Application of rhodamine B thiolactone to fluorescence imaging of Hg in Arabidopsis thaliana

Exposure to mercury causes severe damage to plants, animals and even humans. Concern over mercury toxicity has encouraged the development of efficient, sensitive, and selective methods for the in vivo detection of mercury. Although a variety of studies have been published describing fluorescence imaging of mercury in animal cells and tissues, no in vivo monitoring has been developed for plant systems until now. In this paper, we report the semi-quantitative fluorescence imaging of Hg²⁺ ions in a common model plant Arabidopsis thaliana (A. thaliana), with rhodamine B thiolactone (RBS) as a novel Hg²⁺ probe. The experimental results show that RBS is plant cell wall and cell membrane permeable, and the probe responds selectively to Hg²⁺ ions instead of the other species in plant systems. Real-time monitoring of Hg²⁺ absorption in roots of A. thaliana by RBS shows that saturation of Hg²⁺ uptake could occur in a short period of 3 days at most. The transportation and accumulation of Hg²⁺ ions in roots of A. thaliana have also been studied, revealing that most of Hg²⁺ ions reside in root cap and meristematic zone, and only a small amount of Hg²⁺ ions can reach the maturation zone. This indicates that the interaction of Hg²⁺ ions with any Hg²⁺-philic species including proteins in these regions may be responsible for plant poisoning and even death.     

Gradient convergence filters and a phase congruency approach for in vivo cell nuclei detection

Computational methods used in microscopy cell image analysis have largely augmented the impact of imaging techniques, becoming fundamental for biological research. The understanding of cell regulation processes is very important in biology, and in particular confocal fluorescence imaging plays a relevant role for the in vivo observation of cells. However, most biology researchers still analyze cells by visual inspection alone, which is time consuming and prone to induce subjective bias. This makes automatic cell image analysis essential for large scale, objective studies of cells. While the classic approach for automatic cell analysis is to use image segmentation, for in vivo confocal fluorescence microscopy images of plants, such approach is neither trivial nor is it robust to image quality variations. To analyze plant cells in in vivo confocal fluorescence microscopy images with robustness and increased performance, we propose the use of local convergence filters (LCF). These filters are based in gradient convergence and as such can handle illumination variations, noise and low contrast. We apply a range of existing convergence filters for cell nuclei analysis of the Arabidopsis thaliana plant root tip. To further increase contrast invariance, we present an augmentation to local convergence approaches based on image phase information. Through the use of convergence index filters we improved the results for cell nuclei detection and shape estimation when compared with baseline approaches. Using phase congruency information we were able to further increase performance by 11% for nuclei detection accuracy and 4% for shape adaptation. Shape regularization was also applied, but with no significant gain, which indicates shape estimation was good for the applied filters.     

Integration of a hematopoietic progenitor cell program using the ACT/DB database system

Infusion of hematopoietic progenitor cells following high-dose chemotherapy is frequently used to treat patients with hematological malignancies and solid tumors. We have developed a comprehensive software system to monitor these patients once they are entered into an experimental protocol. The captured data encompasses all phases of progenitor cell therapy including progenitor cell mobilization and collection, stem cell processing, as well as cell infusion and engraftment kinetics. Particular attention was paid to the quality assurance and quality control functionality of the software during development of data entry forms and reports. The system was developed using the ACT/DB client-server database, which utilizes Microsoft Access as a front-end and accesses either an Oracle or SQL Server database. ACT/DB has been modified for deployment on the Internet in order to take advantage of Web-based technology. Information technology can help to integrate the diverse data requirements of complex therapeutic trials.     

模数混合控制光伏并网型逆变电源的设计

描述了一种模数混合控制实现的光伏并网逆变器,介绍了在CPU速度受限制的条件下,采用模数混合电路实现运算控制并跟踪并网电流的方案。同时对光伏并网型逆变电源的结构及组成做了详细讨论。根据所提出的方案研制了一台三相并网逆变器,并网电流跟踪具有较高的分辨率,电流谐波小、噪声低,该逆变器已成功应用于20KWp的大型光伏并网示范电站。     

基于微流控芯片的钙离子浓度检测系统的实现

在细胞内物质定量分析中引入微流控芯片.利用微流控芯片完成细胞的培养、染色、试剂的进给等生物实验功能.设计了用于细胞内钙离子浓度检测的微流控荧光检测系统.通过双波长激发,利用荧光检测系统完成荧光强度和图像的采集.同时研究比值荧光法,计算出定量检测的钙离子浓度.实验结果表明,此检测装置可靠性高,检测结果准确.这一研究,提供了一种细胞检测新手段.为细胞研究提供更加便捷的细胞培养、检测、试剂进给一体化检测装置.     

Multiple hypothesis tracking based on the Shiryayev sequential probability ratio test基于Shiryayev序贯概率比检测的多假设跟踪算法

To date, Wald sequential probability ratio test (WSPRT) has been widely applied to track management of multiple hypothesis tracking (MHT). But in a real situation, if the false alarm spatial density is much larger than the new target spatial density, the original track score will be very close to the deletion threshold of the WSPRT. Consequently, all tracks, including target tracks, may easily be deleted, which means that the tracking performance is sensitive to the tracking environment. Meanwhile, if a target exists for a long time, its track will have a high score, which will make the track survive for a long time even after the target has disappeared. In this paper, to consider the relationship between the hypotheses of the test, we adopt the Shiryayev SPRT (SSPRT) for track management in MHT. By introducing a hypothesis transition probability, the original track score can increase faster, which solves the first problem. In addition, by setting an independent SSPRT for track deletion, the track score can decrease faster, which solves the second problem. The simulation results show that the proposed SSPRT-based MHT can achieve better tracking performance than MHT based on the WSPRT under a high false alarm spatial density.     

Automated analysis of two- and three-color fluorescent Elispot (Fluorospot) assays for cytokine secretion

The Elispot effectively measures the frequencies of cells secreting particular molecules, especially low-frequency cells such as antigen-specific T cells. The Fluorospot assay adapted this analysis to two products per cell, and this has now been extended to three-color measurement of both mouse and human cytokine-secreting cells. Due to the increased data complexity, and particularly the need to define single-, double- and triple-producing cells, it is critical to objectively quantify spot number, size, intensity, and coincidence with other spots. An automated counting program, Exploraspot, was therefore developed to detect and quantify Fluorospots in automated fluorescence microscope images. Morphological parameters, including size, intensity, location, circularity and others are calculated for each spot, exported in FCS format, and further analyzed by gating and graphical display in popular flow cytometry analysis programs. The utility of Exploraspot is demonstrated by identification of single-, double- and triple-secreting T cells; tolerance of variable background fluorescence; and estimation of the numbers of genuine versus random multiple events.     

介电泳芯片的研制及不同细胞介电分离的应用

制备了包括指状交叉、城墙状和梯形的微电极阵列芯片装置.并用这些芯片探索了生物细胞的介电响应.另外观察了酵母和鸡血红细胞的迁移、旋转和融合以及几种细胞收集图片.发现了两种细胞的正、负介电泳现象,确定了这两种细胞的分离条件.讨论了两种细胞正、负介电泳的原因.利用同一芯片在相同的条件下一种细胞移向强场区(正介电泳),另一种细胞移向弱场区(负介电泳).因此可用同一芯片分离不同的细胞.有望建立一种非接触式细胞分离技术,而且在分离过程中不需要添加任何试剂.     

Microfluidic immobilization and subcellular imaging of developing <Emphasis Type="Italic">Caenorhabditis elegans</Emphasis>

Caenorhabditis elegans has been an essential model organism in the fields of developmental biology, neuroscience, and aging. However, these areas have been limited by our ability to visualize and track individual C. elegans worms, especially at the subcellular scale, over the course of their lifetime. Here we present a microfluidic device to culture individual C. elegans in parallel throughout post-embryonic development. The device allows for periodic mechanical immobilization of the worm, enabling 3D imaging at subcellular precision. The immobilization is sufficient to enable fluorescence recovery after photobleaching (FRAP) measurements on organelles and other substructures within the same specific cells throughout larval development, without the use of chemical anesthetics. Using this device, we measure FRAP recovery of two nucleolar proteins in specific intestinal cells within the same worms during larval development. We show that these proteins exhibit different fluorescence recovery as the worm grows, suggesting differential protein interactions during development. We anticipate that this device will help expand the possible uses of C. elegans as a model organism, enabling its use in addressing fundamental questions at the subcellular scale.     

超声波在干细胞治疗中的应用进展

超声波是指频率在 20 kHz 以上的机械波,具有机械效应、热效应和空化效应。不同强度的超声波作用于生物体,会对其器官、组织和细胞等产生不同程度的生物学效应。随着工程技术的发展,超声波已在医学显像、实体肿瘤治疗等生物医学领域具有广泛应用,低强度超声波也成为辅助干细胞治疗各类疾病的重要工程手段。深入了解超声波在干细胞治疗中所起的作用及机制,将促进广大研究者对超声波技术的了解,从而重视并加大超声波在该领域的研究力度及广泛应用。该文综述了超声波在干细胞治疗中的研究进展,对超声促进干细胞增殖、分化和迁移、细胞示踪显影及靶向药物传递等前沿研究进展进行了全面介绍,并对超声波在干细胞治疗领域的应用前景提出了设想。     

A new instrument for passive remote sensing: 1. Measurements of sunlight-induced chlorophyll fluorescence

Under natural sunlight illumination, the chlorophyll fluorescence emitted by the vegetation represents less than 3% of the reflected light in the near infrared part of the spectrum. This small amount is difficult to quantify except at certain wavelengths, where the solar spectrum is attenuated (Fraunhofer lines). An instrument measuring the in-filling of the atmospheric oxygen absorption band at 760 nm by chlorophyll fluorescence has been designed and constructed at the “Laboratoire pour l'Utilisation du Rayonnement Electromagnetique” in Orsay, France. The system was calibrated against a pulsed fluorimeter (FIPAM), especially developed for monitoring chlorophyll fluorescence at distance. The penetration of diuron, a herbicide acting on photosynthesis, was monitored by the passive instrument for several days on a corn canopy. A good agreement was found between gas exchange and variable chlorophyll fluorescence at the canopy level and variable fluorescence at the leaf level. The potential application of the passive chlorophyll fluorescence measurements for long range vegetation remote sensing is discussed.     

3D level set method for blastomere segmentation of preimplantation embryos in fluorescence microscopy images

Detailed and accurate characteristics of preimplantation embryos are fundamental for a deep understanding of their development. Recent studies indicate that various geometric features of cells, such as size, shape, volume, and position play a significant role in embryo growth. However, a quantitative assessment of these characteristics first needs a segmentation of the individual cells. The manual separation and labeling of cells is extremely inefficient, and an automated approach is highly desirable. This paper presents an automatic method for early stage embryo segmentation into its constituent cells and membranes using three-dimensional (3D) data. The input data consist of two Z-stacks of fluorescence microscope images containing nuclei and membranes. The method uses a 3D level set segmentation algorithm. Its evaluation is based on a dataset composed of 20 mouse embryos, each with 4–32 blastomeres. Segmentation accuracy was evaluated by calculating F-scores with ground truth obtained by manually labeling desired regions. We also compared output of our method with the one acquired with a watershed algorithm. The proposed approach was able to achieve more than \(90\%\) accuracy for embryos with 4 and 8 cells, while for embryos with higher number of cells it was lower, reaching \(75\%\) for 32-cell embryo.     

Development and application of a three-dimensional artificial visual system

A three-dimensional artificial visual system has been developed to aid in the analysis of 3-D fluorescence images of smooth muscle cells. The system consists of three sets of 3-D spatial filters that decompose the image to enable a simple recombination algorithm to locate the discrete bodies of protein concentration in a cell, classify the concentration bodies as globular or oval, and determine the 3-D orientation of the oval bodies. A graphic model of the protein concentration is created from the data provided by the artificial visual system. Patterns of organization in the distribution of the protein bodies are investigated using an interactive graphics system.     

Industrially oriented voice control system

The purpose of my research was to develop a novel voice control system for the use in the robotized manufacturing cells as well as to create tools providing its simple integration into manufacturing. A comprehensive study of existing problems and their possible solutions has been performed. Unlike some other works, it focused on the specific requirements that should be fulfilled by industrially oriented voice control systems. Analysis of existing solutions related to the natural language processing and those related to various voice control applications has been performed. Its goal was to establish the optimal method of voice command analysis for industrially oriented systems. Finally, a voice control system for manufacturing cells has been developed, implemented and practically verified in the laboratory. Unlike many other solutions, it takes into consideration almost all aspects of voice command processing (speech recognition, syntactic and semantic analysis and spontaneous speech effects) and – most importantly – their mutual influence. To provide the simple system customization (integration into any particular manufacturing cells), a special format for quasi-natural sublanguage syntax definition has been developed. A novel algorithm for semantic analysis, using specific features of voice commands used for controlling industrial devices and machines, has been incorporated into the system. Successful implementation in the educational robotized machining cell shows that industrial applications should be possible in the very next future.     

An optimal track maintenance scheduling model analysis taking the risk of accidents into consideration

This paper presents an optimal track maintenance scheduling model developed for maintaining adequate conditions of the railway tracks and accomplishing an efficient management of the railway service. Our model has been used in several major Japanese railway companies to develop optimal track maintenance schedules (OTMS). Since train accidents cause enormous losses (not only for the railway company but also for society), we propose to take into account the risk of train accidents while developing the OTMS. Thus, it is critically important to identify measures to quantitatively evaluate the risk of accidents to ensure safe and stable transportation. We propose a method to estimate track maintenance costs and the related costs associated with the risk of train derailment due to longitudinal level irregularities of the railway track. We attempt to show a desirable condition of railway tracks by applying the accident cost estimation method and the OTMS model to an actual railway section. We also demonstrate how to apply these models to actual railway networks in general by validating the estimation results using actual numerical data.     

一种基于微流控芯片技术的流式细胞计数系统

采用聚二甲基硅氧烷（PDMS）材料制作微流控流式细胞计数芯片,利用负压驱动与鞘液夹流技术实现样品的水力聚焦,达到了10μm的样品聚焦宽度。基于激光诱导荧光技术,制作了小型化的检测装置。以488nm固体激光器为光源,激光束以45°方向穿过一个水平狭缝,以线光源形式汇聚到微流控芯片的检测区域,并与微通道垂直交叉,细胞样品的荧光信号通过光电倍增管收集。整个分析系统结构简单,操作方便,灵敏度高,可初步实现细胞的计数。     

Computer-assisted analysis of the vascular endothelial cell motile response to injury

We have developed an automated, user-friendly method to track vascular endothelial cell migration in vitro using an IBM PC/XT with MS DOS. Analog phase-contrast images of the bovine aortic endothelial cells are converted into digital images (8 bit, 250 x 240 pixel resolution) using a Tecmar Video VanGogh A/D board. Digitized images are stored at selected time points following mechanical injury in vitro. FORTRAN and assembly language subroutines have been implemented to automatically detect the wound edge and the edge of each cell nucleus in the phase-contrast, light-microscope field. Detection of the wound edge is accomplished by intensity thresholding following noise reduction in the image and subsequent sampling of the wound. After the range of wound intensities is determined, the entire image is sampled and a histogram of intensities is formed. The histogram peak corresponding to the wound intensities is subtracted, leaving a histogram peak that gives the range of intensities corresponding to the cell nuclei. Rates of cell migration, as well as cellular trajectories and cell surface areas, can be automatically quantitated and analyzed. This inexpensive, automated cell-tracking system should be widely applicable in a variety of cell biologic applications.     

在体生物光学成像技术的研究进展

在体生物发光成像和在体荧光成像是近年来新兴的在体生物光学成像技术, 能够无损实时动态监测被标记细胞在活体小动物体内的活动及反应, 在肿瘤检测、基因表达、蛋白质分子检测、药物受体定位、药物筛选和药物疗效评价等方面具有很大的应用潜力. 本文详细介绍了在体生物发光成像和在体荧光成像的特点、系统及应用, 比较了它们的异同, 综述了在体生物光学成像技术的基本原理和应用领域, 讨论了将其应用于临床的进一步发展方向.     

基于深度学习网络模型的端到端航迹关联

为提高雷达数据处理中航迹关联的智能性,充分利用目标的特征信息,并简化系统处理流程,提出了一种基于深度学习网络模型的端到端航迹关联算法。首先分析了基于神经网络的航迹关联存在样本细节少、处理流程繁杂的问题,然后提出了端到端的深度学习模型。该模型根据航迹关联数据的处理特征,改进了卷积神经网络结构用于特征提取,充分利用了长短期记忆网络对历史信息和将来信息的处理能力,并分析了前后航迹的关联性。在对原始数据进行卡尔曼滤波后,将全部航迹信息特征作为输入,并由基于卷积神经网络特征提取的长短期记忆深度神经网络模型直接输出航迹关联结果。仿真结果表明,提出的模型可以充分学习推演目标的多个特征信息,具有较高的航迹关联准确率,对航迹关联的智能化分析具有一定的参考价值。     

基于模糊膨胀模型的细胞核轮廓提取方法

针对细胞核之间经常出现重叠聚堆的现象，提出了一种新的基于模糊膨胀模型的细胞核轮廓提取方法．结合细胞核的椭圆边界信息，将图像数据映射到反映与细胞核颜色和位置关系的多个模糊域；基于这些模糊映射关系，建立了一种新的主动轮廓模型进行细胞核轮廓跟踪．采用一种自适应的膨胀机制帮助曲线克服局部极小值快速膨胀直到收敛到真实边界．多种信息的融合使得模型具有较强的边界跟踪能力．实验表明，对细胞核边界的残缺或重叠部分具有较好的分割效果，且分割性能很稳定．