Transparent Samples Research Articles

Study on Measurement Method of Three-dimensional Position of Unlabeled Microspheres under Bright Background

Abstract: Using computer vision technology to obtain the position and trajectory data of particle probe microspheres from microscope images has significance and value in the molecular field. However, most of the existing microsphere measurement methods are based on transmission, which can only be measured under transparent samples and substrates and are not suitable for the application scenario of living cell measurement. In this paper, a method based on reflectivity imaging is proposed to measure the three-dimensional position of the dark microspheres in the bright field. Based on the outermost ring radius method, the relationship between the inner ring radius of the microsphere spot and the out-of-focus distance was explored to measure the coordinates in the Z direction. Cardiomyocytes were combined with 10um size silica microspheres. Experiments show that in a bright field with a high perturbation environment, it can achieve high precision measurement of dark microspheres and achieve three-dimensional position measurement with an accuracy of 50nm in XY direction and 100nm in Z direction.

High pressure and high temperature Brillouin scattering measurements of pyrope single crystals using flexible CO2 laser heating systems

Single-crystal Brillouin scattering measurements are important for interpreting seismic velocities within the Earth and other planetary interiors. These measurements are rare, however, at temperatures above 1000 K, due to the fact that the transparent samples cannot be heated by common laser heating systems operating at a wavelength on the order of 1 μm. Here we present Brillouin scattering data on pyrope collected at pressures up to 23.8 GPa and temperatures between 850 and 1900 K using a novel CO2 laser heating system confined in either a flexible hollow silica waveguide or an articulated arm with mirrors mounted in each junction to direct the laser to the exit point. Pyrope has been chosen because it has been extensively studied at high pressures and moderate temperatures and therefore it is an excellent sample for bench-marking the CO2 laser heating system. The new high-temperature velocity data collected in this study allow the room pressure thermal parameters of pyrope to be constrained more tightly, resulting in values that reproduce the temperature dependence of the unit-cell volume of pyrope measured in recent studies at ambient pressure. Aggregate wave velocities of pyrope calculated along an adiabat using the thermoelastic parameters determined in this study are larger than those obtained using published values, implying that velocities for many mantle components may be underestimated at mantle temperatures because high temperature experimental data are lacking.

Effect of Eu Ions Concentration in Y2O3-Based Transparent Ceramics on the Electron Irradiation Induced Luminescence and Damage

Eu3+-doped Y2O3-based luminescent materials can be used as a scintillator for electron or high energy β-ray irradiation, which are essential for applications such as electron microscopy and nuclear batteries. Therefore, it is essential to understand their defect mechanisms and to develop materials with excellent properties. In this paper, Y2O3-based transparent ceramics with different Eu3+ doping concentrations were prepared by solid-state reactive vacuum sintering. This series of transparent ceramic samples exhibits strong red emission under electron beam excitation at the keV level. However, color change appears after the high-energy electron irradiation due to the capture of electrons by the traps in the Y2O3 lattice. Optical transmittance, laser-excited luminescence, X-ray photoelectron spectroscopy (XPS), and other analyses indicated that the traps, or the color change, mainly originate from the residual oxygen vacancies, which can be suppressed by high Eu doping. Seen from the cathodoluminescence (CL) spectra, higher doping concentrations of Eu3+ showed stronger resistance to electron irradiation damage, but also resulted in lower emissions due to concentration quenching. Therefore, 10% doping of Eu was selected in this work to keep the high emission intensity and strong radiation resistance both. This work helps to enhance the understanding of defect formation mechanisms in the Y2O3 matrix and will be of benefit for the modification of scintillation properties for functional materials systems.

Near‐infrared scintillation properties of Nd-doped BaO-Bi2O3-P2O5 glasses

We synthesized BaO-Bi2O3-P2O5 glass samples with different Nd concentrations by the melt-quenching technique. The concentrations were 1.0, 5.0, 10, 15, and 20 %. We obtained transparent and homogeneous samples for 1.0–10 % Nd concentrations. The 15 % Nd-doped sample was inhomogeneous, and the 20 % Nd-doped sample contained a crystalline phase. The photoluminescence (PL) and scintillation properties of the 1.0–10 % Nd-doped samples were investigated. The samples showed PL and scintillation peaks in the near-infrared region, which were due to 4f–4f transitions of Nd3+. We tested the X-ray dose rate response functions of the samples. The 10 % Nd-doped samples showed the most intensive response among the examined samples, and the lower detection limit of dose rate was 0.06 Gy/h with our setup.

Visible/solar light active transparent BiB3O6 glass- ceramics multi-catalysis

Borate based non-centrosymmertic crystals are known for their non-linear optical properties. This study reports BiB3O6 glass-ceramics for photocatalysis and piezocatalysis applications. These were fabricated via. melt-quench technique and yellow transparent glass samples were made. Effectiveness of a bismuth-triborate glass-ceramic (BiB3O6) in degrading methylene blue (MB) and methyl violet (MV) dye undergoing photocatalysis in the presence of visible light and solar light for MB dye was studied. Degradation observed in photocatalysis was ∼62 % and ∼50 % for MB and MV dye respectively in 240 min whereas, ∼72 % of MB dye was degraded under solar light. Piezocatalysis was examined to assess the extended potential of this material, resulting in approximately ∼76 % degradation of MB dye. Moreover, in synergistic photo-piezocatalysis degradation of ∼88 % was observed. Phytotoxicity was also assessed using the germination index (GI) showing ∼76 %, using heat treated sample.

Label-free 3D molecular imaging of living tissues using Raman spectral projection tomography

The ability to image tissues in three dimensions (3D) with label-free molecular contrast at the mesoscale would be a valuable capability in biology and biomedicine. Here, we introduce Raman spectral projection tomography (RSPT) for volumetric molecular imaging with optical sub-millimeter spatial resolution. We have developed a RSPT imaging instrument capable of providing 3D molecular contrast in transparent and semi-transparent samples. We also created a computational pipeline for multivariate reconstruction to extract label-free spatial molecular information from Raman projection data. Using these tools, we demonstrate imaging and visualization of phantoms of various complex shapes with label-free molecular contrast. Finally, we apply RSPT as a tool for imaging of molecular gradients and extracellular matrix heterogeneities in fixed and living tissue-engineered constructs and explanted native cartilage tissues. We show that there exists a favorable balance wherein employing Raman spectroscopy, with its advantages in live cell imaging and label-free molecular contrast, outweighs the reduction in imaging resolution and blurring caused by diffuse photon propagation. Thus, RSPT imaging opens new possibilities for label-free molecular monitoring of tissues.

Off-axis digital lensless holographic microscopy based on spatially multiplexed interferometry.

Digital holographic microscopy (DHM) is a label-free microscopy technique that provides time-resolved quantitative phase imaging (QPI) by measuring the optical path delay of light induced by transparent biological samples. DHM has been utilized for various biomedical applications, such as cancer research and sperm cell assessment, as well as for in vitro drug or toxicity testing. Its lensless version, digital lensless holographic microscopy (DLHM), is an emerging technology that offers size-reduced, lightweight, and cost-effective imaging systems. These features make DLHM applicable, for example, in limited resource laboratories, remote areas, and point-of-care applications. In addition to the abovementioned advantages, in-line arrangements for DLHM also include the limitation of the twin-image presence, which can restrict accurate QPI. We therefore propose a compact lensless common-path interferometric off-axis approach that is capable of quantitative imaging of fast-moving biological specimens, such as living cells in flow. We suggest lensless spatially multiplexed interferometric microscopy (LESSMIM) as a lens-free variant of the previously reported spatially multiplexed interferometric microscopy (SMIM) concept. LESSMIM comprises a common-path interferometric architecture that is based on a single diffraction grating to achieve digital off-axis holography. From a series of single-shot off-axis holograms, twin-image free and time-resolved QPI is achieved by commonly used methods for Fourier filtering-based reconstruction, aberration compensation, and numerical propagation. Initially, the LESSMIM concept is experimentally demonstrated by results from a resolution test chart and investigations on temporal stability. Then, the accuracy of QPI and capabilities for imaging of living adherent cell cultures is characterized. Finally, utilizing a microfluidic channel, the cytometry of suspended cells in flow is evaluated. LESSMIM overcomes several limitations of in-line DLHM and provides fast time-resolved QPI in a compact optical arrangement. In summary, LESSMIM represents a promising technique with potential biomedical applications for fast imaging such as in imaging flow cytometry or sperm cell analysis.

Magneto-Optical Ceramics with High Transparency for Highly Sensitive Magnetometer via Quantum Weak Measurement.

Sensitive magnetometer technology is desirable for biomagnetic field detection and geomagnetic field measuring. Signal amplification materials such as magneto-optical crystals or ceramics are crucial for enhancing detection sensitivity, but severe optical scattering and low Verdet constant further limit its application. To develop high-sensitivity magnetometers for quantum weak measurement schemes, we have conducted investigations on the powder calcining dynamics and prepared a series of high-optical-quality (Ho/Dy)2Zr2O7 transparent ceramic samples. The Verdet constant of magneto-optical materials was measured across a continuous wavelength spectrum, exhibiting a peak at 283 ± 5 rad/(T·m). We further established an electron transition mechanism to elucidate the exceptional magneto-optical attributes of dysprosium. In addition, samples demonstrated superior performance in weak-value amplification, reaching a low detectable magnetic field threshold of 3.5 × 10-8 T and continuously worked over 6 h with high stability. Our work developed a highly sensitive magnetometer using optimized magneto-optical ceramics and provided guidance on design, fabrication, and application for magneto-optical ceramics in quantum weak measurement.

Research on additive manufacturing technology of high solid loading silica glass slurry

In this work, digital light processing (DLP) 3D printing technology was used to fabricate transparent silica glass with complex structures. The effects of different particle sizes and mixing ratios of silica powders on the viscosity of the prepared slurries were investigated. The results showed that when 100–200 nm and 800 nm silica powders were mixed in a 1:4 vol ratio, the slurry exhibited the lowest viscosity. The highest solid loading of the slurry prepared with this mixed silica powder reached 75 wt%, with a viscosity of only 1245 mPa·s. The high solid loading slurry was subjected to 3D printing, debinding, and sintering treatment, and the transparent silica glass samples with complex structures was obtained.

High-quality phase imaging by phase-shifting digital holography and deep learning

Quantitative phase imaging (QPI) technology is widely used in biomedical imaging and other fields because it can realize exact imaging for transparent phase-type samples, which is of great research significance. The complex amplitude distribution of the object wave obtained by phase-shifting digital holography (PSDH) reproduction can provide phase information for QPI, but its existence of phase wrapping and other problems limits its practical application. Although the traditional phase unwrapping algorithm provides a solution, it has problems such as low unwrapping accuracy or long time running. To solve these problems in QPI, a high-quality phase imaging (HQPI) method by PSDH and deep learning (DL) is proposed, where QPI is achieved by extracting the unknown phase shift using a generalized non-iterative phase shift extraction algorithm and unwrapping the wrapped phase by a DL network. Both numerical simulations and optical experiments verify the feasibility of the method. By comparing with the traditional phase unwrapping algorithm, it is demonstrated that the DL unwrapping method has higher unwrapping accuracy and more efficiency. The results show that the method of HQPI is capable of realizing comparatively fast and accurate QPI.

Simultaneous mapping of cathodoluminescence spectra and backscatter diffraction patterns in a scanning electron microscope

Electron backscatter diffraction and cathodoluminescence are complementary scanning electron microscopy modes widely used in the characterisation of semiconductor films, respectively revealing the strain state of a crystalline material and the effect of this strain on the light emission from the sample. Conflicting beam, sample and detector geometries have meant it is not generally possible to acquire the two signals together during the same scan. Here, we present a method of achieving this simultaneous acquisition, by collecting the light emission through a transparent sample substrate. We apply this combination of techniques to investigate the strain field and resultant emission wavelength variation in a deep-ultraviolet micro-LED. For such compatible samples, this approach has the benefits of avoiding image alignment issues and minimising beam damage effects.

Assessing the light scattering properties of c-Si PV module materials for agrivoltaics: Towards more homogeneous light distribution in crop canopies

Increasing the diffusivity of transmitted light at crop canopies in agrivoltaic (AV) systems remains a desired objective. This is because diffuse light increases the uniformity of light distribution and penetrates deeper into compact crop canopies thereby enhancing the photosynthesis rate and crop yields. Current approaches in greenhouses and open horticultural systems involve the use of diffusing films, diffuse glass, and light diffusing coatings. While these methods are effective, their combination with photovoltaic (PV) modules in AV greenhouses and other AV farming practices remain relatively unknown. However, little to no work has been done to assess the light scattering properties of the existing PV module structural materials such as the glass, encapsulants and the transparent backsheet. This work therefore investigates the light scattering behaviour of c-Si PV module materials through haze and Hortiscatter measurements. 18 samples with the structural layout of glass/encapsulant/encapsulant/back cover (glass or transparent polymer backsheet), applying different commercial encapsulants, were manufactured and tested experimentally. Light in the photosynthetic spectrum was used in the optical characterization of the samples. Furthermore, the impact of UV degradation on the haziness was also tested and the uniformity of the light distribution was further assessed to obtain the Hortiscatter. The findings indicated that (i) Transparent backsheets increased the light diffusivity. (ii) UV degradation reduced the light scattering of most of the PV materials. (iii) For the haziest transparent backsheet sample, the distribution of the transmitted light increased with the incidence light angle and reduced with increasing wavelength in the visible spectrum (iv) Highest haze and Hortiscatter values of up to 80% and 84% respectively were obtained for a sample with glass/TPO/TPO/transparent backsheet layout. (v) Haze and Hortiscatter values could help in optimising the PV module bill of materials for AV applications.

Near 2π solid angle fluorescence collection for laser-induced breakdown spectroscopy.

Detecting heavy metal and radioactive elements distributed in the environment and human body is crucial for life and environmental safety. A lens with a high numerical aperture (NA) is used in laser-induced breakdown spectroscopy to collect the plasma fluorescence as much as possible and improve the limit of detection. However, even a lens with NA up to 0.6, only the fluorescence in a solid angle of 0.4π can be collected. In this work, a novel, to our knowledge, fluorescence collecting scheme composed of a parabolic mirror and a lens is proposed which can collect the plasma fluorescence in a solid angle of ∼1.4π for an opaque material and ∼1.9π for a transparent material. Simulation results show that for opaque samples, this method can improve the fluorescence collection ability by 3.8 times compared to a single lens (NA = 0.5) collection scheme, and for transparent samples 4.5 times may be achieved. In experiments, a 2.8-fold enhancement in the fluorescence collection ability is demonstrated, and the signal-to-noise ratio is increased by 2.5 times for opaque samples.

Velocimetry of GHz elastic surface waves in quartz and fused silica based on full-field imaging of pump–probe reflectometry

This study reports an imaging method for gigahertz surface acoustic waves in transparent layers using infrared subpicosecond laser pulses in the ablation regime and an optical pump–probe technique. The reflectivity modulations due to the photoelastic effect of generated multimodal surface acoustic waves were imaged by an sCMOS camera illuminated by the time-delayed, frequency-doubled probe pulses. Moving the delay time between 6.0nsto11.5ns, image stacks of wave field propagation were created.Two representative samples were investigated: wafers of isotropic fused silica and anisotropic x-cut quartz. Rayleigh (SAW) and longitudinal dominant high-velocity pseudo-surface acoustic wave (HVPSAW) modes could be observed and tracked along a circular grid around the excitation center, allowing the extraction of angular profiles of the propagation velocity. In quartz, the folding of a PSAW was observed. A finite element simulation was developed to predict the measurement results. The simulation and measurement were in good agreement with a relative error of 2% to 5%.These results show the potential for fast and full-field imaging of laser-generated ultrasonic surface wave modes, which can be utilized for the characterization of thin transparent samples such as semiconductor wafers or optical crystals in the gigahertz frequency range.

Double hysteresis (P-E) loops in the transparent composite containing BaTiO3 at room temperature

A transparent composite sample containing BaTiO3 was synthesized by using the melting quenching method. The presence of a glassy phase in this composite sample was detected through XRD analysis, and this was further confirmed by DSC study. TEM and SAED analyses provided evidence that the BaTiO3 nanoparticles/clusters are embedded within the borate glass matrix, establishing the composite nature of this sample. FTIR spectrum of the present sample revealed that the glass matrix is composed of two structural groups (BO3 with NBO’s, and BO4), along with the distinct groups for BaTiO3. XPS spectra of the present composite sample indicated the presence of more than one type of boron, barium, titanium and oxygen. DSC and dielectric studies of the present composite sample revealed the presence of the phase transition temperature (Tc). Dielectric constant (ɛr) and dielectric loss (tan δ) curves of the present composite sample displayed an anomaly peak in the vicinity of TC. The optical transmission spectrum of the present composite sample exhibit two transmission bands of Ti3+ (3d1) ions in tetragonal distorted sites. At room temperature, the present transparent composite sample exhibited double hysteresis loops for BaTiO3 at low electric fields. The results obtained can be used for the development of lead-free ferroelectric material.

Speckle-enabled in vivo demixing of neural activity in the mouse brain.

Functional imaging of neuronal activity in awake animals, using a combination of fluorescent reporters of neuronal activity and various types of microscopy modalities, has become an indispensable tool in neuroscience. While various imaging modalities based on one-photon (1P) excitation and parallel (camera-based) acquisition have been successfully used for imaging more transparent samples, when imaging mammalian brain tissue, due to their scattering properties, two-photon (2P) microscopy systems are necessary. In 2P microscopy, the longer excitation wavelengths reduce the amount of scattering while the diffraction-limited 3D localization of excitation largely eliminates out-of-focus fluorescence. However, this comes at the cost of time-consuming serial scanning of the excitation spot and more complex and expensive instrumentation. Thus, functional 1P imaging modalities that can be used beyond the most transparent specimen are highly desirable. Here, we transform light scattering from an obstacle into a tool. We use speckles with their unique patterns and contrast, formed when fluorescence from individual neurons propagates through rodent cortical tissue, to encode neuronal activity. Spatiotemporal demixing of these patterns then enables functional recording of neuronal activity from a group of discriminable sources. For the first time, we provide an experimental, in vivo characterization of speckle generation, speckle imaging and speckle-assisted demixing of neuronal activity signals in the scattering mammalian brain tissue. We found that despite an initial fast speckle decorrelation, substantial correlation was maintained over minute-long timescales that contributed to our ability to demix temporal activity traces in the mouse brain in vivo. Informed by in vivo quantifications of speckle patterns from single and multiple neurons excited using 2P scanning excitation, we recorded and demixed activity from several sources excited using 1P oblique illumination. In our proof-of-principle experiments, we demonstrate in vivo speckle-assisted demixing of functional signals from groups of sources in a depth range of 220-320 µm in mouse cortex, limited by available speckle contrast. Our results serve as a basis for designing an in vivo functional speckle imaging modality and for maximizing the key resource in any such modality, the speckle contrast. We anticipate that our results will provide critical quantitative guidance to the community for designing techniques that overcome light scattering as a fundamental limitation in bioimaging.

Off-Axis Co-Optical Path Large-Range Line Scanning Chromatic Confocal Sensor

This article proposes a line scanning chromatic confocal sensor to solve the problem of limited chromatic confocal measurement due to the small measurement range and low measurement efficiency in the industrial inspection process. To obtain an extensive dispersion range, the advantages of a simple single-axis structure are combined with the advantages of a large luminous flux of a biaxial structure. Considering large-scale measurement, our sensor uses off-axis rays to limit the illumination path and imaging path to the same optical path structure. At the same time, the field of view is expanded, and a symmetrical structure is adopted to provide a compact optical path and improve space utilization. The simulation and physical system test results shows that the sensor scanning line length is 12.5 mm, and the axial measurement range in the 450 nm to 750 nm band is better than 20 mm. The axial resolution of the detector is ±1 µm combined with the subpixel centroid extraction data processing method, and the maximum allowable tilt angle for specular reflection samples is ±7°. The thicknesses of transparent standard flat glass and the wet collagen membrane are measured. The maximum average error is 1.3 µm, and the relative error is within 0.7%. The constructed sensor is of great significance for rapidly measuring the three-dimensional profile, flatness, and thickness in the fields of transparent biological samples, optics, micromechanics, and semiconductors.

Intrinsic and Extrinsic Scattering and Absorption Coefficients New Equations in Four-flux and Two-flux Models Used for Determining Light Intensity Gradients

Collimated, diffuse and total light intensity gradients, for forward and backward light senses, were determined in a three substrate layers – glass/electrolyte/glass – almost transparent sample, using optical constants and new equations for intrinsic and extrinsic scattering and absorption coefficients. These new equations were obtained for the inner electrolyte layer from the systems of differential equations of the four-flux and two-flux radiative transfer models, used for determining intrinsic and extrinsic coefficients, respectively, once knowing the optical constants of outer glass layers, from a single glass substrate sample measured in advance. Extinction coefficients were determined from optical constants and considering the wavelength compression of light when it enters into a material, decreasing its speed with respect to the vacuum. The same extinction coefficients for glass and electrolyte layers were computed in three different ways. First, from optical constants, determined using collimated transmittance and reflectance solutions of four-flux model. From them and as intermediate parameters for glass and electrolyte layers, collimated interface reflectance and attenuation due to extinction were computed since they were required for determining inner collimated light intensities at the interfaces, which were used at the collimated forward and backward differential equations, solving for the forward and backward extinction coefficients. The three-extinction matching requirement was successfully satisfied for the glass and electrolyte layers. Two average crossing parameters equations, for each sense, and four forward scattering ratios equations, for collimated and for diffuse light intensities for each sense, were used in the system of diffuse differential equations for intrinsic coefficients. For them, intuitive equations were proposed, based on collimated and diffuse light intensities at each interface. New equations for extrinsic parameters were determined by equalizing the system of total differential equations of the two-flux model to the sum of the systems of collimated and diffuse differential equations of the four-flux model.

Single-shot differential phase contrast microscopy using ring-shaped polarisation multiplexing illumination.

We propose a differential phase contrast microscopy that enables single-shot phase imaging for unstained biological samples. The proposed approach employs a ring-shaped LED array for polarisation multiplexing illumination and a polarisation camera for image acquisition. As such, multiple images of different polarisation angles can be simultaneously captured with a single shot. Through polarisation demultiplexing, the sample phase can therefore be recovered from the single-shot measurement. Both simulations and experiments demonstrate the effectiveness of the approach. We also demonstrate that ring-shaped illumination enables higher contrast and lower-distortion imaging results than disk-shaped illumination does. The proposed single-shot approach potentially enables phase contrast imaging for live cell samples in vitro. Lay Description: We propose a microscopy that enables imaging of transparent samples, unstained cells, etc. We demonstrate that the proposed method enables higher contrast and lower-distortion imaging results than conventional methods, and significantly improves imaging efficiency. The proposed method potentially enables dynamic imaging for live cell samples in vitro.

Incoherent partial superposition modeling for single-shot angle-resolved ellipsometry measurement of thin films on transparent substrates

Ellipsometric measurement of transparent samples suffers from substrate backside reflection challenges, including incoherent and partial superposition issues. The recently developed angle-resolved ellipsometry (ARE) can naturally eliminate the backside reflections of substrates with a micro-spot equivalent thickness or thicker; however, for thinner substrates, ARE working with general incoherent backside reflection models shows significant inaccuracy or measurement failure. In this paper, an incoherent partial superposition (IPS) model is proposed to characterize the optical superposition effect between the frontside and uncertain backside reflections from an unknown substrate. IPS introduces a cosine-like correction of the backside reflection, corresponding to the overlapping-area change of backside and frontside reflections along with incident angles. Benefiting from ARE’s wide-angle spectral imaging capability, IPS achieves single-shot measurement of thin film thicknesses on transparent substrates of unknown thickness. An ARE system was built and calibrated regarding the linear relationship between the cosine-corrected angular frequencies and substrate thicknesses. Then, commercial ITO films on glasses of different thicknesses ranging from 200 to 1000 µm were measured. Experimental results show that IPS-ARE results in a root-mean-square accuracy error of ∼1 nm in film thickness measurement and provides a ∼77% error reduction from general incoherent backside reflection models.