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Raman microspectroscopy has become an effective method for analyzing the molecular appearance of biomarkers in skin tissue. For the first time, we acquired in vitro Raman spectra of healthy and malignant skin tissues, including basal cell carcinoma (BCC) and squamous cell carcinoma (SCC), at 532 and 785 nm laser excitation wavelengths in the wavenumber ranges of 900-1800 cm-1 and 2800-3100 cm-1 and analyzed them to find spectral features for differentiation between the three classes of the samples. The intensity ratios of the bands at 1268, 1336, and 1445 cm-1 appeared to be the most reliable criteria for the three-class differentiation at 532 nm excitation, whereas the bands from the higher wavenumber region (2850, 2880, and 2930 cm-1) were a robust measure of the increased protein/lipid ratio in the tumors at both excitation wavelengths. Selecting ratios of the three bands from the merged (532 + 785) dataset made it possible to increase the accuracy to 87% for the three classes and reach the specificities for BCC + SCC equal to 87% and 81% for the sensitivities of 95% and 99%, respectively. Development of multi-wavelength excitation Raman spectroscopic techniques provides a versatile non-invasive tool for research of the processes in malignant skin tumors, as well as other forms of cancer.
Assuntos
Carcinoma Basocelular , Carcinoma de Células Escamosas , Neoplasias Cutâneas , Análise Espectral Raman , Análise Espectral Raman/métodos , Humanos , Neoplasias Cutâneas/patologia , Neoplasias Cutâneas/diagnóstico , Neoplasias Cutâneas/metabolismo , Carcinoma Basocelular/diagnóstico , Carcinoma Basocelular/patologia , Carcinoma Basocelular/metabolismo , Carcinoma de Células Escamosas/diagnóstico , Carcinoma de Células Escamosas/patologia , Carcinoma de Células Escamosas/metabolismo , Feminino , Masculino , Pessoa de Meia-Idade , Pele/patologia , Pele/metabolismo , IdosoRESUMO
Inscription of embedded photoluminescent microbits inside micromechanically positioned bulk natural diamond, LiF and CaF2 crystals was performed in sub-filamentation (geometrical focusing) regime by 525 nm 0.2 ps laser pulses focused by 0.65 NA micro-objective as a function of pulse energy, exposure and inter-layer separation. The resulting microbits were visualized by 3D-scanning confocal Raman/photoluminescence microscopy as conglomerates of photo-induced quasi-molecular color centers and tested regarding their spatial resolution and thermal stability via high-temperature annealing. Minimal lateral and longitudinal microbit separations, enabling their robust optical read-out through micromechanical positioning, were measured in the most promising crystalline material, LiF, as 1.5 and 13 microns, respectively, to be improved regarding information storage capacity by more elaborate focusing systems. These findings pave a way to novel optomechanical memory storage platforms, utilizing ultrashort-pulse laser inscription of photoluminescent microbits as carriers of archival memory.
RESUMO
Tightly focused 515-nm, 0.3-ps laser pulses modify in a laser filamentation regime the crystalline structure of an Ib-type high-pressure, high-temperature (HPHT) synthesized diamond in a thin-plate form. The modified microregions (micromarks) in the yellow and colorless crystal zones, possessing different concentrations of elementary substitutional nitrogen (N) impurity atoms (C-centers), exhibit their strongly diminished local IR absorption (upon correction to the thickness scaling factor). Simultaneously, local visible-range (400-550 nm) absorption coefficients were increased, and photoluminescence (PL) yield was strongly enhanced in the broad range of 450-800 nm. The strong yellow-red PL enhancement saturates with laser exposure, implying the complete conversion of C-centers into nitrogen-vacancy (NV0,-) ones due to the laser-induced generation of Frenkel "interstitial-vacancy" I-V carbon pairs. The other emerging blue-green (>470 nm) and green-yellow (>500 nm) PL bands were also simultaneously saturated versus the laser exposure. The observed IR/optical absorption and PL spectral changes enlighten the ultrashort pulse laser inscription of NV0--based quantum-emitter centers in synthetic diamonds and enable the evaluation of the productivity of their inscription along with the corresponding I-V generation rates.
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An ultrashort-pulse laser inscription of embedded birefringent microelements was performed inside bulk fluorite in pre-filamentation (geometrical focusing) and filamentation regimes as a function of laser wavelength, pulsewidth and energy. The resulting elements composed of anisotropic nanolattices were characterized by retardance (Ret) and thickness (T) quantities, using polarimetric and 3D-scanning confocal photoluminescence microscopy, respectively. Both parameters exhibit a monotonous increase versus pulse energy, going over a maximum at 1-ps pulsewidth at 515 nm, but decrease versus laser pulsewidth at 1030 nm. The resulting refractive-index difference (RID) Δn = Ret/T ~ 1 × 10-3 remains almost constant versus pulse energy and slightly decreases at a higher pulsewidth, generally being higher at 515 nm. The birefringent microelements were visualized using scanning electron microscopy and chemically characterized using energy-dispersion X-ray spectroscopy, indicating the increase of calcium and the contrary decrease of fluorine inside them due to the non-ablative inscription character. Dynamic far-field optical diffraction of the inscribing ultrashort laser pulses also demonstrated the accumulative inscription character, depending on the pulse energy and the laser exposure. Our findings revealed the underlying optical and material inscription processes and demonstrated the robust longitudinal homogeneity of the inscribed birefringent microstructures and the facile scalability of their thickness-dependent retardance.
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A rather narrow field of view (FOV) has always been considered as an essential limitation of spectral imagers based on acousto-optical tunable filters (AOTFs). We demonstrate a computational technique to overcome this constraint. It is based on preliminary precise spectral-angular characterization of beam transformation caused by light diffraction on an acoustic wave and consequent correction of acquired stack of spectral images. This technique is applicable for any geometry of acousto-optic interaction and opens the way for the development of AOTFs with significantly expanded FOV.
Assuntos
Óptica e FotônicaRESUMO
We report on wavelength-multiplexed digital holographic imaging based on simultaneous Bragg diffraction of wideband light by several ultrasound waves of different frequencies in crystalline media. This technique is easy to implement, avoids spectral scanning, and is applicable in various digital holography schemes. It also enables single-shot acquisition of a few spectral fringe patterns by a single monochrome sensor and wavelength demultiplexing of the resulting interferogram. We have assembled a Mach-Zehnder interferometer with an acousto-optical tunable filter operating in the multifrequency mode and have validated the proposed technique by dual-, three-, and four-wavelength quantitative phase imaging of the test samples.
RESUMO
Spectral image filtration by means of acousto-optical tunable filters (AOTFs) has multiple applications. For its implementation, a few different optical schemes are in use. They differ in image quality, number of coupling components, dimensions and alignment complexity. To choose the optical system of AOTF-based spectral imager properly, many factors have to be considered. Though various schemes of acousto-optic (AO) filtration have been tested and discussed, their comparative analysis has not been reported up to now. In this study, we assembled the four most popular schemes (confocal, collimating, tandem and double-path) using the same AO cells and experimentally compared their main features. Depending on the application, each scheme may be the basis of compact cost-effective spectral imaging devices.
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We address the optical design procedure of prism-based stereoscopic imaging systems. Conventional approach includes two sequential stages: selection of the hardware and development of the proper digital image processing algorithms. At each of these stages, specific techniques are applied, which are almost unrelated to each other. The main requirements to the imaging system include only the key parameters and the image quality. Therefore, the insufficient measurement accuracy may be revealed only after the prototype is assembled and tested. In this case, even applying complex time-consuming image processing and calibration procedures does not ensure the necessary precision. A radical solution of this issue is to include the measurement error estimation into the optical design stage. In this research, we discuss a simplified implementation of this approach and demonstrate the capabilities of optical design software for this purpose. We demonstrate the effectiveness of this approach by the analysis and optimization of a prism-based stereoscopic imager with respect to required 3D measurement accuracy. The results are meaningful for the development of 3D imaging techniques for machine vision, endoscopic and measurement systems.
RESUMO
Stereoscopic video endoscopes are widely used for remote visual inspection and precise three-dimensional (3D) measurements in industrial and biomedical applications. The reconstruction of 3D points from the corresponding image points requires a geometrical calibration procedure, the accuracy of which affects the measurement uncertainty. We propose to perform an optimal choice of the calibration technique and the calibration target parameters using a computer simulation at the design stage. The effectiveness of this approach is demonstrated via the design of a self-developed miniature prism-based stereoscopic system. We simulated acquisition of calibration and measurement data using optical design software. The conventional calibration technique, requiring many positions of the flat target with arbitrary displacements and rotations, was compared with another one that uses the translation stage to provide pure translation of the target. We analyzed the impact of the translation uncertainty, the number of positions, the number of targets, and the uncertainty of image point coordinates on the uncertainty of calibration parameters and 3D measurements. We have shown that the second technique could provide acceptable measurement accuracy using only two images. The results of the computer simulation were confirmed experimentally using the prototype of the stereoscopic endoscope. The proposed approach may be used to optimize calibration techniques and reduce the cost of calibration equipment for various stereoscopic measurement systems.
RESUMO
Stereoscopic imagers are widely used in machine vision for three-dimensional (3D) visualization as well as in non-destructive testing for quantitative characterization of cracks, delamination and other defects. Measurement capability in these systems is provided by a proper combination of the optical parameters and data processing techniques. Conventional approach to their design consists of two sequential stages: optical system design and optimization of calibration and image processing algorithms. Such two-stage procedure often complicates both the hardware and the software, and results in a time-ineffective design procedure and cost-ineffective solution. We demonstrate a more effective approach and propose to estimate errors of 3D measurements at the early optical design stage. We show that computer simulation using optical design software allows not only optimizing optical parameters of the imager but also choosing the most effective mathematical model of the system and the equipment necessary for calibration. We tested the proposed approach on the design of miniature prism-based stereoscopic system and analyzed the impact of various factors (aberrations, tolerances, etc.) as on the image quality, so on the quality of calibration and 3D measurements accuracy. The proposed joint design approach may be highly effective for various measurement systems and applications when both optical parameters and image processing algorithms are not defined in advance and are necessary to be optimized.
RESUMO
We propose a new technique for three-dimensional (3-D) imaging in arbitrary spectral intervals. It is based on a simultaneous diffraction of two divergent stereoscopic light beams on a single acoustic wave propagating in a uniaxial birefringent crystal. We discuss in detail this configuration of acousto-optic (AO) interaction, derive basic relations, and experimentally demonstrate the applicability of the proposed approach to 3-D spectral imaging. A stereo-imager of this type may be produced as an ultra-compact embeddable optical element, which is promising for many imaging applications.