. Gas detection is accomplished by monitoring the spectral shift of the output signal induced by a change in the effective refractive index of the coated waveguide. Results pave the way to novel integrated solutions for environmental safety monitoring.
Demand to reveal fundamental micro-mechanical properties is driven by a growing evidence that altered cellular processes in aging-associated disease environments are caused by a change in the regulating biomechanics. Unlike standard elastography techniques, Brillouin microscopy has shown great capabilities to non-invasively assess the biomechanics in the volume of biological samples, such as the lens cornea, atherosclerotic plaques and cells.
Spectral contrast is key in Brillouin microscopy to optically probe biological systems, where the elastic Rayleigh scattering and specular reflection are orders of magnitude greater than the Brillouin signal. Here, we developed a noncontact and label-free imaging method, named background-deflection Brillouin (BDB) microscopy, to investigate the three-dimensional intracellular biomechanics at a sub-micron resolution. Our method exploits diffraction to achieve an unprecedented 10,000-fold enhancement in the spectral contrast of single-stage spectrometers, enabling the first direct biomechanical analysis on intracellular stress granules containing ALS mutant FUS protein in fixed cells. Our results provide insights on an aberrant liquid-to-solid phase transition observed in in-vitro reconstituted droplets of FUS protein, which has been recently proposed as a possible pathogenic mechanism for ALS.
Spontaneous Brillouin scattering is an inelastic scattering process arising from inherent thermal density fluctuations, or acoustic phonons, propagating in a medium. The recent development of high throughput efficiency Virtually Imaged Phased Array (VIPA) etalons and high sensitivity CCD cameras has dramatically reduced the data acquisition time, in turn enabling the extension of Brillouin spectroscopy from a point sampling technique to an imaging modality. Hitherto Brillouin microscopy has shown great capabilities to non-invasively assess the biomechanics in the volume of biological samples, such as the lens cornea, atherosclerotic plaques and cells.
Spectral contrast is key to optically probe biological systems, where the elastic Rayleigh scattering and specular reflection are orders of magnitude greater than the Brillouin signal. In VIPA spectrometers, the elastic background light introduces crosstalk signals that overwhelms the weak Brillouin peaks, thus impeding the acquisition of biomechanical images. One method to increase the contrast is to add more etalons in tandem and crossed with respect to each other. Nevertheless, this comes at the cost of a reduced throughput efficiency and a significantly increase system complexity. Here we demonstrate a method to increase the contrast by more than 30dB respect to standard VIPA spectrometers without the requirement of any additional optical or dispersive components. Our method was demonstrated by acquiring Brillouin images of single cells at a sub-micron spatial resolution, where the biomechanical properties of individual cellular structures were investigated.
Spontaneous Brillouin scattering is an inelastic scattering process arising from inherent thermal density fluctuations, or acoustic phonons, propagating in a medium. Over the last few years, Brillouin spectroscopy has shown great potential to become a reliable non-invasive diagnostic tool due to its unique capability of retrieving viscoelastic properties of materials such as strain and stiffness. The detection of the weak scattered light, in addition to the resolution of the Brillouin peaks (typically shifted by few GHz from the central peak) represent one of the greatest challenges in Brillouin. The recent development of high sensitivity CCD cameras has brought Brillouin spectroscopy from a point sampling technique to a new imaging modality. Furthermore, the application of Virtually Imaged Phased Array (VIPA) etalons has dramatically reduced insertion loss simultaneously allowing fast (<1s) collection of the entire spectrum. Hitherto Brillouin microscopy has been shown the ability to provide unique stiffness maps of biological samples, such as the human lens, in a non-destructive manner. In this work, we present results obtained using our Brillouin microscope to map the stiffness variations in the walls of blood vessels in particular when atherosclerotic plaques are formed. The stiffness of the membrane that covers the plaques is critical in developing acute myocardial infarction yet it is not currently possible to credibly assess its stiffness due to lack of suitable methods.
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