Quantitative Phase Contrast Imaging Research Articles

Simple implementation of aperture modulation quantitative differential phase contrast imaging

As one of the quantitative phase imaging methods, aperture modulation quantitative differential phase contrast (amqDPC) imaging has isotropic lateral resolution, being widely used in many fields such as biological cell morphology detection. However, the devices required to implement it are poorly controlled and very expensive. In addition, due to the weak phase approximation, there is a large axial phase error, and the full spectrum center pass during aperture modulation will cause distortion. Therefore, a method that enables simple and low-cost implementation of amqDPC, high axial phase accuracy, and isotopic lateral resolution is reported. Through simulations and experimental verifications, axial accuracy is greatly improved in the phase range of [0, π], and an isotropic lateral resolution of 870 nm can be achieved. In addition, experiments on transparent glass circular step sample and mouse kidney tissue slice also prove the practicability of this method. According to this study, researchers can accurately and simply measure the axial height of a slightly thick sample, which is achieved by capturing two intensity images to reconstruct a quantitative phase.

Investigation of an Improved Angular Spectrum Method Based on Holography

Digital holography (DH) is a novel, real-time, non-destructive, and quantitative phase-contrast imaging method that is particularly suitable for label-free live biological cell imaging and real-time dynamic monitoring. It is currently a research hotspot in the interdisciplinary field of optics and biomedical sciences, both domestically and internationally. This article proposes an improved angle spectrum algorithm based on holographic technology, which reconstructs a cellular hologram based on phase information. Optical images and chromosome cell images, reconstructed using holographic technology at different diffraction distances under the improved angle spectrum algorithm, were analyzed and compared. The optimal diffraction distance for reconstructing chromosome cell images was selected, and chromosome cell images reproduced using traditional angle spectrum algorithms, angle spectrum algorithms combined with GS, and improved angle spectrum algorithms were compared. Comparative experiments with the different models show that the proposed algorithm is superior to traditional angle spectrum algorithms in reconstructing cell images based on phase information. Furthermore, experiments have shown that images reconstructed using the improved algorithm can resolve high signal-to-noise ratio information. This algorithmic improvement provides new applications for cellular detection in clinical diagnostics and is more suitable for cell phase reconstruction in practical applications.

Single-shot quantitative differential phase contrast imaging combined with programmable polarization multiplexing illumination.

We propose a single-shot quantitative differential phase contrast method with polarization multiplexing illumination. In the illumination module of our system, a programmable LED array is divided into four quadrants and covered with polarizing films of four different polarization angles. We use a polarization camera with polarizers before the pixels in the imaging module. By matching the polarization angle between the polarizing films over the custom LED array and the polarizers in the camera, two sets of asymmetric illumination acquisition images can be calculated from a single-shot acquisition image. Combined with the phase transfer function, we can calculate the quantitative phase of the sample. We present the design, implementation, and experimental image data demonstrating the ability of our method to obtain quantitative phase images of a phase resolution target, as well as Hela cells.

Single-shot quantitative phase contrast imaging based on deep learning.

Quantitative differential phase-contrast (DPC) imaging is one of the commonly used methods for phase retrieval. However, quantitative DPC imaging requires several pairwise intensity measurements, which makes it difficult to monitor living cells in real-time. In this study, we present a single-shot quantitative DPC imaging method based on the combination of deep learning (DL) and color-encoded illumination. Our goal is to train a model that can generate an isotropic quantitative phase image (i.e., target) directly from a single-shot intensity measurement (i.e., input). The target phase image was reconstructed using a linear-gradient pupil with two-axis measurements, and the model input was the measured color intensities obtained from a radially asymmetric color-encoded illumination pattern. The DL-based model was trained, validated, and tested using thirteen different cell lines. The total number of training, validation, and testing images was 264 (10 cells), 10 (1 cell), and 40 (2 cells), respectively. Our results show that the DL-based phase images are visually similar to the ground-truth phase images and have a high structural similarity index (>0.98). Moreover, the phase difference between the ground-truth and DL-based phase images was smaller than 13%. Our study shows the feasibility of using DL to generate quantitative phase imaging from a single-shot intensity measurement.

Quantitative differential phase contrast imaging in an LED array microscope: errata.

This errata corrects typos found in the derived transfer functions in [Opt. Express23, 11394 (2015)10.1364/OE.23.011394].

Single-shot color-coded LED microscopy for quantitative differential phase contrast imaging

Common quantitative differential phase contrast microscopy imaging usually uses multistep circular or annular illuminations and color multiplexed illuminations, bringing about low capture efficiency and poor phase reconstruction, respectively. We present a new single-shot quantitative differential phase contrast microscopy imaging method, which uses R/B-G annular LED multiplexed illumination to achieve better phase reconstruction by capturing 1 colored image. By deriving the phase transfer function of the illumination and simulation verification, the superiority of it is confirmed. Through phase reconstruction simulations for circular step and gradual samples, we verify that the R/B-G annular multiplexed illumination has better axial phase reconstruction accuracy and weaker lateral anisotropy than other color multiplexed illuminations. As a demonstration, quantitative phase imaging is performed on Quantitative Phase Target, validating the axial accuracy, lateral resolution, and lateral anisotropy of our method. Imaging and phase reconstruction were also performed on mouse kidney tissue slice and motional silica microspheres to visually express the utility and practicability of this method. Our research provides a better color multiplexed avenue for the real-time quantitative phase imaging in dispersion-free, weak-phase samples, especially living biological tissues.

Quantitative phase contrast imaging with a nonlocal angle-selective metasurface

Phase contrast microscopy has played a central role in the development of modern biology, geology, and nanotechnology. It can visualize the structure of translucent objects that remains hidden in regular optical microscopes. The optical layout of a phase contrast microscope is based on a 4 f image processing setup and has essentially remained unchanged since its invention by Zernike in the early 1930s. Here, we propose a conceptually new approach to phase contrast imaging that harnesses the non-local optical response of a guided-mode-resonator metasurface. We highlight its benefits and demonstrate the imaging of various phase objects, including biological cells, polymeric nanostructures, and transparent metasurfaces. Our results showcase that the addition of this non-local metasurface to a conventional microscope enables quantitative phase contrast imaging with a 0.02π phase accuracy. At a high level, this work adds to the growing body of research aimed at the use of metasurfaces for analog optical computing.

Quantitative Phase and Absorption Contrast Imaging

We present an algorithm for coherent diffractive imaging with phaseless measurements. It treats the forward model as a combination of coherent and incoherent waves. The algorithm reconstructs absorption and phase contrast that quantifies the attenuation and the refraction of the waves propagating through an object. It requires coherent or partially coherent illuminations, and several detectors to record the intensity of the distorted wave that passes through the object under inspection. The diversity of illuminations, obtained by putting masks between the source and the object, provides enough information for imaging. The computational cost of our algorithm is linear in the number of pixels of the image. Therefore, it is efficient for high-resolution imaging. Our algorithm guarantees exact recovery if the image is sparse for a given basis. Numerical experiments in the setting of phaseless diffraction imaging of sparse objects validate the efficiency and the precision of the suggested algorithm.

Factors limiting quantitative phase retrieval in atomic-resolution differential phase contrast scanning transmission electron microscopy using a segmented detector

Quantitative differential phase contrast imaging of materials in atomic-resolution scanning transmission electron microscopy using segmented detectors is limited by various factors, including coherent and incoherent aberrations, detector positioning and uniformity, and scan-distortion. By comparing experimental case studies of monolayer and few-layer graphene with image simulations, we explore which parameters require the most precise characterisation for reliable and quantitative interpretation of the reconstructed phases. Coherent and incoherent lens aberrations are found to have the most significant impact. For images over a large field of view, the impact of noise and non-periodic boundary conditions are appreciable, but in this case study have less of an impact than artefacts introduced by beam deflections coupling to beam scanning (imperfect tilt-shift purity).

Patch-Based U-Net Model for Isotropic Quantitative Differential Phase Contrast Imaging.

Quantitative differential phase-contrast (qDPC) imaging is a label-free phase retrieval method for weak phase objects using asymmetric illumination. However, qDPC imaging with fewer intensity measurements leads to anisotropic phase distribution in reconstructed images. In order to obtain isotropic phase transfer function, multiple measurements are required; thus, it is a time-consuming process. Here, we propose the feasibility of using deep learning (DL) method for isotropic qDPC microscopy from the least number of measurements. We utilize a commonly used convolutional neural network namely U-net architecture, trained to generate 12-axis isotropic reconstructed cell images (i.e. output) from 1-axis anisotropic cell images (i.e. input). To further extend the number of images for training, the U-net model is trained with a patch-wise approach. In this work, seven different types of living cell images were used for training, validation, and testing datasets. The results obtained from testing datasets show that our proposed DL-based method generates 1-axis qDPC images of similar accuracy to 12-axis measurements. The quantitative phase value in the region of interest is recovered from 66% up to 97%, compared to ground-truth values, providing solid evidence for improved phase uniformity, as well as retrieved missing spatial frequencies in 1-axis reconstructed images. In addition, results from our model are compared with paired and unpaired CycleGANs. Higher PSNR and SSIM values show the advantage of using the U-net model for isotropic qDPC microscopy. The proposed DL-based method may help in performing high-resolution quantitative studies for cell biology.

Dual-energy phase retrieval algorithm for inline phase sensitive x-ray imaging system.

Phase retrieval is vital for quantitative x-ray phase contrast imaging. This work presents an iterative method to simultaneously retrieve the x-ray absorption and phase images from a single x-ray exposure. The proposed approach uses the photon-counting detectors' energy-resolving capability in providing multiple spectrally resolved phase contrast images from a single x-ray exposure. The retrieval method is derived, presented, and experimentally tested with a multi-material phantom in an inline phase contrast imaging setup. By separating the contributions of photoelectric absorption and Compton scattering to the attenuation, the authors divide the phase contrast image into two portions, the attenuation map arises from photoelectric absorption and a pseudo phase contrast image generated by electron density. This way one can apply the Phase Attenuation Dualiby (PAD) algorithm and Fresnel propagation for the iteration. The retrieval results from the experimental images show that this iterative method is fast, accurate, robust against noise, and thus yields noticeable enhancement in contrast to noise ratios.

3D monitoring of the surface slippage effect on micro-particle sedimentation by digital holographic microscopy

In several phenomena in biology and industry, it is required to understand the comprehensive behavior of sedimenting micro-particles in fluids. Here, we use the numerical refocusing feature of digital holographic microscopy (DHM) to investigate the slippage effect on micro-particle sedimentation near a flat wall. DHM provides quantitative phase contrast and three-dimensional (3D) imaging in arbitrary time scales, which suggests it as an elegant approach to investigate various phenomena, including dynamic behavior of colloids. 3D information is obtained by post-processing of the recorded digital holograms. Through analysis of 3D trajectories and velocities of multiple sedimenting micro-particles, we show that proximity to flat walls of higher slip lengths causes faster sedimentation. The effect depends on the ratio of the particle size to (1) the slip length and (2) its distance to the wall. We corroborate our experimental findings by a theoretical model which considers both the proximity and the particle interaction to a wall of different hydrophobicity in the hydrodynamic forces.

Spatial coherence control and analysis via micromirror-based mixed-state ptychography

Flexible and fast control of the phase and amplitude of coherent light, enabled by digital micromirror devices (DMDs) and spatial light modulators, has been a driving force for recent advances in optical tweezers, nonlinear microscopy, and wavefront shaping. In contrast, engineering spatially partially coherent light remains widely elusive due to the lack of tools enabling a joint analysis and control sequence. Here, we report an approach to coherence engineering that combines a quasi-monochromatic, thermal source and a DMD together with a ptychographic scanning microscope. The reported method opens up new routes to low-cost coherence control, with applications in micromanipulation, nanophotonics, and quantitative phase contrast imaging.

Transmission Structured Illumination Microscopy for Quantitative Phase and Scattering Imaging

In this paper, we demonstrate a digital micromirror device (DMD) based optical microscopic apparatus for quantitative differential phase contrast (qDIC) imaging, coherent structured illumination microscopy (SIM), and dual-modality (scattering/fluorescent) imaging. For both the qDIC imaging and the coherent SIM, two sets of fringe patterns with orthogonal orientations and five phase-shifts for each orientation, are generated by a DMD and projected on a sample. A CCD camera records the generated images in a defocusing manner for qDIC and an in-focus manner for coherent SIM. Both quantitative phase images and super-resolved scattering/fluorescence images can be reconstructed from the recorded intensity images. Moreover, fluorescent imaging modality is integrated, providing specific biochemical structures of the sample once using fluorescent labeling.

Sample phase gradient and fringe phase shift in triple phase grating X-ray interferometry.

Triple phase grating X-ray interferometry is a promising new technique of grating based X-ray differential phase contrast imaging. Accurate retrieval of sample phase gradients from measured interference fringe shifts is a key task in X-ray interferometry. To fulfill this task in triple phase grating X-ray interferometry with monochromatic X-ray sources, the authors derived exact formulas relating sample phase gradient to fringe phase shift. These formulas not only provide a design optimization tool for triple phase grating interferometry, but also lay a foundation for quantitative phase contrast imaging.

Quantitative phase contrast imaging of a shock-wave with a laser-plasma based X-ray source

X-ray phase contrast imaging (XPCI) is more sensitive to density variations than X-ray absorption radiography, which is a crucial advantage when imaging weakly-absorbing, low-Z materials, or steep density gradients in matter under extreme conditions. Here, we describe the application of a polychromatic X-ray laser-plasma source (duration ~0.5 ps, photon energy >1 keV) to the study of a laser-driven shock travelling in plastic material. The XPCI technique allows for a clear identification of the shock front as well as of small-scale features present during the interaction. Quantitative analysis of the compressed object is achieved using a density map reconstructed from the experimental data.

Quantitative Clinical Cardiac Magnetic Resonance Imaging.

Cardiac magnetic resonance imaging (MRI) represents the established reference standard method for the assessment of cardiac function and non-invasive evaluation of myocardial tissue in a variety of clinical questions, wherein quantification of cardiac parameters gains growing diagnostic and differential-diagnostic importance. This review aims to summarize established and newly emerging quantitative parameters, which are assessed in routine cardiac MRI. Interrelations and interdependencies between metrics are explained, and common factors affecting quantitative results are discussed. The review is based on a PubMed literature research using the search terms "cardiac magnetic resonance" and "quantification", "recommendations", "quantitative evaluation/assessment", "reference method", "reference/normal values", "pitfalls" or "artifacts" published between 2000-2019. Quantitative functional, phase contrast, and perfusion imaging, as well as relaxation time mapping techniques give opportunity for assessment of a large number of quantitative cardiac MRI parameters in clinical routine. Application of these techniques allows for characterization of function, morphology and perfusion of the heart beyond visual analysis of images, either in primary evaluation and comparison to normal values or in patients' follow-up and treatment monitoring. However, with implementation of quantitative parameters in clinical routine, standardization is of particular importance as different acquisition and evaluation strategies and algorithms may substantially influence results, though not always immediately apparent. · Clinical cardiac MRI provides numerous functional and morphological quantitative parameters.. · Quantitative cardiac MRI enables assessment of diffuse and global myocardial alterations.. · Standardized data acquisition/evaluation is the prerequisite for diagnostic use of quantitative cardiac MRI parameters.. · Reiter U, Reiter C, Kräuter C et al. Quantitative Clinical Cardiac Magnetic Resonance Imaging. Fortschr Röntgenstr 2020; 192: 246 - 256.

Multi-wavelength quantitative differential phase contrast imaging by radially asymmetric illumination.

A new approach for achieving isotropic differential phase contrast imaging by applying multi-wavelength asymmetric illumination is demonstrated. Multi-wavelength isotropic differential phase contrast scheme (MW-iDPC) can be implemented using an add-on module in any commercial inverted microscope. Isotropy of intensity transfer function is achieved using three axis measurements. The expression for MW-iDPC imaging is presented, and detailed mathematical analysis is performed for transfer function. By applying color leakage correction, image sensor responses can be calibrated. Asymmetric illumination masks are designed, and simulation studies for intensity of the transfer function are performed. We utilize the MW-iDPC system to reconstruct quantitative phase images of standard microspheres and live breast cancer cells. The optical thickness ofcells can be measured with high accuracy and image acquisition time is reduced significantly.

No-search focus prediction at the single cell level in digital holographic imaging with deep convolutional neural network.

Digital propagation of an off-axis hologram can provide the quantitative phase-contrast image if the exact distance between the sensor plane (such as CCD) and the reconstruction plane is correctly provided. In this paper, we present a deep-learning convolutional neural network with a regression layer as the top layer to estimate the best reconstruction distance. The experimental results obtained using microsphere beads and red blood cells show that the proposed method can accurately predict the propagation distance from a filtered hologram. The result is compared with the conventional automatic focus-evaluation function. Additionally, our approach can be utilized at the single-cell level, which is useful for cell-to-cell depth measurement and cell adherent studies.

Quantitative Phase-Contrast Imaging for Distinction between Different States of Human Breast Cancer Cells

We present label-free phase imaging to differentiate human breast cancer cells for breast cancer diagnosis. Low-coherent quantitative phase microscopy (QPM) is exploited to provide speckle-free interference images of live cells adhered to a plate, from which the phase information of the individual cells can be extracted. The phase measurement performance of QPM is validated by the phase-derived optical thickness of a sub-wavelength featured sample, consistent with the results when using conventional atomic force microscopy (AFM). The phase sensitivity is recorded as ~44 mrad, corresponding to ~2.3 nm depth fluctuation during measurement. Phase-contrast imaging is performed with three breast epithelial cell lines: non-tumorous (MCF-10A), weakly malignant (MCF-7), and highly malignant (MDA-MB-231), which mimic an in vivo environments in development of human breast cancer. Our results indicate that MCF-10A and MDA-MB-231 exhibited significantly lower phase-shifts than those of MCF-7, but no difference in phase was observed between MCF-10A and MDA-MB-231. We expect that the use of the QPM approach would be beneficial for clinicians to quantitatively evaluate the breast cancer states from the unmodified specimen of breast biopsy.