Spiral-phase-objective for a compact spiral-phase-contrast microscopy.

Spiral-phase-contrast imaging, which utilizes a spiral phase optical element, has proven to be effective in enhancing various aspects of imaging, such as edge contrast and shadow imaging. Typically, the implementation of spiral-phase-contrast imaging requires the formation of a Fourier plane through a 4f optical configuration in addition to an existing optical microscope. In this study, we present what we believe to be a novel single spiral-phase-objective, integrating a spiral phase plate, which can be easily and simply applied to a standard microscope, such as a conventional objective. Using a new hybrid design approach that combines ray-tracing and field-tracing simulations, we theoretically realized a well-defined and high-quality vortex beam through the spiral-phase-objective. The spiral-phase-objective was designed to have conditions that are practically manufacturable while providing predictable performance. To evaluate its capabilities, we utilized the designed spiral-phase-objective to investigate isotropic spiral phase contrast and anisotropic shadow imaging through field-tracing simulations, and explored the variation of edge contrast caused by changes in the thickness of the imaging object.

Spectrally encoded dual-mode interferometry with orthogonal scanning.

Spectrally encoded confocal microscopy (SECM) is a high-speed reflectance confocal microscopy technique. Here, we present a method to integrate optical coherence tomography (OCT) and SECM for complementary imaging by adding orthogonal scanning to the SECM configuration. The co-registration of SECM and OCT is automatic, as all system components are shared in the same order, eliminating the need for additional optical alignment. The proposed multimode imaging system is compact and cost-effective while providing the benefits of imaging aiming and guidance. Furthermore, speckle noise can be suppressed by averaging the speckles generated by shifting the spectral-encoded field in the direction of dispersion. Using a near infrared (NIR) card and a biological sample, we demonstrated the capability of the proposed system by showing SECM imaging at depths of interest guided by the OCT in real time and speckle noise reduction. Interfaced multimodal imaging of SECM and OCT was implemented at a speed of approximately 7 frames/s using fast-switching technology and GPU processing.

Label-free photothermal optical coherence microscopy to locate desired regions of interest in multiphoton imaging of volumetric specimens.

Biochip-based research is currently evolving into a three-dimensional and large-scale basis similar to the in vivo microenvironment. For the long-term live and high-resolution imaging in these specimens, nonlinear microscopy capable of label-free and multiscale imaging is becoming increasingly important. Combination with non-destructive contrast imaging will be useful for effectively locating regions of interest (ROI) in large specimens and consequently minimizing photodamage. In this study, a label-free photothermal optical coherence microscopy (OCM) serves as a new approach to locate the desired ROI within biological samples which are under investigation by multiphoton microscopy (MPM). The weak photothermal perturbation in sample by the MPM laser with reduced power was detected at the endogenous photothermal particles within the ROI using the highly sensitive phase-differentiated photothermal (PD-PT) OCM. By monitoring the temporal change of the photothermal response signal of the PD-PT OCM, the hotspot generated within the sample focused by the MPM laser was located on the ROI. Combined with automated sample movement in the x-y axis, the focal plane of MPM could be effectively navigated to the desired portion of a volumetric sample for high-resolution targeted MPM imaging. We demonstrated the feasibility of the proposed method in second harmonic generation microscopy using two phantom samples and a biological sample, a fixed insect on microscope slide, with dimensions of 4 mm wide, 4 mm long, and 1 mm thick.

Wavelength-tunable spiral-phase-contrast imaging.

Wavelength-tunable spiral-phase-contrast (SPC) imaging was experimentally accomplished in the visible wavelengths spanning a broad bandwidth of ∼200 nm based on a single off-axis spiral phase mirror (OSPM). By the rotation of an OSPM, which was designed with an integer orbital angular momentum (OAM) of l = 1 at a wavelength of 561 nm and incidence angle of 45°, high-quality SPC imaging was obtained at different wavelengths. For the comparison with wavelength-tunable SPC imaging using an OSPM, SPC imaging using a spiral phase plate (manufactured to generate an OAM of l = 1 at 561 nm) was performed at three wavelengths (473, 561, and 660 nm), resulting in clear differences. Theoretically, based on field tracing simulations, high-quality wavelength-tunable SPC imaging could be demonstrated in a very broad bandwidth of ∼400 nm, which is beyond the bandwidth of ∼200 nm obtained experimentally. This technique contribute to developing high-performance wavelength-tunable SPC imaging by simply integrating an OSPM into the current optical imaging technologies.

Generation of wavelength-tunable optical vortices using an off-axis spiral phase mirror: publisher's note.

This publisher's note contains corrections to Opt. Lett.46, 4216 (2021)OPLEDP0146-959210.1364/OL.432413.

Generation of wavelength-tunable optical vortices using an off-axis spiral phase mirror.

Wavelength-tunable optical vortices with a topological charge equal to l=1 of orbital angular momentum (OAM) were experimentally realized using a single off-axis spiral phase mirror (OSPM) with lasers of various visible-light wavelengths. Using an OSPM designed for 561 nm and an incidence angle of 45°, circular doughnut-shaped l=1 optical vortices were obtained at 561, 473, and 660 nm by rotating the OSPM to modify the laser incidence angle. Wavelength-tunable l=1 optical vortices were obtained at the respective incidence angles of 45°, 53.4°, and 33.7°, because the effective geometrical thickness of the OSPM, which determines the order of OAM, was identical at each wavelength. This flexible OSPM which operates over a wide wavelength range will provide continuously wavelength-tunable optical vortices for applications in the fields of advanced optics and photonics in which optical vortices with wide wavelength tunability are in demand.

Quantitative Photothermal Characterization with Bioprinted 3D Complex Tissue Constructs for Early-Stage Breast Cancer Therapy Using Gold Nanorods.

Plasmonic photothermal therapy (PPTT) using gold nanoparticles (AuNPs) has shown great potential for use in selective tumor treatment, because the AuNPs can generate destructive heat preferentially upon irradiation. However, PPTT using AuNPs has not been added to practice, owing to insufficient heating methods and tissue temperature measurement techniques, leading to unreliable and inaccurate treatments. Because the photothermal properties of AuNPs vary with laser power, particle optical density, and tissue depth, the accurate prediction of heat generation is indispensable for clinical treatment. In this report, bioprinted 3D complex tissue constructs comprising processed gel obtained from porcine skin and human decellularized adipose tissue are presented for characterization of the photothermal properties of gold nanorods (AuNRs) having an aspect ratio of 3.7 irradiated by a near-infrared laser. Moreover, an analytical function is suggested for achieving PPTT that can cause thermal damage selectively on early-stage human breast cancer by regulating the heat generation of the AuNRs in the tissue.

Design, Fabrication, and Performance Evaluation of Portable and Large-Area Blackbody System.

In this study, a portable and large-area blackbody system was developed following a series of processes including design, computational analysis, fabrication, and experimental analysis and evaluation. The blackbody system was designed to be lightweight (5 kg), and its temperature could exceed the ambient temperature by up to 15 °C under operation. A carbon-fiber-based heat source was used to achieve a uniform temperature distribution. A heat shield fabricated from an insulation material was embedded at the opposite side of the heating element to minimize heat loss. A prototype of the blackbody system was fabricated based on the design and transient coupled electro-thermal simulation results. The operation performance of this system, such as the thermal response, signal transfer function, and noise equivalent temperature difference, was evaluated by employing an infrared imaging system. In addition, emissivity was measured during operation. The results of this study show that the developed portable and large-area blackbody system can be expected to serve as a reliable reference source for the calibration of aerial infrared images for the application of aerial infrared techniques to remote sensing.

Estimation and compensation of phase errors induced by axial bulk motion of a sample in wavelength-sweeping parallel Fourier domain OCT.

We report a new, to the best of our knowledge, approach to correct image blurring due to the axial bulk motion of a sample in wavelength-sweeping Fourier domain parallel optical coherence tomography (OCT). This approach can estimate phase errors changing rapidly in time through direct measurements of the apparent axial shift during the sampling interval using common phase changes in parallel detection without additional hardware. To demonstrate the performance of the proposed algorithm, a single reflection and scattering sample were imaged with wavelength-sweeping parallel OCT implemented by scanning a spectrally dispersed line-field along the line direction. In addition, we quantitatively demonstrated that even a small axial movement of the sample could cause serious image blur at a high nonlinear degree of movement.

Bragg scattering from a millimeter-scale periodic structure with extremely small aspect ratios.

The periodic structure on the optical surface affects the beam shape and its propagation. As the size of the optical elements becomes larger and its shape becomes complicated, the quantitative analysis of the effect of the periodic structure on the optical surface becomes indispensable given that it is very difficult to completely eliminate the microscopic periodic structures. Herein, we have experimentally investigated Bragg scattering from an optical surface with extremely small aspect ratios (~10-5) and groove densities (0.5 lines/mm). We observed the period of the constructive interference formed due to the propagation of the 0th, 1st, and -1st beam modes caused by Bragg scattering. When the periodic structure has a modulation depth of ± 50 nm, the intensity increase of constructive interference between the beam modes formed by Bragg scattering was > 10 times greater than the intensity of a flat surface at the propagation distance at which constructive interference was most pronounced. This study is envisaged to open new avenues for the quantification of the effect of periodic structures based on the observation of the interference on the beam profile formed by Bragg scattering during the beam propagation.

HDAC6 regulates thermogenesis of brown adipocytes through activating PKA to induce UCP1 expression.

Mitochondrial uncoupling protein 1 (UCP1) is responsible for nonshivering thermogenesis in brown adipose tissue (BAT). UCP1 increases the conductance of the inner mitochondrial membrane (IMM) for protons to make BAT mitochondria generate heat rather than ATP. HDAC6 is a cytosolic deacetylase for non-histone substrates to regulate various cellular processes, including mitochondrial quality control and dynamics. Here, we showed that the body temperature of HDAC6 knockout mice is slightly decreased in normal hosing condition. Interestingly, UCP1 was downregulated in BAT of HDAC6 knockout mice, which extensively linked mitochondrial thermogenesis. Mechanistically, we showed that cAMP-PKA signaling plays a key role in HDAC6-dependent UCP1 expression. Notably, the size of brown adipocytes and lipid droplets in HDAC6 knockout BAT is increased. Taken together, our findings suggested that HDAC6 contributes to mitochondrial thermogenesis in BAT by increasing UCP1 expression through cAMP-PKA signaling pathway.

Laser Scanning Confocal Thermoreflectance Microscope for the Backside Thermal Imaging of Microelectronic Devices.

In this paper, we report on a confocal thermoreflectance imaging system that can examine the thermal characteristics of microelectronic devices by penetrating the backside of a device through the substrate. In this system, the local reflectivity variations due to heat generation in the device are measured point by point by a laser scanning confocal microscope capable of eliminating out-of-focus reflections and the thermoreflectance is extracted via Fourier-domain signal processing. In comparison to the conventional widefield thermoreflectance microscope, the proposed laser scanning confocal thermoreflectance microscope improves the thermoreflectance sensitivity by ~23 times and the spatial resolution by ~25% in backside thermoreflectance measurements.

3D Defect Localization on Exothermic Faults within Multi-Layered Structures Using Lock-In Thermography: An Experimental and Numerical Approach.

Micro-electronic devices are increasingly incorporating miniature multi-layered integrated architectures. However, the localization of faults in three-dimensional structure remains challenging. This study involved the experimental and numerical estimation of the depth of a thermally active heating source buried in multi-layered silicon wafer architecture by using both phase information from an infrared microscopy and finite element simulation. Infrared images were acquired and real-time processed by a lock-in method. It is well known that the lock-in method can increasingly improve detection performance by enhancing the spatial and thermal resolution of measurements. Operational principle of the lock-in method is discussed, and it is represented that phase shift of the thermal emission from a silicon wafer stacked heat source chip (SSHSC) specimen can provide good metrics for the depth of the heat source buried in SSHSCs. Depth was also estimated by analyzing the transient thermal responses using the coupled electro-thermal simulations. Furthermore, the effects of the volumetric heat source configuration mimicking the 3D through silicon via integration package were investigated. Both the infrared microscopic imaging with the lock-in method and FE simulation were potentially useful for 3D isolation of exothermic faults and their depth estimation for multi-layered structures, especially in packaged semiconductors.

Spectrally encoded common-path fiber-optic-based parallel optical coherence tomography.

We demonstrate a fiber-optic-based parallel optical coherence tomography (OCT) using spectrally encoded extended illumination with a common-path handheld probe, where the flexibility and robustness of the system are significantly improved, which is critical in the clinical environment. To the best of our knowledge, we present the first parallel OCT based on fiber optics including a fiber coupler with a sensitivity of 94 dB, which is comparable to that of point-scanning OCT. We also investigated the effect of the phase stability of the fiber-based interferometry on the parallel OCT system by comparing the common-path OCT with two-arm OCT. Using the homemade common-path handheld probe based on a Mirau interferometer, the phase stability was 32 times better than that of the two-arm OCT. The axial resolution of the common-path OCT was measured as 5.1±0.3 µm. To demonstrate the in vivo imaging performance of the fiber-optic-based parallel OCT, human skin was imaged.

Parallelized multi-graphics processing unit framework for high-speed Gabor-domain optical coherence microscopy.

Gabor-domain optical coherence microscopy (GD-OCM) is a volumetric high-resolution technique capable of acquiring three-dimensional (3-D) skin images with histological resolution. Real-time image processing is needed to enable GD-OCM imaging in a clinical setting. We present a parallelized and scalable multi-graphics processing unit (GPU) computing framework for real-time GD-OCM image processing. A parallelized control mechanism was developed to individually assign computation tasks to each of the GPUs. For each GPU, the optimal number of amplitude-scans (A-scans) to be processed in parallel was selected to maximize GPU memory usage and core throughput. We investigated five computing architectures for computational speed-up in processing 1000×1000 A-scans. The proposed parallelized multi-GPU computing framework enables processing at a computational speed faster than the GD-OCM image acquisition, thereby facilitating high-speed GD-OCM imaging in a clinical setting. Using two parallelized GPUs, the image processing of a 1×1×0.6 mm3 skin sample was performed in about 13 s, and the performance was benchmarked at 6.5 s with four GPUs. This work thus demonstrates that 3-D GD-OCM data may be displayed in real-time to the examiner using parallelized GPU processing.

Maximum-likelihood estimation in Optical Coherence Tomography in the context of the tear film dynamics.

Understanding tear film dynamics is a prerequisite for advancing the management of Dry Eye Disease (DED). In this paper, we discuss the use of optical coherence tomography (OCT) and statistical decision theory to analyze the tear film dynamics of a digital phantom. We implement a maximum-likelihood (ML) estimator to interpret OCT data based on mathematical models of Fourier-Domain OCT and the tear film. With the methodology of task-based assessment, we quantify the tradeoffs among key imaging system parameters. We find, on the assumption that the broadband light source is characterized by circular Gaussian statistics, ML estimates of 40 nm +/- 4 nm for an axial resolution of 1 µm and an integration time of 5 µs. Finally, the estimator is validated with a digital phantom of tear film dynamics, which reveals estimates of nanometer precision.

Phantom study of tear film dynamics with optical coherence tomography and maximum-likelihood estimation.

In this Letter, we implement a maximum-likelihood estimator to interpret optical coherence tomography (OCT) data for the first time, based on Fourier-domain OCT and a two-interface tear film model. We use the root mean square error as a figure of merit to quantify the system performance of estimating the tear film thickness. With the methodology of task-based assessment, we study the trade-off between system imaging speed (temporal resolution of the dynamics) and the precision of the estimation. Finally, the estimator is validated with a digital tear-film dynamics phantom.

Three-dimensional imaging of normal skin and nonmelanoma skin cancer with cellular resolution using Gabor domain optical coherence microscopy.

We investigate morphological differences in three-dimensional (3-D) images with cellular resolution between nonmelanoma skin cancer and normal skin using Gabor domain optical coherence microscopy. As a result, we show for the first time cellular optical coherence images of 3-D features differentiating cancerous skin from normal skin. In addition, in vivo volumetric images of normal skin from different anatomic locations are shown and compared.

Observation of the Gouy phase anomaly in astigmatic beams.

The Gouy phase anomaly, well established for stigmatic beams, is validated here for astigmatic beams. We simulate the predicted Gouy phase anomaly near astigmatic foci using a beam propagation algorithm integrated within lens design software. We then compare computational results with experimental data acquired using a modified Mertz-Sagnac interferometer. Both in simulation and in experiment, results show that a π/2-phase change occurs as the beam passes through each of the astigmatic foci, experimentally validating results derived in a recent paper by Visser and Wolf [Opt. Commun. 283, 3371-3375 (2010)].

Virtual skin biopsy with Gabor Domain optical coherence microscopy.

We report in-vivo volumetric optical coherence microscopy images of the skin, with resolution at the cellular level. With resolution of 2 µm both laterally and axially, structures below the skin as deep as 1 mm may be imaged at various anatomic locations. Custom optical instrumentation was designed, built, and integrated to achieve this unprecedented optical imaging resolution, in three dimensions, at clinically feasible configuration and speed.