Spectral-domain optical coherence phase microscopy for quantitative phase-contrast imaging

We describe a novel microscopy technique for quantitative phase-contrast imaging of a transparent specimen. The technique is based on depth-resolved phase information provided by common path spectral-domain optical coherence tomography and can measure minute phase variations caused by changes in refractive index and thickness inside the specimen. We demonstrate subnanometer level path-length sensitivity and present images obtained on reflection from a known phase object and human epithelial cheek cells.     

基于白光干涉的自稳定定量相位成像系统

提出和建立了基于白光干涉的自稳定定量相位成像系统.该系统以柯勒照明的白光迈克耳孙干涉计为基础,使用自稳定锁相电路以减少相位噪音,引入相移装置获得定量的相化位图像.实验分别以平面镜、标准测试图和聚苯乙烯小球为样品.获得了清晰的定量相位图像,实现了对样品的实时观测.该装置采用卤素灯照明,达到轴向分辨力5 nm,系统中的柯勒照明结构使其具有高空间相干性.实验结果表明,该装置能有效避免系统中其他光学表面(如系统中的透镜表面)干扰信号的影响.     

Quantitative phase imaging with a scanning transmission x-ray microscope

We obtain quantitative phase reconstructions from differential phase contrast images obtained with a scanning transmission x-ray microscope and 2.5 keV x rays. The theoretical basis of the technique is presented along with measurements and their interpretation.     

Quantitative phase imaging using actively stabilized phase-shifting low-coherence interferometry

We describe a quantitative phase-imaging interferometer in which phase shifting and noise cancellation are performed by an active feedback loop using a reference laser. Depth gating via low-coherence light allows phase measurement from weakly reflecting biological samples. We demonstrate phase images from a test structure and living cells.     

Phase-sensitive acoustic microscopy of polymer thin films

The three-dimensional images obtained by scanning acoustic microscopy with vector contrast (PSAM), contain significant qualitative and quantitative information that is not easily obtainable by other methods. We employ this technique to examine homopolymer and polymer blend thin films. The complex V(z) functions derived from the images, and the results obtained by image processing and meticulous analysis are employed to render the morphology, composition and micro-mechanical properties of the polymer films. In addition, ways by which the information inherent in the phase images can be extracted are examined. This is highly desirable, as the phase images contain very useful additional information.     

Hilbert phase microscopy for investigating fast dynamics in transparent systems

We introduce Hilbert phase microscopy (HPM) as a novel optical technique for measuring high transverse resolution quantitative phase images associated with optically transparent objects. Because of its single-shot nature, HPM is suitable for investigating rapid phenomena that take place in transparent structures such as biological cells. The potential of this technique for studying biological systems is demonstrated with measurements of red blood cells, and its ability to quantify dynamic processes on a millisecond scale is exemplified with measurements of evaporating micrometer-sized water droplets.     

Deep-learning-based prediction of living cells mitosis via quantitative phase microscopy

We present a deep learning approach for living cells mitosis classification based on label-free quantitative phase imaging with transport of intensity equation methods. In the approach, we applied a pretrained deep convolutional neural network using transfer learning for binary classification of mitosis and non-mitosis. As a validation, we demonstrated the performances of the network trained by phase images and intensity images, respectively. The convolutional neural network trained by phase images achieved an average accuracy of 98.9% on the validation data, which outperforms the average accuracy 89.6% obtained by the network trained by intensity images. We believe that the quantitative phase microscopy in combination with deep learning enables researchers to predict the mitotic status of living cells noninvasively and efficiently.     

Tissue refractometry using Hilbert phase microscopy

We present, for the first time to our knowledge, quantitative phase images associated with unstained 5 mum thick tissue slices of mouse brain, spleen, and liver. The refractive properties of the tissue are retrieved in terms of the average refractive index and its spatial variation. We find that the average refractive index varies significantly with tissue type, such that the brain is characterized by the lowest value and the liver by the highest. The spatial power spectra of the phase images reveal power law behavior with different exponents for each tissue type. This approach opens a new possibility for stain-free characterization of tissues, where the diagnostic power is provided by the intrinsic refractive properties of the biological structure. We present results obtained for liver tissue affected by a lysosomal storage disease and show that our technique can quantify structural changes during this disease development.     

Jones phase microscopy of transparent and anisotropic samples

We developed an interferometric microscopy technique, referred to as Jones phase microscopy, capable of extracting the spatially resolved Jones polarization matrix associated with transparent and anisotropic samples. This is a generalization of quantitative phase imaging, which is recovered from one diagonal element of the measured matrix. The principle of the technique is demonstrated with measurements of a liquid crystal spatial light modulator and the potential for live cell imaging with experiments on live neurons in culture.     

Full-field quantitative phase imaging by white-light interferometry with active phase stabilization and its application to biological samples

We report a Koehler-illumination-based full-field, actively stabilized, low-coherence phase-shifting interferometer, which is built on a white-light Michelson interferometer. By using a phase-stepping technique we can obtain full-field phase images of the sample. An actively stabilized phase-lock circuit is employed in the system to reduce phase noise. An application to human epithelial cells (HeLa cells) is achieved in our experiment. The advancement of this technique rests in its ability to take images of unstained biological samples quantitatively and on a nanometer scale.     

Confocal diffraction phase microscopy of live cells

We present a new quantitative phase microscopy technique, confocal diffraction phase microscopy, which provides quantitative phase measurements from localized sites on a sample with high sensitivity. The technique combines common-path interferometry with confocal microscopy in a transmission geometry. The capability of the technique for static imaging is demonstrated by imaging polystyrene microspheres and live HT29 cells, while dynamic imaging is demonstrated by quantifying the nanometer scale fluctuations of red blood cell membranes.     

Curvature measurement using three-aperture digital shearography and fast Fourier transform

Curvature measurement using a three-aperture digital shearography (DS) system is reported in this paper. The outer apertures are covered with wedge plates for introducing shear. Four images by sequentially blocking the outer apertures are used for quantitative measurement. Fourier transform technique is used to determine two sheared slope phase maps from two images at a time representing initial and deformed states. Subtraction of these two-phase maps yields the curvature phase map. Experimental results are presented for a circular diaphragm clamped along the edges and loaded at the center.     

Extended depth of focus in tomographic phase microscopy using a propagation algorithm

Tomographic phase microscopy is a laser interferometry technique in which a 3D refractive index map of a biological sample is constructed from quantitative phase images collected at a set of illumination angles. Although the resulting tomographic images provide valuable information, their resolution declines at axial distances beyond about 1 microm from the focal plane. We describe an improved 3D reconstruction algorithm in which the field at the focal plane is numerically propagated to depths throughout the sample. Diffraction is thus incorporated, extending the depth of focus to more than 10 mum. Tomograms with improved focal depth are demonstrated for single HT29 cells.     

Full-field swept-source phase microscopy

We present a full-field phase microscopy technique for quantitative nanoscale surface profiling of samples in reflection. This technique utilizes swept-source optical coherence tomography in a full-field common path interferometer for phase-stable cross-sectional acquisition without scanning. Subwavelength variations in surface sample features are measured without interference from spurious reflections by processing the interferometric phase at a selected depth plane, providing a 1.3 nm stability for high signal-to-noise ratio surface features. Nanoscale imaging was demonstrated by measuring the location of receptor sites on a DNA assay biochip and the surface topography of erythrocytes in a blood smear.     

Quantitative HRTEM investigation of nanoplatelets

High resolution transmission electron microscopy experiments were performed to investigate nanoplatelets induced by ion implantation into a germanium wafer. Atomistic models were used for image simulation in order to get quantitative information from the experimental images. The geometrical phase shift analysis technique was also employed to measure the strain field induced by such defects. We discuss the limits and artefacts imposed by each approach and show how these approaches can be combined to study the atomic structure of such defects and the strain field they induce.     

The study and application of a new filtering method on electronic speckle pattern interferometric fringe

It is important for the study of digital image processing to remove noises and enhance images because of various noises in images. The article is concerned with the recursive algorithm-filtering technique, which is capable of effectively removing noise and rapidly leading to a continuous gray-level distribution of a random image. By applying the method on speckle images of defects of rubber material, it is shown that this technique is the most effective and speedy method for the filtering of ESPI fringe patterns. Comparing the images before and after filtering, we can see the advantages of the novel technique clearly. It can increase the accuracy of measurement both in phase measurement and in quantitative evaluations of defects.     

Enhanced quantitative X-ray phase-contrast images using Foucault differential filters

Enhanced quantitative X-ray phase-contrast(QXPC) imaging is implemented with a Foucault knife-edge array filter(FKAF), which is a real differential spatial filter. The intensities of Foucault differential filtering(FDF) are acquired according to the linear translation of the FKAF along the axes. The FDF using the FKAF scheme for obtaining the QXPC images is demonstrated by a stereoscopic rendering of the quantitative phase images of the tail fin of an anchovy containing soft and hard components in specimen. FDF is a noninterferometric quantitative phase-imaging method that depicts quantitative phase images and renders stereoscopic images.     

A new method of three-dimensional measurement by differential interference contrast microscope

Based on weak phase approximation and the partial coherence theory, we analyze the image characteristics of a phase object using a microscope. We show that the image of the phase object is formed by the interplay between the phase distribution and the defocus.Using this theory, we also show the image characteristics of a differential interference contrast (DIC) microscope.We develop a method for extracting the phase component from the DIC image using two images with different retardation to reconstruct the phase distribution of the object. We call our new microscope a “retardation-modulated DIC (RM-DIC) microscope”. We describe the RM-DIC microscope and confirm our method using grating samples with depths of 20 and 50 nm.To measure the three-dimensional (3D) figures of the microstructures on the object using a DIC microscope we need to extract the phase component from the DIC image and to deconvolute the phase component by means of the modulation transfer function (MTF) of the DIC microscope.We conclude that our RM-DIC microscope can take quantitative measurements of the phase distribution, making it a very useful tool for 3D measurement of an object’s microstructures.     

Adaptive quadrature filters for multiple phase-stepping images

We present a new technique for the recovery of local phase from multiple phase-stepping fringe images that uses adaptive quadrature filters constructed by use of Bayesian estimation theory and complex-valued Markov random fields as prior models. It is shown that with this technique it is possible to perform accurate phase reconstructions even for extremely noisy fringe images and that the performance of this technique is nearly independent of the particular noise model, as long as the noise spectrum is wideband.     

Zernike phase contrast in scanning microscopy with X-rays

Scanning X-ray microscopy focuses radiation to a small spot and probes the sample by raster scanning. It allows information to be obtained from secondary signals such as X-ray fluorescence, which yields an elemental mapping of the sample not available in full-field imaging. The analysis and interpretation from these secondary signals can be considerably enhanced if these data are coupled with structural information from transmission imaging. However, absorption often is negligible and phase contrast has not been easily available. Originally introduced with visible light, Zernike phase contrast(1) is a well-established technique in full-field X-ray microscopes for visualization of weakly absorbing samples(2-7). On the basis of reciprocity, we demonstrate the implementation of Zernike phase contrast in scanning X-ray microscopy, revealing structural detail simultaneously with hard-X-ray trace-element measurements. The method is straightforward to implement without significant influence on the resolution of the fluorescence images and delivers complementary information. We show images of biological specimens that clearly demonstrate the advantage of correlating morphology with elemental information.