Angular optimization for cancer identification with circularly polarized light

Depolarization of circularly polarized light scattered from biological tissues depends on structural changes in cell nuclei, which can provide valuable information for differentiating cancer tissues concealed in healthy tissues. In this study, we experimentally verified the possibility of cancer identification using scattering of circularly polarized light. We investigated the polarization of light scattered from a sliced biological tissue with various optical configurations. A significant difference between circular polarizations of light scattered from cancerous and healthy tissues is observed, which is sufficient to distinguish a cancerous region. The line-scanning experiments along a region incorporating healthy and cancerous parts indicate step-like behaviors in the degree of circular polarization corresponding to the state of tissues, whether cancerous or normal. An oblique and perpendicular incidence induces different resolutions for identifying cancerous tissues, which indicates that the optical arrangement can be selected according to the priority of resolution.     

Good,better, best? The effects of polarization on photobiomodulation therapy

Photobiomodulation therapy (PBMT) is a widely adopted form of phototherapy used to treat many chronic conditions that effect the population at large. The exact physiological mechanisms of PBMT remain unsolved; however, the prevailing theory centres on changes in mitochondrial function. There are many irradiation parameters to consider when investigating PBMT, one of which is the state of polarization. There is some evidence to show that polarization of red and near‐infrared light may promote different and/or increased biological activity when compared to otherwise identical non‐polarized light. These enhanced cellular effects may also be present when the polarized light is applied linear to the tissue direction. Herein, we synthesize the current experimental and clinical evidence pertaining to polarized photobiomodulation therapy; ultimately, to better inform future research into this area of phototherapy.     

Colon cancer detection by using Poincaré sphere and 2D polarimetric mapping of ex vivo colon samples

This work is dedicated to the diagnosis and grading of colon cancer by a combined use of Poincaré sphere and 2D Stokes vector polarimetry mapping approaches. The major challenge consists in exploring the applicability of polarized light for noninvasive screening of the histological abnormalities within the samples of biological tissues. Experimental studies were conducted in ex vivo colon sample, excised after surgical procedure for colon tumor removal of G2‐adenocarcinoma lesion. Polarimetric measurements in linear and circular regime were carried via personally developed polarimetric, optical set‐up, using supercontinuous fiber laser with irradiation fixed at 635 nm. We apply the Poincaré sphere and two‐dimensional Stokes vector scanning approach for screening the corresponding tissue samples. A comparison between linear and circular polarization states is made both for quantitative and qualitative evaluations. It is shown that circular polarization has better diagnostic capabilities than linear polarization, with higher dynamic ranges of the polarimetric parameters and better values of the diagnostic quantities. In addition to the standard polarimetry parameters, utilized as essential diagnostic markers, we apply statistical analysis to obtain more detailed information in frame of the applied diagnostic approach.     

Optical coherence hyperspectral microscopy with a single supercontinuum light source

In the paper, we have developed an optical coherence hyperspectral microscopy with a single supercontinuum light source. The microscopy consists of optical coherence tomography (OCT) and hyperspectral imaging (HSI), which can visualize the structural and functional characteristics of biological tissues. The 500 to 700 nm band is selected for HSI and OCT imaging, where HSI enables imaging of oxygen saturation and hemoglobin (Hb) content, while OCT acquires structural characteristics to assess the morphology of biological tissues. The system performance of the optical coherence hyperspectral microscopy is verified by normal mice ears, and the practical applications of the microscopy is further performed in 4T1 and inflammation Balb/c mice ears in vivo. The experimental results demonstrate that the microscopy has potential to provide complementary information for clinical applications.     

Effects of the phenotypic polarization state of human leukocytes on the optical absorbance spectrum

This study evaluated the optical absorbance spectrum of human monocytes, neutrophils and lymphocytes polarized, or not, to the inflammatory or immunoregulatory phenotypes. Peripheral human blood leukocytes were isolated and polarized (10 ng/mL) with LPS or IL-4 + LPS for 2 hours. After polarization, cells were washed and incubated for an additional 24 hours (monocytes and lymphocytes) or 12 hours (neutrophils). Next, cells were collected to evaluate the optical absorbance spectrum. The three types of leukocytes exhibited absorbance in the region from 450 to 900 nm, with greater absorbance at wavelengths lower than 570 nm. Lymphocytes had a second region of greater absorbance between 770 and 900 nm. Inflammatory monocytes and lymphocytes showed increased absorbance of blue, green and yellow wavelengths (monocytes), as well as red and infrared wavelengths (monocytes and lymphocytes). Immunoregulatory polarization altered the absorbance of monocytes and lymphocytes very little. Neutrophils treated with LPS or LPS + IL-4 exhibited lower absorbance at wavelengths higher than 575 nm compared to untreated cells. The present findings showed that leukocytes exhibit greater absorbance in regions of the spectrum that have not been much used in photobiomodulation (PBM), and the polarization of these cells can affect their capacity to absorb light. Taken together, these results suggest new perspectives in the use of PBM in the clinical setting depending on the wavelengths and the stage of the inflammatory process.     

Photobiomodulation at a wavelength of 633 nm leads to faster functional recovery than 804 nm after facial nerve injury

We analyzed the effects of photobiomodulation (PBM) of various wavelengths on regeneration of the facial nerve using in vitro and in vivo experimental models. We assessed the antioxidative effect of PBM in geniculate ganglion neurons irradiated with a diode laser at 633 nm, 780 nm and 804 nm. Wavelengths of 633 and 780 nm but not 804 nm inhibited cell death by oxidative stress. We assessed the effects of PBM on functional and morphologic recovery in rats divided into control, facial nerve damage (FND) and FND irradiated with a 633 nm or 804 nm lasers. Injured rats treated with 633-nm light had better facial palsy scores, larger axon diameter and higher expression of Schwann cells compared with the FND group. No positive results were observed in rats irradiated at 804-nm light. These findings indicate that 633-nm PBM promotes accelerated nerve regeneration and improved functional recovery in an injured facial nerve.     

Instant polarized light microscopy for imaging collagen microarchitecture and dynamics

Collagen fibers are a primary load-bearing component of connective tissues and are therefore central to tissue biomechanics and pathophysiology. Understanding collagen architecture and behavior under dynamic loading requires a quantitative imaging technique with simultaneously high spatial and temporal resolutions. Suitable techniques are thus rare and often inaccessible. In this study, we present instant polarized light microscopy (IPOL), in which a single snapshot image encodes information on fiber orientation and retardance, thus fulfilling the requirement. We utilized both simulation and experimental data from collagenous tissues of chicken tendon, sheep eye, and porcine heart to evaluate the effectiveness of IPOL as a quantitative imaging technique. We demonstrate that IPOL allows quantitative characterization of micron-scale collagen fiber architecture at full camera frame rates (156 frames/second herein).     

Pathological crystal imaging with single‐shot computational polarized light microscopy

Pathological crystal identification is routinely practiced in rheumatology for diagnosing arthritis disease such as gout, and relies on polarized light microscopy as the gold standard method used by medical professionals. Here, we present a single‐shot computational polarized light microscopy method that reconstructs the transmittance, retardance and slow‐axis orientation of a birefringent sample using a single image captured with a pixelated‐polarizer camera. This method is fast, simple‐to‐operate and compatible with all the existing standard microscopes without extensive or costly modifications. We demonstrated the success of our method by imaging three different types of crystals found in synovial fluid and reconstructed the birefringence information of these samples using a single image, without being affected by the orientation of individual crystals within the sample field‐of‐view. We believe this technique will provide improved sensitivity, specificity and speed, all at low cost, for clinical diagnosis of crystals found in synovial fluid and other bodily fluids.     

Dual‐ and single‐shot susceptibility ratio measurements with circular polarizations in second‐harmonic generation microscopy

Polarization‐resolved second‐harmonic generation (P‐SHG) microscopy is a technique capable of characterizing nonlinear optical properties of noncentrosymmetric biomaterials by extracting the nonlinear susceptibility tensor components ratio , with z‐axis parallel and x‐axis perpendicular to the C₆ symmetry axis of molecular fiber, such as a myofibril or a collagen fiber. In this paper, we present two P‐SHG techniques based on incoming and outgoing circular polarization states for a fast extraction of : A dual‐shot configuration where the SHG circular anisotropy generated using incident right‐ and left‐handed circularly‐polarized light is measured; and a single‐shot configuration for which the SHG circular anisotropy is measured using only one incident circular polarization state. These techniques are used to extract the of myosin fibrils in the body wall muscles of Drosophila melanogaster larva. The results are in good agreement with values obtained from the double Stokes‐Mueller polarimetry. The dual‐ and single‐shot circular anisotropy measurements can be used for fast imaging that is independent of the in‐plane orientation of the sample. They can be used for imaging of contracting muscles, or for high throughput imaging of large sample areas.     

生物组织的折射和折射率

光在生物组织中的传播与组织的光学性质有关。光通过组织时,光强和光的偏振状态会发生变化。而折射率是组织光学用来评价组织改变光线行进方向的基本参量。本文以菲涅耳公式为理论依据,用空气一组织界面的反射率、生物组织薄膜的反射率和生物组织反射光的倔振分量,推算生物组织的折射率。     

Spatial orientation of trout to partially polarized light

Summary The results of this study reveal that rainbow trout (Oncorhyncus mykiss formerly Salmo gairdneri) are capable of orienting to polarized light fields, and that the degree of polarization of the polarized light field affects the accuracy of orientation behavior. As previously shown, rainbow trout can accurately orient to a plane polarized light field after several sessions of food-rewarded training. The present data demonstrate that the accuracy of such orientation decreases the degree of polarization of the plane-polarized light field is lowered. In testing sessions, different concentrations of latex beads were introduced into a cuvette positioned below the light source to degrade the degree of polarization. There was evidence that trout could still detect the evector and use it in making orienting responses when the light was only 65% polarized. However, most of the test trout did not demonstrate orienting ability at levels of polarization below the 75% level.     

Optical plasticity of mammalian cells

Transparency is widespread in nature, ranging from transparent insect wings to ocular tissues that enable you to read this text, and transparent marine vertebrates. And yet, cells and tissue models in biology are usually strongly light scattering and optically opaque, precluding deep optical microscopy. Here we describe the directed evolution of cultured mammalian cells toward increased transparency. We find that mutations greatly diversify the optical phenotype of Chinese Hamster Ovary cells, a cultured mammalian cell line. Furthermore, only three rounds of high-throughput optical selection and competitive growth are required to yield fit cells with greatly improved transparency. Based on 15 monoclonal cell lines derived from this directed evolution experiment, we find that the evolved transparency frequently goes along with a reduction of nuclear granularity and physiological shifts in gene expression profiles. In the future this optical plasticity of mammalian cells may facilitate genetic clearance of living tissues for in vivo microscopy.     

Orthogonal-polarization-gating optical coherence tomography for human sweat ducts in vivo

We propose an orthogonal-polarization-gating optical coherence tomography (OPG-OCT) for human sweat ducts in vivo. OPG-OCT is composed of the orthogonal linearly polarized light of a sample arm individually interfering with orthogonal linearly polarized lights of the reference arms, where OPG-OCT induces two images, one reflecting the projection intensity and the other the horizontal linear diattenuation (HLD). The results demonstrate that OPG-OCT projection intensity could improve the image quality of sweat ducts. HLD also clearly illustrates the spiral shape of the sweat ducts. Finally, sweat ducts in intensity image are segmented by employing convolutional neural networks (CNN). The proportions of left-handed and right-handed ducts are extracted to characterize the sweat ducts based on HLD. Therefore, the OPG-OCT technique employing CNN for the human sweat glands has the potential to automatically identify the human sweat ducts in vivo.     

Automatic imaging of Drosophila embryos with light sheet fluorescence microscopy on chip

We present a microscope on chip for automated imaging of Drosophila embryos by light sheet fluorescence microscopy. This integrated device, constituted by both optical and microfluidic components, allows the automatic acquisition of a 3D stack of images for specimens diluted in a liquid suspension. The device has been fully optimized to address the challenges related to the specimens under investigation. Indeed, the thickness and the high ellipticity of Drosophila embryos can degrade the image quality. In this regard, optical and fluidic optimization has been carried out to implement dual-sided illumination and automatic sample orientation. In addition, we highlight the dual color investigation capabilities of this device, by processing two sample populations encoding different fluorescent proteins. This work was made possible by the versatility of the used fabrication technique, femtosecond laser micromachining, which allows straightforward fabrication of both optical and fluidic components in glass substrates.     

Hyperspectral imaging as a diagnostic tool to differentiate between amalgam tattoos and other dark pigmented intraoral lesions

The goal of this project is to identify any in-depth benefits and drawbacks in the diagnosis of amalgam tattoos and other pigmented intraoral lesions using hyperspectral imagery collected from amalgam tattoos, benign, and malignant melanocytic neoplasms. Software solutions capable of classifying pigmented lesions of the skin already exist, but conventional red, green and blue images may be reaching an upper limit in their performance. Emerging technologies, such as hyperspectral imaging (HSI) utilize more than a hundred, continuous data channels, while also collecting data in the infrared. A total of 18 paraffin-embedded human tissue specimens of dark pigmented intraoral lesions (including the lip) were analyzed using visible and near-infrared (VIS–NIR) hyperspectral imagery obtained from HE-stained histopathological slides. Transmittance data were collected between 450 and 900 nm using a snapshot camera mounted to a microscope with a halogen light source. VIS–NIR spectra collected from different specimens, such as melanocytic cells and other tissues (eg, epithelium), produced distinct and diagnostic spectra that were used to identify these materials in several regions of interest, making it possible to distinguish between intraoral amalgam tattoos (intramucosal metallic foreign bodies) and melanocytic lesions of the intraoral mucosa and the lip (each with P < .01 using the independent t test). HSI is presented as a diagnostic tool for the rapidly growing field of digital pathology. In this preliminary study, amalgam tattoos were reliably differentiated from melanocytic lesions of the oral cavity and the lip.     

Simulation and Analytical Study of Optical Complex Field in Nano-corral Slits Plasmonic Lens

Although spiral plasmonic lens has been proposed as circular polarization analyzer, there is no such plasmonic nanostructure available for linear polarization. In the current work, we have designed nano-corral slits (NCS) plasmonic lens, which focuses the x- and y-polarized light into spatially distinguished plasmonic fields. We have calculated analytically and numerically the electric field intensity and phase of the emission from nano-corral slits plasmonic lens with different pitch lengths under various polarizations of the illumination. It has been shown that one can control the wave front of the output beam of these plasmonic lenses by manipulating the illumination of both circular and linear polarization. Our theoretical study in correlation with FDTD simulation has shown that NCS plasmonic lens with pitch length equal to λ_spp produces scalar vortex beam having optical complex fields with helical wave front and optical singularity at the center under circular polarization of light. When NCS lens (pitch = λ_spp) is illuminated with linearly polarized light, it exhibits binary distribution of phase with same electric field intensity around the center. However, with pitch length of 0.5λ_spp, NCS shows linear dichroism under linearly polarized illumination unlike spiral plasmonic lens (SPL) eliminating the use of circularly polarized light. Optical complex fields produced by these NCS plasmonic lenses may find applications for faster quantum computing, data storage, and telecommunications.

     

Computed tomography for in vivo deep over‐1000 nm near‐infrared fluorescence imaging

This study aims to develop a novel cross‐sectional imaging of fluorescence in over‐1000 nm near‐infrared (OTN‐NIR), which allows in vivo deep imaging, using computed tomography (CT) system. Cylindrical specimens of composite of OTN‐NIR fluorophore, NaGdF₄ co‐doped with Yb³⁺ and Ho³⁺ (ex: 980 nm, em: 1150 nm), were embedded in cubic agar (10.5–12 mm) or in the peritoneal cavity of mice and placed on a rotatable stage. When the fluorescence from inside of the samples was serially captured from multiple angles, the images were disrupted by the reflection and refraction of emitted light on the sample‐air interface. Immersing the sample into water filled in a rectangular bath suppressed the disruption at the interface and successfully reconstructed the position and concentration of OTN‐NIR fluorophores on the cross‐sectional images using a CT technique. This is promising as a novel three‐dimensional imaging technique for OTN‐NIR fluorescent image projections of small animals captured from multiple angles.     

Cone photoreceptor mechanisms and the detection of polarized light in fish

Summary Although numerous studies have demonstrated the detection of polarized light in vertebrates, little is known of the photoreceptor mechanisms involved. Recent evidence, however, indicates that cyprinid fishes possess both ultraviolet (UV) and polarization sensitivity suggesting that some vertebrates, like many invertebrates, may employ UV-sensitive cone receptors in polarization sensitivity. In this report, we describe experiments that determine which spectral types of receptors participate in the detection of polarized light. We used a heart-rate conditioning technique to measure increment thresholds of immobilized goldfish for plane-polarized, narrow-band (10 nm half max.) spectral stimuli (380 nm, 460 nm, 540 nm, 660 nm). A typical experiment involved isolating the activity of a cone photoreceptor mechanism by chromatic adaptation and measuring increment thresholds for spectral stimuli at e-vector orientations of the polarizer between 0° to 180° in 30° steps. The UV-, green- and red-sensitive cone receptor mechanisms showed clear evidence of polarization sensitivity while the blue-sensitive cone receptor mechanism was polarizationally insensitive. The average amplitude (base to peak height on Fig. 4) of the polarization sensitivity curves (UV-, green- and red-curves) was 0.67 log unit (standard deviation of 0.12 log unit), with the UV-sensitive cone receptor mechanism most sensitive to the vertical e-vector axis and the green- and red-sensitive cone receptor mechanisms most sensitive to the horizontal e-vector axis. The observation that different cone photoreceptor mechanisms have orthogonal polarization sensitivity in fish suggests that the perception of polarized light may enhance the capacity for visual discrimination in lower vertebrates.     

Excitation energy transfer and chlorophyll orientation in the green alga Chlamydomonas reinhardi

We have investigated the process of intermolecular excitation energy transfer and the relative orientation of the chlorophyll molecules in the unicellular green alga Chlamydomonas reinhardi. The principal experiments involved in vivo measurements of the fluorescence polarization as a function of the exciting-light wavelength in the presence and in the absence of 3-(3,4-dichlorophenyl)-1,1-dimethylurea. We found that as the fluorescence lifetime increases upon the addition of 3-(3,4-dichlorophenyl)-1,1-dimethylurea that the degree of fluorescence polarization decreases over the excitation region from 600 to 660 nm. This result, we argue, implies that a Förster mechanism of excitation energy transfer is involved for Photosystem II chlorophyll molecules absorbing primarily below 660 nm. We must add that our results do not exclude the possibility of a delocalized transfer process from being involved as well. Fluorescence polarization measurements using chloroplast fragments are also discussed in terms of a Förster transfer mechanism. As the excitation wavelength approaches 670 nm the fluorescence polarization is nearly constant upon the addition of 3-(3,4-dichlorophenyl)-1,1-dimethylurea.Experiments performed using either vertically or horizontally polarized exciting light show that the fluorescence polarization increases as the exciting light wavelength increases from 650 to 673 nm. This suggests the possibility that chlorophyll molecules absorbing at longer wavelengths have a higher degree of relative order. Furthermore, these studies imply that chlorophyll molecules exist in discrete groups that are characterized by different absorption maxima and by different degrees of the fluorescence polarization. In view of these results we discuss different models for the Photosystem II antenna system and energy transfer between different groups of optically distinguishable chlorophyll molecules.     

Comparison between high definition FT‐IR,Raman and AFM‐IR for subcellular chemical imaging of cholesteryl esters in prostate cancer cells

The family of vibrational spectroscopic imaging techniques grows every few years and there is a need to compare and contrast new modalities with the better understood ones, especially in the case of demanding biological samples. Three vibrational spectroscopy techniques (high definition Fourier‐transform infrared [FT‐IR], Raman and atomic force microscopy infrared [AFM‐IR]) were applied for subcellular chemical imaging of cholesteryl esters in PC‐3 prostate cancer cells. The techniques were compared and contrasted in terms of image quality, spectral pattern and chemical information. All tested techniques were found to be useful in chemical imaging of cholesterol derivatives in cancer cells. The results obtained from FT‐IR and Raman imaging showed to be comparable, whereas those achieved from AFM‐IR study exhibited higher spectral heterogeneity. It confirms AFM‐IR method as a powerful tool in local chemical imaging of cells at the nanoscale level. Furthermore, due to polarization effect, p‐polarized AFM‐IR spectra showed strong enhancement of lipid bands when compared to FT‐IR.