Digital image processing of crystalline specimens examined by electron microscopy

Crystalline specimens imaged in the electron microscope are analysed using digital processing. Some principles of structural analysis using the method of Fourier decomposition are discussed. Complementary techniques, such as enhancement by gradient and Laplacian operators, have been found useful in analysing electron micrographs. The application of these techniques to some problems in Materials Science and Biology are reviewed. By selecting and phase-correcting spots in the computed diffraction pattern, it was possible to localize atoms, molecules, and their defective arrangement in evaporated gold, sputter-deposited tungsten films, and single crystals of cadmium selenide. Digital processing based on the theory of helical diffraction was used to explore the three-dimensional arrangement of molecules in cellular components of alveolar soft part sarcoma, Hirano bodies, and neurofibrillar tangles in the human brain.     

数字图像中傅里叶变换的研究分析与仿真

为探索新的数字图像处理方法,研究了数字图像的频谱,特别研究了幅值谱对图像的影响。从高频和低频两个角度,分别研究了频谱。以一幅图像为例,通过Mat Lab进行仿真和分析。尤其是对幅值谱的理论研究,为实现新的数字图像处理方法奠定了基础。     

周期信号离散傅里叶变换的误差分析及其应用

实际的系统都是不满足或不完全满足取样定理条件的,频率混迭是不可避免的。因此ＤＦＴ（ＦＦＴ）给出的频谱是有误差的,本文给出了表达式及估算方法。     

Determining image distortion and PBS (point of best symmetry) in digital images using straight line matrices

It is impossible to take accurate measurements in photogrammetry without first removing the distortion in images. This paper presents a methodology for correcting radial and tangential distortion and for determining the PBS (Point of Best Symmetry) without knowledge of the interior orientation parameters (IOPs). An analytical plumb-line calibration method is used, measuring only the coordinates of points on straight lines, regardless of the position and direction of these lines within the image. Points belonging to multiple lines can also be used since the effects on their X and Y coordinates are calculated independently. The results obtained on an image of a common scene, taken with a handheld non-metric camera, show a high degree of accuracy even with a minimum number of observables. And its application on a calibrated grid for engineering purposes with a semi-metric camera, results optimal even using a single image.     

Lateral performance evaluation of laser micromachining by high precision optical metrology and image analysis

Today several techniques are available for micro-manufacturing. Yet, it is difficult to assess the precision and lateral X,Y accuracy of these techniques. The available accuracy information is usually based on specifications given by machine suppliers. This information is based on in-house laboratory tests performed by dedicated machine operators and within an adapted environment. In practice, the accuracy is likely to vary due to environmental conditions, materials and operator skills. In order to check the specifications in realistic environments the EUMINAfab infrastructure consortium initiated a set of independent high precision onsite verification tests on different laser micromachining installations. In addition to providing performance verification, it gave the participating partners real capability information of their equipment and possibilities to improve machining performance to a higher level. In this study a comprehensive verification test was designed and carried out by using a high precision metrology method for 2D measurements based on subpixel resolution image analysis. This methodology improved our knowledge of the capabilities of three laser micromachining installations, and showed that specifications at single micron levels are hard to obtain.     

The distribution of caveolin-3 immunofluorescence in skeletal muscle fibre membrane defined by dual channel confocal laser scanning microscopy, fast Fourier transform and image modelling

Membrane domains rich in caveolin‐3 overlie sarcomeric actin in skeletal muscle. The membrane exhibits a regular array of caveolin‐3 immunofluorescence using confocal laser scanning microscopy (CLSM). Fourier analysis of tissue imaged by CLSM accurately defines a repeating intensity with a long‐axis spacing of 1.48 µm confirmed by measurement of direct images. Reverse fast Fourier transform (FFT) and image‐modelling allow reconstruction of the pattern. Mathematical modelling has allowed replication of several features of the FFT, including the second order maxima that confirm the relatively high information content of the original images. Measurements of membrane‐pattern primary long‐axis spacings are consistent with our measurements of the I‐band sarcomere repeat in similarly prepared specimens labelled with fluorescent phalloidin or imaged using differential interference contrast microscopy. Dual‐channel CLSM analysis of the sarcomeric banding pattern of actin and the repeating pattern of muscle fibre membrane caveolin showed that caveolae overlie the I‐band. The anti‐caveolin immunofluorescence is deficient over the Z‐disc and maximal toward each of the I‐band extremities. A mechanism of membrane shape change in which membrane–lipid molecules are interposed between more stable anchored rafts associated with caveolae can be envisaged. Thus, increasing girth and reducing length of the sarcolemma in rapid contraction may be explained.     

Condition monitoring of first mode of metal transfer in friction stir welding by image processing techniques

This paper discusses a method for online condition monitoring of the friction stir welding (FSW) process using image processing techniques. Many FSW experiments are carried out at different process parameters and images of the first mode of the weld zone are captured. They are subsequently digitally processed and analysed using MATLAB to study the variations in the quality of weld, subjected to various conditions such as pin failure, and pin depth. This facilitates development of a methodology for online condition monitoring.     

基于UHPLC-LTQ/Orbitrap MS法的苦荬菜内酯Z和11,13α-二氢苦荬菜内酯Z的大鼠肝微粒体体外代谢产物鉴定

苦荬菜内酯Z和11,13α-二氢苦荬菜内酯Z是苦碟子(Ixeris sonchifolia(Bunge)Hance)中2种重要的倍半萜内酯类成分。为研究它们在大鼠肝微粒体中的代谢产物,采用Waters Acquity UPLC BEH C18色谱柱(2.1mm×50mm×1.7μm)分离,以0.1%甲酸水(A)-乙腈(B)为流动相进行梯度洗脱,采用高效液相色谱-线性离子阱静电轨道场质谱(UHPLC-LTQ/Orbitrap MS)技术,在ESI负离子模式下分析样品。筛选出18个化合物(包括苦荬菜内酯Z和11,13α-二氢苦荬菜内酯Z,及其16个代谢产物),并对其中15个代谢产物进行结构鉴定。结果表明,苦荬菜内酯Z和11,13α-二氢苦荬菜内酯Z经大鼠肝微粒体主要发生了羟基化、加氢和脱氢的Ⅰ相代谢反应。该结果初步阐明了苦荬菜内酯Z和11,13α-二氢苦荬菜内酯Z在大鼠肝微粒体中的代谢规律,可为新药、毒理学研究提供支持。     

Detection of small displacement of atoms in crystals by atom resolution electron microscopy

Four kinds of works on the detection of displacement of atoms in crystals are shown. The irregular small displacement of atoms has been detected, with an accuracy of about 0.1 Å around dislocations and stacking faults in Au crystals as shown by their electron microscope images. The displacement of the atomic images is recorded by aberration-free focus (AFF). Even when the periodic displacement of atoms in SiC and TaS₂ crystals is around 0.1% of the lattice constant, this displacement has been revealed as the weak-contrast anomaly in the images. Using in situ observations by a TV system attached to an electron microscope, the rapid movements of atoms that have taken place within 1/30 sec have been recorded. Using the technique of successive subtraction of the images by TV system and image sigma, only the images of moving atoms in Au crystal have been recorded each 1/10 sec.     

Analysis of two-phase flow regimes and parameter distribution in 5 x 5 rod bundle using image processing techniques

With the recent developments in image processing and analysis, this paper presents bubble characteristics distribution in adiabatic air-water two-phase flow through a 5 × 5 rod bundle. The experiment covered water superficial velocities (J_l = 0.012 m/s – 0.421 m/s) and air superficial velocities (J_g = 0.042 m/s – 0.987 m/s) in which three distinct flow regimes were identified. The flow regime map was compared with existing flow regime transition criteria for vertical rod bundles. Distinct features from the two-phase flow images were extracted to train a classifier model to distinguish between regimes from a separate experiment. The model distinguishes between the bubbly flow regime and others accurately. The void fraction and velocity distributions were also extracted from the R–CCN masked images. Bubble-induced turbulence that was dominant in the subchannel at (J_l = 0.28 m/s) shifted to the outer subchannels and gaps when the flow rate increased (J_l = 0.42 m/s). These methods over-predicted the void faction around the surfaces of the inner rods.     

Quantitative characterization of chitosan in the skin by Fourier‐transform infrared spectroscopic imaging and ninhydrin assay: application in transdermal sciences

The chitosan has been used as the primary excipient in transdermal particulate dosage form design. Its distribution pattern across the epidermis and dermis is not easily accessible through chemical assay and limited to radiolabelled molecules via quantitative autoradiography. This study explored Fourier‐transform infrared spectroscopy imaging technique with built‐in microscope as the means to examine chitosan molecular distribution over epidermis and dermis with the aid of histology operation. Fourier‐transform infrared spectroscopy skin imaging was conducted using chitosan of varying molecular weights, deacetylation degrees, particle sizes and zeta potentials, obtained via microwave ligation of polymer chains at solution state. Both skin permeation and retention characteristics of chitosan increased with the use of smaller chitosan molecules with reduced acetyl content and size, and increased positive charge density. The ratio of epidermal to dermal chitosan content decreased with the use of these chitosan molecules as their accumulation in dermis (3.90% to 18.22%) was raised to a greater extent than epidermis (0.62% to 1.92%). A larger dermal chitosan accumulation nonetheless did not promote the transdermal polymer passage more than the epidermal chitosan. A small increase in epidermal chitosan content apparently could fluidize the stratum corneum and was more essential to dictate molecular permeation into dermis and systemic circulation. The histology technique aided Fourier‐transform infrared spectroscopy imaging approach introduces a new dimension to the mechanistic aspect of chitosan in transdermal delivery.     

Potential of functional image processing technique for the measurements of contact area and contact pressure of a radial ply tire in a soil bin testing facility

The exploration principally deals with a promising applicability of image processing method for contact area determination. A digital camera mounted on a single-wheel tester was utilized to take images of contact area at four levels of tire inflation pressure (i.e. 70, 100, 140, and 175 kPa) and six levels of wheel load (i.e. 0.75, 1.75, 2.75, 3.75, 4.75, and 5.75 kN). Contact pressure measurements, subsequently, were performed utilizing seven embedded load cells incorporated in an instrumented housing in longitudinal and perpendicular direction of traversing. It was observed that contact area is a polynomial (with order of two) function of wheel load while there was a reverse linear relation between contact area and inflation pressure. Furthermore, contact pressure has linear relation with vertical load and inflation pressure. Models for contact area and contact pressure were developed by multiple regression analysis while a classical predicting model was assessed using contact area results of image processing technique. The models indicated acceptable coefficient of determinations amounts of 0.974 and 0.953 for contact area and contact pressure models, respectively. However, applied image processing method denoted R²-value of 0.988 for contact pressure equation.     

Automatic high-precision measurements of the location and width of Kikuchi bands in electron backscatter diffraction patterns

The demands for reliability and precision of crystal orientation data obtained through automatic analysis of electron backscattering patterns (EBSPs) in the SEM result in similar demands on the quality of the band position data which is provided by an image analysis procedure. This paper describes a new image processing procedure which is capable of providing accurate measurements of the location and width of typically 10–15 bands in digitized EBSPs of average quality. The new procedure is based on the Hough transform (HT) for line detection, and employs a special backmapping technique for generating two simplified HTs which separately focus on bright and dark lines in the images. A coordinated search for peaks in the two HTs leads to precise estimates of both the position and the width of bands in the patterns. A visual evaluation of the data produced by the new procedure shows that it performs significantly better than the conventional procedure with regard to both reliability and precision. Additionally, the measured band width data are fairly precise and can be used for obtaining a more robust and reliable indexing of the bands. Finally, the computational costs of the new procedure are smaller than for the conventional procedure.     

Evaluation of lanthanide tracer methods in the study of mammalian pulmonary parenchyma and cardiac muscle by electron energy-loss spectroscopy

Lanthanum (La) has widely been used as a tracer to study the integrity of plasma membranes. With conventional transmission electron microscopy (cTEM), the absence of electron scattering deposits from the cytoplasm has generally been assumed to reflect an intact cell membrane. However, the application of electron spectroscopic imaging (ESI) and electron energy-loss spectroscopy (EELS) reveals that electron scattering deposits may be present which do not contain La. However, La could be detected in regions of pulmonary parenchyma and cardiac muscle that were devoid of electron scattering deposits. Therefore, to exclude misinterpretations based on cTEM the application of microanalytical techniques is strongly recommended for the study of the integrity of plasma membranes by means of La tracers. In addition, ESI and EELS are shown to distinguish between different tracers in simultaneous applications of La and terbium (Tb) which were used at the different faces of the pulmonary air-blood barrier. The analysis of the distribution of both tracers which form electron scattering deposits, indistinguishable by cTEM, may help us to understand the different functional significances of cellular alterations of both cellular borders of the barrier. As was shown for La, however, strictly controlled conditions are mandatory during the fixation procedure because an increase in the incubation time to more than 1 h in samples of pulmonary parenchyma may result in the occurrence of La deposits within the cytoplasm. In the absence of electron scattering deposits, the presence of La in glycogen granules and ribosome-containing areas of various types of alveolar septal cells even after 15 min incubation indicates that the absence of deposits does not necessarily correspond to the absence of the tracer.