Object-wave reconstruction by carbon film-based Zernike- and Hilbert-phase plate microscopy: a theoretical study not restricted to weak-phase objects

Transmission electron microscopy phase-contrast images taken by amorphous carbon film-based phase plates are affected by the scattering of electrons within the carbon film causing a modification of the image-wave function. Moreover, image artefacts are produced by non-centrosymmetric phase plate designs such as the Hilbert-phase plate. Various methods are presented to correct phase-contrast images with respect to the scattering of electrons and image artefacts induced by phase plates. The proposed techniques are not restricted to weak-phase objects and linear image formation. Phase-contrast images corrected by the presented methods correspond to those taken by an ideal centrosymmetric, matter-free phase plate and are suitable for object-wave reconstruction.     

An autonomous phase-boundary detection technique for colloidal hard sphere suspension experiments

Colloidal suspensions of monodisperse spheres are used as physical models of thermodynamic phase transitions and as precursors to photonic band gap materials. Current techniques for identifying the phase boundaries involve manually identifying the phase transitions, which is very tedious and time-consuming. In addition, current image analysis techniques are not able to distinguish between densely packed phases within conventional microscope images, which are mainly characterized by degrees of randomness or order with similar grayscale value properties. We have developed an intelligent machine vision technique that automatically identifies colloidal phase boundaries. The technique utilizes intelligent image processing algorithms that accurately identify and track phase changes vertically or horizontally for a sequence of colloidal hard sphere suspension images. This technique is readily adaptable to any imaging application wherein regions of interest are distinguished from the background by differing patterns of motion over time.     

基于复合Zernike矩相角估计的图像配准

提出了一种基于复合Zernike矩相角估计的图像配准方法.首先,利用尺度不变检测子Harris-laplace检测图像中的兴趣点作为初始特征点,计算以兴趣点为中心、邻域具有尺度不变性的Zernike矩;提出一种鲁棒的相角估计方法,用于估计两个归一化区域的旋转角度值.然后,利用Zernike矩的幅值和相角信息,通过比较每个兴趣点邻域Zernike矩的相似度提取出初始匹配点.最后,提出一种迭代角度修正算法用于精确估计变换参数,并对输入图像进行几何变换后将两幅图像配准.实验结果表明,该算法可在尺度缩放、任意角度旋转以及噪声等复杂条件下实现图像的高精度配准.当旋转角度误差小于20°时,图像的平均覆盖率达到94.125％,有效降低了误匹配的概率.     

A focusing algorithm for high magnification cell imaging

An algorithm to produce a uniformly focused image in digital acquisition of high magnification light microscopy images is presented. In very high magnification microscopic imaging the specimen surface cannot be considered ideally flat so that capturing a single image frame is usually not sufficient to capture an image that is focused everywhere. An image formation model for light microscopic images is presented, and based on this model an algorithm to construct a uniformly focused image is presented. The algorithm requires that multiple frames of the image at different focal planes be processed to combine their information to obtain an estimated of the desired image which is more completely focused than any of the individual frames. Experimental results show that the proposed algorithm is very effective in approximating the desired image in high magnification microscopic imaging and highly robust comparing to the gradient method.     

Object wave reconstruction by phase-plate transmission electron microscopy

A method is described for the reconstruction of the amplitude and phase of the object exit wave function by phase-plate transmission electron microscopy. The proposed method can be considered as in-line holography and requires three images, taken with different phase shifts between undiffracted and diffracted electrons induced by a suitable phase-shifting device. The proposed method is applicable for arbitrary object exit wave functions and non-linear image formation. Verification of the method is performed for examples of a simulated crystalline object wave function and a wave function acquired with off-axis holography. The impact of noise on the reconstruction of the wave function is investigated.     

一种适用于特征稀少零件的图像精确拼接方法

针对特征稀少零件的图像精确拼接难题,提出了一种基于相位相关法和闭环运动控制的图像精确拼接方法,以充分发挥软硬件的综合优势。该方法以具有足够特征信息的零件为对象,获取成像系统分别沿X轴和Y轴运动时的零件等距序列图像并预处理,再利用相位相关法求解图像配准参数。在闭环运动控制系统的良好重复定位精度支持下,将上述图像配准参数视为系统配准参数,以进行特征稀少零件的图像拼接。典型零件的图像拼接实验表明,该拼接方法用于特征稀少零件的图像拼接具有无像素级拼接错位和拼接速度快等优点。     

Automated Surface Inspection Using Gabor Filters

In this paper we present a machine vision system for the automatic inspection of defects in textured surfaces found in industry. The defects to be inspected are those that appear as local anomalies embedded in a homogeneous texture. The proposed method is based on a Gabor filtering scheme that computes the output response of energy from the convolution of a textured image with a specific Gabor filter. The best parameters of a Gabor filter are selected so that the energy of the homogeneous texture is zero, and any unpredictable defeats will generate significantly large energy values. A simple thresholding scheme then follows to discriminate between homogeneous regions and defective regions in the filtered image. This transforms texture differences into detectable filter output. The experiments on structural textures such as leather and sandpaper have shown the effectiveness of the proposed method.     

The quantitative measurement of magnetic moments from phase images of nanoparticles and nanostructures—I. Fundamentals

An approach that can be used to measure the magnetic moment of a magnetized nanoparticle or nanostructure from an electron-optical phase image is introduced. The measurement scheme is based on integration of the gradient of the measured phase image within a circular boundary that contains the structure of interest. The quantity obtained is found to be directly proportional to the magnetic moment of the particle, with a constant of proportionality that does not depend on the particle's shape or magnetization state. The measurement of magnetic moments from both simulated and experimental phase images is demonstrated, and strategies are presented that can be utilized to overcome sources of error associated with, for example, the presence of neighboring magnetic particles and the perturbation of the holographic reference wave.     

Electron crystallography at atomic resolution: The structure of the odd-chain paraffin n-tritriacontane

The crystal structure of the odd-chain paraffin, n-tritriacontane, nC₃₃H₆₈, is determined directly by using low-dose electron microscope images and electron diffraction intensity data from epitaxially grown microcrystals. Phases of the most intense “polyethylene” reflections are determined from triplet structure-invariant relationships often used in X-ray crystallography. Low-dose electron microscopic images provide phases of the low-angle “lamellar” reflections and these can be used with one-dimensional structure-invariant relationships to determine other phases on the 00? reciprocal row. The phase set is sufficient to calculate an electrostatic potential map which is directly interpretable as a structure image at atomic resolution.     

相位相关配准法及其在射线图像数字减影中的应用

介绍了一种基于相位相关法射线图像配准的方法。该方法利用傅立叶变换的平移特性对发生平移和旋转的两幅图像进行精确的配准。提出了将该方法用在数字减影中图像的配准 ,首先用相位相关配准法对发生平移和旋转的两幅图像进行配准 ,然后 ,对配准好的图像进行数字减影 ,正确地检测出了构件的内部状态。实验证明 ,该方法完全不受人工干预 ,具有较高的精度和较强的抗噪能力 ,匹配速度也有很大提高。同时 ,旋转校正算法简单 ,易于实现 ,能够满足系统的实时性要求     

SLM彩色显微图像的亚像素边缘检测方法

微操作技术的迅速崛起迫切要求显微图像处理技术的发展。在HSI彩色模型的基础上,根据SLM视觉系统的彩色显微图像特点,基于小波变换提出了一种适用于SLM彩色显微图像的边缘检测方法。根据三次样条拟合曲线的一阶导数零点,获取精确的亚像素边缘。试验证明,该方法适用于各放大倍率下的SLM彩色显微图像,能较好地滤除噪声,检测出完整的亚像素边缘。给出了利用该算法处理噪声仿真图像和真实SLM彩色显微图像的试验结果。     

Sequential processing of quantitative phase images for the study of cell behaviour in real‐time digital holographic microscopy

Transmitted light holographic microscopy is particularly used for quantitative phase imaging of transparent microscopic objects such as living cells. The study of the cell is based on extraction of the dynamic data on cell behaviour from the time‐lapse sequence of the phase images. However, the phase images are affected by the phase aberrations that make the analysis particularly difficult. This is because the phase deformation is prone to change during long‐term experiments. Here, we present a novel algorithm for sequential processing of living cells phase images in a time‐lapse sequence. The algorithm compensates for the deformation of a phase image using weighted least‐squares surface fitting. Moreover, it identifies and segments the individual cells in the phase image. All these procedures are performed automatically and applied immediately after obtaining every single phase image. This property of the algorithm is important for real‐time cell quantitative phase imaging and instantaneous control of the course of the experiment by playback of the recorded sequence up to actual time. Such operator's intervention is a forerunner of process automation derived from image analysis. The efficiency of the propounded algorithm is demonstrated on images of rat fibrosarcoma cells using an off‐axis holographic microscope.     

高阶混合正则化图像盲复原方法

提出了一种高阶混合正则化图像盲复原方法,用于实现模糊噪声图像的清晰化盲复原。根据自然图像边缘的稀疏特性,对图像的边缘细节成分进行了全变差(total variation TV)正则化约束,根据自然图像同性质平滑区域内像素值的变化规律,将一种高阶的类Tikhonov正则化约束运用于图像的平滑区域中,提出了一种新的高阶混合正则化模型。最后,提出一种多变量分裂布雷格曼(Multi-variable Split Bregman MSB)最优化迭代策略对提出的模型进行最优化求解。实验结果表明,提出的方法能够很好地保护图像的边缘细节,同时有效地消除图像平滑区域内的阶梯和假边缘瑕疵。与近几年的一些较好的图像盲复原方法相比,本文方法的信噪比增量(increase of the signal to noise ratio ISNR)增加了0.03~2.5 dB。     

Structure of purple membrane from halobacterium halobium: recording,measurement and evaluation of electron micrographs at 3.5 Å resolution

Electron micrographs of the purple membrane have been recorded using liquid nitrogen and liquid helium cooling on three cryoelectron microscopes. The best micrographs show optical diffraction spots, arising from the two-dimensional crystal, out to resolutions of around 6 Å. Large areas of several of these micrographs have been analysed using a procedure which determines the strength of the very weak high resolution Fourier components of the image of the crystal. The procedure consists of reciprocal space filtering followed by real space correlation analysis to characterise image distortions, removal of the distortions by interpolation, and finally extraction of the amplitudes and phases of the Fourier components from the distortion-corrected image of the crystal. These raw image amplitudes and phases are then used, together with previously measured amplitude and phase information, to refine the beam tilt and crystal tilt, phase origin and amount of defocus and astigmatism of the image. The phases can then be corrected for the effects of the contrast transfer function, beam tilt and phase origin. The amplitudes of all the spots which are expected to be strong from their known electron diffraction intensity are observed to be significantly above the background noise level, and the independent phases from different images, and from symmetry-related directions in the same image, show excellent agreement out to a resolution of 3.5 Å. Although only images from untilted or slightly tilted ( < 5°) crystals have been analysed using the procedure described in this paper, a simple additional step enables analysis of images at any tilt angle, providing a complete practical method for high resolution analysis of images of two-dimensional crystalline arrays.     

Reconstruction of images of reflectors using the correlation method at an arbitrary number of reflections of an ultrasonic pulse from the boundary of a test object that consists of regions with different acoustic properties

An algorithm for reconstructing images of reflectors from echo signals that propagate in a test object, which consists of several regions with different acoustic properties, is considered. The ray trajectories are calculated using the method of direct construction of the family of rays that escape from the point where a transmitter is located but not the Fermat variational principle. After the family of rays is constructed, their belonging to a certain acoustic scheme can be analyzed and the approximation of the calculated delays on the spatial mesh of the image reconstruction region (IRR) can be performed. This will allow the reconstruction of both the pulse travel time from the transmitter to any point in the IRR and back to the receiver and the attenuation of the pulse amplitude caused by the divergence, reflection, and refraction effects at the boundaries of the regions that constitute the test object. Numerical and model experiments show the working capacity of the proposed algorithm for reconstructing reflector images.     

面阵CCD在大尺寸形位误差测量中的应用

本文介绍了在半导体激光光纤和位相结合的准直情况下,利用面阵ＣＣＤ探测器测量大尺寸形位误差的原理,并针对二维位相板衍射图像的特点,提出了一种处理此类衍射图样的图像处理新方法－正交投影法,消除了ＣＣＤ表面保护玻璃形式的干涉条纹的影响,对空气扰动也起到了很好的抑制作用,并且使二维图像处理变形为一维计算,大大提高了计算速度。此面阵ＣＣＤ测量系统与双频激光干涉仪测量直线度误差的比对实验结果表明,二者的测量结     

相位法曲面三维全场无接触检测的研究

采用光学函数分析法和傅里叶频谱分析技术，研究了相位法曲面三维全场无接触检测的基本原理和本质特点。在分析条纹类测量图象的幅频待性的基础上，实现了被测曲面调制栅线图象的精确解相和三维信息的求解，从而建立了计算机曲面三维自动检测系统，并应用于汽车覆盖件的检测，获得较好的测量精度。     

Application of convolutional neural network to acquisition of clear images for objects with large vertical size in stereo light microscope vision system

For an object with large vertical size that exceeds the certain depth of a stereo light microscope (SLM), its image will be blurred. To obtain clear images, we proposed an image fusion method based on the convolutional neural network (CNN) for the microscopic image sequence. The CNN was designed to discriminate clear and blurred pixels in the source images according to the neighborhood information. To train the CNN, a training set that contained correctly labeled clear and blurred images was created from an open‐access database. The image sequence to be fused was aligned at first. The trained CNN was then used to measure the activity level of each pixel in the aligned source images. The fused image was obtained by taking the pixels with the highest activity levels in the source image sequence. The performance was evaluated using five microscopic image sequences. Compared with other two fusion methods, the proposed method obtained better performance in terms of both visual quality and objective assessment. It is suitable for fusion of the SLM image sequence.     

Performance evaluation of image segmentation algorithms on microscopic image data

In our paper, we present a performance evaluation of image segmentation algorithms on microscopic image data. In spite of the existence of many algorithms for image data partitioning, there is no universal and ‘the best’ method yet. Moreover, images of microscopic samples can be of various character and quality which can negatively influence the performance of image segmentation algorithms. Thus, the issue of selecting suitable method for a given set of image data is of big interest. We carried out a large number of experiments with a variety of segmentation methods to evaluate the behaviour of individual approaches on the testing set of microscopic images (cross‐section images taken in three different modalities from the field of art restoration). The segmentation results were assessed by several indices used for measuring the output quality of image segmentation algorithms. In the end, the benefit of segmentation combination approach is studied and applicability of achieved results on another representatives of microscopic data category – biological samples – is shown.     

Compensation of inhomogeneous fluorescence signal distribution in 2D images acquired by confocal microscopy

In images acquired by confocal laser scanning microscopy (CLSM), regions corresponding to the same concentration of fluorophores in the specimen should be mapped to the same grayscale levels. However, in practice, due to multiple distortion effects, CLSM images of even homogeneous specimen regions suffer from irregular brightness variations, e.g., darkening of image edges and lightening of the center. The effects are yet more pronounced in images of real biological specimens. A spatially varying grayscale map complicates image postprocessing, e.g., in alignment of overlapping regions of two images and in 3D reconstructions, since measures of similarity usually assume a spatially independent grayscale map. We present a fast correction method based on estimating a spatially variable illumination gain, and multiplying acquired CLSM images by the inverse of the estimated gain. The method does not require any special calibration of reference images since the gain estimate is extracted from the CLSM image being corrected itself. The proposed approach exploits two types of morphological filters: the median filter and the upper Lipschitz cover. The presented correction method, tested on images of both artificial (homogeneous fluorescent layer) and real biological specimens, namely sections of a rat embryo and a rat brain, proved to be very fast and yielded a significant visual improvement. Microsc. Res. Tech., 2011. © 2010 Wiley‐Liss, Inc.