Combination of thresholding and fitting methods for measuring nanoparticle sizes and size distributions in (S)TEM

The practical need for a simple and reliable tool for routine size analysis of nanoparticles with diameters down to a few nm embedded in a polymer matrix motivated the development of a new approach. The idea underlying the method proposed in this work is to combine intensity thresholding and contrast fitting procedures in the same software for particle recognition and measurements of sizes and size distributions of nanoparticles in transmission and scanning transmission electron microscopy images. Particle recognition in images is performed in an interactive process of manual setting the numerical threshold level after image preprocessing. We show that fitting the calculated gray level distribution to the real images is able to provide a maximum accuracy in measurements of the particle diameters in contrast to thresholding approaches. The fitting procedure is applied in the vicinity of nanoparticle images with the mass‐thickness, diffraction, and chemical contrast. The grayscale function associated to the nanoparticle thickness is described using polynomial with degree ? 2 and undetermined coefficients. The program for particle detection and size measurement— An alyzer of Na noparticles ( AnNa )—has been written and is described here. It was successfully tested on systems containing Ag nanoparticles grown and stabilized in aqueous solutions of different polymers for biomedical use and is available from the authors.     

视觉测量中圆形标志两种提取方法的比较

针对靶场视觉测量相机标定中圆形标志在复杂背景图像上的提取和定位问题,研究了两种不同的圆形标志提取方法,一种是基于感兴趣区域的交互式提取方法,另一种是基于图像灰度和几何形态学的提取方法。对两种方法的圆形标志提取过程进行了研究,并分别采用两种方法对视觉测量图像中的圆形标志进行提取实验,对两种方法的边缘提取效果和定位结果进行了对比,分析了两种方法的优势和不足。圆形标志的提取结果表明,第一种方法能够平滑地提取出圆形标志的边缘,但受圆形标志的成像情况影响较大,第二种方法边缘提取效果受圆形标志的成像情况影响较小,但提取出的边缘为锯齿形,部分圆形标志的像素点被误判为背景,与圆形标志的实际成像有一定误差。     

A comparative evaluation of RPD and ferrographic diagnostic methods for turbine engine lubricant samples

A rotary particle depositor (RPD) was evaluated and compared with analytical (AF) and direct reading (DR) ferrographs to ascertain if the RPD is better suited for analysis of wear debris in turbine engine lubricant samples than the AF and DR. Sieved iron particles were added to synthetic turbine engine lubricants at various concentrations and were analysed by RPD and ferrographic methods and compared. Lubricant samples containing a fine test dust were also analysed by RPD and AF to determine the effectiveness of the RPD's centripetal acceleration and the AF's gravity flow in eliminating non-ferrous contamination from the sample. Finally, comparisons were made between actual lubricant samples from a turbine engine simulator analysed by RPD and ferrographic methods. For lubricant samples with relatively small amounts of non-ferrous contamination or samples where non-ferrous particles are important in machine health condition assessment, the AF and DR ferrographs are equal or superior to RPD for lubricant analyses. For samples with high levels of non-ferrous contamination, the RPD is superior in eliminating unwanted non-magnetic particles and permitting a less obstructed view of the important ferrous wear debris particles.     

Comparison of different preparation methods of biological samples for FIB milling and SEM investigation

When a new approach in microscopy is introduced, broad interest is attracted only when the sample preparation procedure is elaborated and the results compared with the outcome of the existing methods. In the work presented here we tested different preparation procedures for focused ion beam (FIB) milling and scanning electron microscopy (SEM) of biological samples. The digestive gland epithelium of a terrestrial crustacean was prepared in a parallel for FIB/SEM and transmission electron microscope (TEM). All samples were aldehyde-fixed but followed by different further preparation steps. The results demonstrate that the FIB/SEM samples prepared for conventional scanning electron microscopy (dried) is suited for characterization of those intracellular morphological features, which have membranous/lamellar appearance and structures with composition of different density as the rest of the cell. The FIB/SEM of dried samples did not allow unambiguous recognition of cellular organelles. However, cellular organelles can be recognized by FIB/SEM when samples are embedded in plastic as for TEM and imaged by backscattered electrons. The best results in terms of topographical contrast on FIB milled dried samples were obtained when samples were aldehyde-fixed and conductively stained with the OTOTO method (osmium tetroxide/thiocarbohydrazide/osmium tetroxide/thiocarbohydrazide/osmium tetroxide). In the work presented here we provide evidence that FIB/SEM enables both, detailed recognition of cell ultrastructure, when samples are plastic embedded as for TEM or investigation of sample surface morphology and subcellular composition, when samples are dried as for conventional SEM.     

几种波片位相延迟测量方法的比较

建立了用于波片位相延迟测量的高精度多功能测试系统,实现了在同一测试系统上应用不同的测试方法,使测试具有可比性。对光谱扫描法、Soleil补偿器法及两种光强法进行了比较测量,并结合测试结果对各种方法进行了系统的误差分析。分析结果表明,对本测试系统而言,光谱扫描法适于测量λ/2波片,测量精度主要由单色仪的精度所决定,其误差＜0.032%;Soleil补偿器法测量精度主要由Soleil补偿器精度决定,其误差＜0.18%;光强法误差与待测波片的延迟量有关,且测试光路与偏振器的缺陷对测量精度影响较大。光谱扫描法与Soleil补偿器法测量λ/2波片的结果在误差范围内相符,光强法测量结果产生较大差异的原因来自光路、偏振器及方法本身的缺陷。实验结果为不同条件下选择最佳测量方法提供了参考依据。     

发光菌急性毒性分析方法的比较和优化

从发光菌急性毒性测试方法的原理出发,分析现行国标方法的缺陷,并深入比较国外主流仪器的优缺点,进而总结出一套可靠性更高的急性毒性测试方法。方法涵盖实验分析、干扰分析、数据处理、质量控制等方面。同时,针对现有方法研究中的不足,提出建立青海弧菌Q67毒性数据库及真和发光菌研发两个方向,为发光菌急性毒性测试方法新国标的修制定提供参考。     

Development of advanced quantitative analysis methods for wear particle characterization and classification to aid tribological system diagnosis

Classification of wear particles is performed in two steps, i.e. first a particle to be classified is characterized by surface feature parameters such as roughness, directionality, homogeneity, periodicity, etc. and then, the particle is assigned to a specific class using these parameters. However, a significant limitation of this approach is that surface parameters are often not unique to a specific surface topography and their values may change significantly with scale, orientation angle and position at which the particle data was acquired. Various attempts were made to overcome this limitation by selecting a core set of parameters which ensures that wear particles are accurately classified. However, the parameter selection is usually cumbersome and requires lengthy computation. Furthermore, there is no guarantee that parameters selected are sensitive enough to separate particles belonging to different classes. Thus, a new classification technique based entirely on dissimilarity measures (e.g. Euclidean, Baddeley's distances), calculated between surface images of an unclassified particle and classified particles, was developed. The classification process is based on assigning a particle to a class of particles with the smallest dissimilarity measure. This idea arises naturally from the two facts: (i) compactness, similar objects are in close proximity to each other in their representation space, while different objects are far apart, and (ii) true representation, if objects are close to each other in their representation space they belong to the same class. In this paper, an overview of recent advances and developments in the area of particle classification based on dissimilarity measures is presented with a particular emphasis on constructing a simple and accurate classifier.     

Transient suppression in FRF measurement: Comparison and analysis of four state-of-the-art methods

This paper compares four well selected methods for computing the non-parametric Frequency Response Function (FRF) of a periodically excited linear time invariant system. The suppression of the transient is mandatory when its influence in the data is large. Better suppression of the transient leads to a better non-parametric FRF estimate. A good non-parametric FRF estimate can be used to validate the parametric transfer function model in a second step. The suppression of the transient will be highlighted using the mean squared error of the non-parametric FRF estimate. Temperature transients caused by heat diffusion are used as example. The selected methods consist of two standard windowing methods and two methods based on the Local Polynomial Method (LPM). LPM was designed to find a non-parametric FRF estimate in the presence of nonlinearities. This paper will modify LPM to find a non-parametric FRF estimate for linear systems using a single experiment. The mean squared error of the four non-parametric FRF estimates will be compared and analyzed, based on a simulation and a measurement example.     

星载超光谱成像仪摆镜系统轴系的误差分析与设计

摆镜系统轴系的精度会影响星载超光谱成像仪的成像质量。轴系设计时采用了向心推力球轴承来调整轴系的装配误差。分析了摆镜系统工作时影响轴系精度的几个误差源,包括零件的尺寸误差、同轴度误差、配合误差等,按照误差理论设计并计算了各误差值,得到轴系的径向晃动误差和角运动误差。对加工好的轴系进行精度检测,实验结果符合理论结果,验证了该摆镜系统的轴系结构设计的合理性和可靠性,为该类航天运动部件的轴系结构设计提供了参考依据。     

An automatic algorithm for the segmentation and morphological analysis of microvessels in immunostained histological tumour sections

A fully automatic segmentation and morphological analysis algorithm for the analysis of microvessels from CD31 immunostained histological tumour sections is presented. Development of the algorithm exploited the distinctive hues of stained vascular endothelial cells, cell nuclei and background, to provide the seeds for a 'region-growing' method for object segmentation in the 3D hue, saturation, value (HSV) colour model. The segmented objects, identified as microvessels by CD31 immunostaining, were post-processed with three morphological tasks: joining separate objects that were likely to belong to a single vessel, closing objects that had a narrow gap around their periphery, and splitting objects with multiple lumina into individual vessels. The automatic segmentation was validated against a hand-segmented set of 44 images from three different SW1222 human colorectal carcinomas xenografted into mice. 96.3 ± 0.9% of pixels were found to be correctly classified. Automated segmentation was carried out on a further 53 images from three histologically distinct mouse fibrosarcomas (MFs) for morphological comparison with the SW1222 tumours. Four morphometric measurements were calculated for each segmented vessel: vascular area (VA), ratio of lumen area to vascular area (lu/VA), eccentricity (e), and roundness (ro). In addition, the total vascular area relative to tumour tissue area (rVA) was calculated. lu/VA, e and ro were found to be significantly smaller in MF tumours than in SW1222 tumours (p < 0.05; unpaired t-test). The algorithm is available through the website http://www.caiman.org.uk where images can be uploaded, processed and sent back to users. The output from CAIMAN consists of the original image with boundaries of segmented vessels overlaid, the calculated parameters and a Matlab file, which contains the segmentation that the user can use to derive further results.     

Facile green synthesis approach for the production of chromium oxide nanoparticles and their different in vitro biological activities

Green synthesis of nanoparticles using plants has become a promising substitute for the conventional chemical synthesis methods. In the present study, our aim was to synthesize chromium oxide nanoparticles (Cr₂O₃NPs) through a facile, low‐cost, eco‐friendly route using leaf extract of Rhamnus virgata (RV). The formation of Cr₂O₃NPs was confirmed and characterized by spectroscopic profile of UV–Vis, EDX, FTIR, and XRD analyses. The UV‐visible spectroscopy has confirmed the formation of Cr₂O₃NPs by the change of color owing to surface plasmon resonance. The bioactive functional groups present in the leaf extract of RV involved in reduction and stabilization of Cr₂O₃NPs were determined by FTIR analysis. Based on XRD analysis, crystalline nature of Cr₂O₃NPs was determined. The morphological shape and elemental composition of Cr₂O₃NPs were investigated using SEM and EDX analyses, respectively. With growing applications of Cr₂O₃NPs in biological perspectives, Cr₂O₃NPs were evaluated for diverse biopotentials. Cr₂O₃NPs were further investigated for its cytotoxicity potentials against HepG2 and HUH‐7 cancer cell lines (IC₅₀: 39.66 and 45.87 μg/ml), respectively. Cytotoxicity potential of Cr₂O₃NPs was confirmed against promastigotes (IC₅₀: 33.24 μg/ml) and amastigotes (IC₅₀: 44.31 μg/ml) using Leishmania tropica (KMH₂₃). The Cr₂O₃NPs were further evaluated for antioxidants, biostatic, alpha‐amylase, and protein kinase inhibition properties. Biocompatibility assay was investigated against human macrophages which confirmed the nontoxic nature of Cr₂O₃NPs. Overall, the synthesized Cr₂O₃NPs are biocompatible and nontoxic and proved to possess significant biopotentials. In future, different in vivo studies are needed to fully investigate the cytotoxicity and mechanism of action associated with these Cr₂O₃NPs.     

Sequential quantitative X-ray elemental imaging of frozen-hydrated and freeze-dried biological bulk samples in the SEM

Planed frozen-hydrated (FH) bulk biological samples of chicken retina were analysed by X-ray elemental imaging in a scanning electron microscope and reanalysed after freeze-drying in the microscope column. Sequential elemental imaging of the same bulk sample in this way provides improved information on element distributions. There was no evidence of element redistribution during the freeze-drying process. Quantitative elemental images were obtained and interpreted to deduce relative and absolute element concentrations in different regions of the retina. Water concentrations were determined from the difference in oxygen concentrations at 15 kV and 5 kV in FH and freeze-dried (FD) samples, respectively. Two accelerating voltages were used to maintain similar X-ray excitation volumes. Water concentrations were also estimated by relating measured oxygen concentration in FH samples to the concentration of oxygen in solutions of a generalized protein in water and by comparing concentrations of phosphorous or sulphur in the FH and FD states.     

肉制品中淀粉含量测定方法的比较及关键点

本文论述了测定肉制品中淀粉含量的两种方法,一种是食品中淀粉含量的测定方法-淀粉酶水解法,另一种是肉制品中淀粉含量的测定方法-碘量法,详细阐述了这两种方法的原理、试剂设备、步骤,并对这两种方法的稳定性、适用性、优劣、关键点,以及测定结果进行了分析比较.结果表明两种方法均适用于肉制品中淀粉含量检测,但方法一更加简便准确.     

An automated feedback system with the hybrid model of scoring and classification for solving over-segmentation problems in RNAi high content screening

Background: High content screening (HCS) via automated fluorescence microscopy is a powerful technology for generating cellular images that are rich in phenotypic information. RNA interference is a revolutionary approach for silencing gene expression and has become an important method for studying genes through RNA interference‐induced cellular phenotype analysis. The convergence of the two technologies has led to large‐scale, image‐based studies of cellular phenotypes under systematic perturbations of RNA interference. However, existing high content screening image analysis tools are inadequate to extract content regarding cell morphology from the complex images, thus they limit the potential of genome‐wide RNA interference high content screening screening for simple marker readouts. In particular, over‐segmentation is one of the persistent problems of cell segmentation; this paper describes a new method to alleviate this problem. Methods: To solve the issue of over‐segmentation, we propose a novel feedback system with a hybrid model for automated cell segmentation of images from high content screening. A Hybrid learning model is developed based on three scoring models to capture specific characteristics of over‐segmented cells. Dead nuclei are also removed through a statistical model. Results: Experimental validation showed that the proposed method had 93.7% sensitivity and 94.23% specificity. When applied to a set of images of F‐actin‐stained Drosophila cells, 91.3% of over‐segmented cells were detected and only 2.8% were under‐segmented. Conclusions: The proposed feedback system significantly reduces over‐segmentation of cell bodies caused by over‐segmented nuclei, dead nuclei, and dividing cells. This system can be used in the automated analysis system of high content screening images.     

Adaptive Monte Carlo and GUM methods for the evaluation of measurement uncertainty of cylindricity error

Measurement uncertainty is one of the most important concepts in geometrical product specification (GPS). The “Guide to the expression of uncertainty in measurement (GUM)” is the internationally accepted master document for the evaluation of uncertainty. The GUM method (GUMM) requires the use of a first-order Taylor series expansion for propagating uncertainties. However, when the mathematical model of measurand is strongly non-linear the use of this linear approximation may be inadequate. Supplement 1 to GUM (GUM S1) has recently been proposed based on the basis of probability density functions (PDFs) using the Monte Carlo method (MCM). In order to solve the problem that the number of Monte Carlo trials needs to be selected priori, adaptive Monte Carlo method (AMCM) described in GUM S1 is recommended to control over the quality of the numerical results provided by MCM.The measurement and evaluation of cylindricity errors are essential to ensure proper assembly and good performance. In this paper, the mathematical model of cylindricity error based on the minimum zone condition is established and a quasi particle swarm optimization algorithm (QPSO) is proposed for searching the cylindricity error. Because the model is non-linear, it is necessary to verify whether GUMM is valid for the evaluation of measurement uncertainty of cylindricity error. Then, AMCM and GUMM are developed to estimate the uncertainty. The procedure of AMCM scheme and the validation of GUMM using AMCM are given in detail. Practical example is illustrated and the result shows that GUMM is not completely valid for high-precision evaluation of the measurement uncertainty of cylindricity error if only the first-order terms in the Taylor series approximation are taken into account. Compared with conventional methods, not only the proposed QPSO method can search the minimum zone cylindricity error precisely and rapidly, but also the Monte Carlo simulation is adaptive and AMCM can provide control variables (i.e. expected value, standard uncertainty and lower and higher coverage interval endpoints) with an expected numerical tolerance. The methods can be extended to the evaluation of measurement uncertainty of other form errors such as roundness and sphericity errors.     

New methods of track membrane treatment in the preparation of samples for further observation with scanning electron microscopy

Track membranes are porous systems consisting of a polymer foil with thin channels (i.e. pores) piercing it from surface to surface. The creation of non‐cylindrical pores in a track membrane is important for the optimization of membrane characteristics, i.e. the highest productivity at the required selectivity. A new method of cleavage preparation (the irradiation of track membrane samples with accelerated electrons) for the observation of channel shapes directly in the membrane cross‐sections is presented. Diagrams showing the tensile and burst strengths as a function of the irradiation dose, and images of surfaces and cleavages of track membrane samples are presented in this work. The changes in the pore sizes and shapes along the channel were clearly seen. These results can be used for the optimization of track membrane production.     

Current analytical methods for the detection of dityrosine, a biomarker of oxidative stress, in biological samples

Dityrosine is a fluorescent molecule formed as a result of normal posttranslational processing. In many structural proteins, dityrosine confers resistance to proteolysis and physicochemical trauma as a stabilizing crosslink. Dityrosine has also been found in oxidative/nitrative stress under a variety of conditions and biological systems. In this regard, it has been used as an important biomarker for oxidatively modified proteins during UV and gamma-irradiation, aging, and exposure to oxygen free radicals, nitrogen dioxide, peroxynitrite, and lipid hydroperoxides. Renewed interest in dityrosine and other tyrosine oxidation products as clinical indicators of oxidative modification has driven the development of important techniques for the specific analysis and quantification of these molecules. The presence of elevated levels of dityrosine in mammalian tissue and urine samples has been measured by chromatographic separation followed by mass spectrometry GC-MS and HPLC-MS/MS. Increases in dityrosine levels have been associated with pathologies such as eye cataracts, atherosclerosis, acute inflammation, and Alzheimer's disease. The continued development of, and increased accessibility to, improved mass spectrometric instrumentation will expand the capability, feasibility, and sensitivity with which specific biomarkers like dityrosine can be measured.     

Development of a sensitive closed batch vessel hydride generation atomic absorption spectrometry method for the determination of cadmium in aqueous samples

Hydride generation atomic absorption spectrometry was used as a simple, sensitive, and economical method for the determination of cadmium at trace levels. This study includes a stepwise optimization of all parameters that have an impact on the production, transport, and atomization of cadmium hydride for rapid determination due to the unstable nature of the hydride produced. The cell temperature for atomization, pump period, and flow rate of carrier gas were varied, and the best conditions were defined to obtain lower detection limits. In addition, the acid and sodium borohydride concentrations were optimized since they directly affect the production of the cadmium hydride. Under optimum experimental conditions, the hydride generation efficiency was greatly enhanced to record optimized analytical signals. The accuracy of the method was determined by the analysis of a certified reference material, and the results were in good agreement with the certified value. A linear dynamic range for cadmium was observed from 0.10 to 2.0?µg/L. High sensitivity was obtained with a limit of detection of 0.029?µg/L.     

A comparison of some methods for measuring the size distribution of two-dimensionally clustered latex particles in TEM images using mathematical morphology

The particle size distribution of polymer latexes of, for example, polybutadiene can be measured on transmission electron micrographs using image analysis methods. By taking suitable measures in preparing the sample it can be ensured that the image only consists of projections of two-dimensional clusters of adhering latex spheres. For the deagglomeration of these clusters a number of methods based on mathematical morphology are discussed. Speed and accuracy of the methods were tested on a personal-computer-based image analysis system and compared to manual measurements. Repeated openings or erosions applied to a binary image are relatively fast global methods with a low accuracy due to the fact that particle diameters are measured in whole numbers of pixels; only the horizontal diameters of rectangular pixels are measured. With an iterative method the individual components of the clusters are found by repeated conditional thickening of the erosion kernels. The method is accurate but very slow due to the large number of processing steps. The watershed method as applied to the original grey-scale transmission electron micrograph makes use of the sharp local maxima which arise at the interface of the projection of two adhering particles. This method is as accurate as the iterative method, but about eight times as fast.