Laser pressure catapulting followed by B actin gene identification in Japanese quail macrochromosomes and microchromosomes using teflon-coated coverslip slides

Laser microdissection of individual mammalian chromosomes (> 2 µm) has been achieved though the use of a microscope slide coated with a polyethylene naphthalate (PEN) membrane. Although these slides have proved sufficient for larger chromosomes, they are insufficient for small chromosomes (< 1 µm). We have developed a new type of slide which allows laser microdissection of single Japanese quail microchromosomes (0.5 µm) and macrochromosomes (3–4 µm). To test the usefulness of these slides, a Japanese quail single nucleus, a macrochromosome, and a microchromosome were collected with Laser pressure catapulting, the B-actin gene was PCR amplified, and sequenced. The resulting PCR product was confirmed by nucleotide sequencing to be B-actin. These newly developed slides were shown to facilitate the laser microdissection of both Japanese quail macrochromosomes and microchromosomes.     

A new semi-automatic device for autoradiographic coating

A semi-automatic device is described for the coating of slides with nuclear emulsions for autoradiography. The design embodies a metal heatsink for warming the emulsion, and an elevating gantry which withdraws coated slides at a controlled constant speed.     

TOOL LIFE PREDICTION USING RANDOM WALK BAYESIAN UPDATING

According to the Taylor tool life equation, tool life reduces with increasing cutting speed. The influence of additional factors can also be incorporated. However, tool wear is generally considered a stochastic process with uncertainty in the model constants. In this work, Bayesian inference is applied to predict tool life for milling/turning operations using the random walk/surface methods. For milling, Bayesian inference using a random walk approach is applied to the well-known Taylor tool life model. Tool wear tests are performed using an uncoated carbide tool and AISI 1018 steel work material. Test results are used to update the probability distribution of tool life. The updated beliefs are then applied to predict tool life using a probability distribution. For turning, both cutting speed and feed are considered. Bayesian updating is performed using the random surface technique. Turning tests are completed using a coated carbide tool and forged AISI 4137 chrome alloy steel. The test results are applied to update the probability distribution of tool life and the updated beliefs are used to predict tool life. While this work uses the Taylor model, by following the procedures described here, the technique can be applied to other tool life models as well.     

Spot counting to locate fetal cells in maternal blood and tissue: a comparison of manual and automated microscopy

BACKGROUND: Fetal cell detection in maternal tissue requires an accurate, efficient, and reproducible microscopy method. Our objective was to compare manual scoring to a commercially available automated scanning system for the detection of chromosome signals by fluorescence in situ hybridization (FISH). METHODS: X and Y chromosome FISH signals were detected on slides of calibrated mixtures of blood, paraffin-embedded liver sections, and post-termination blood. For manual scoring (400x magnification), the number of cells located and duration of scoring were recorded. For automated scanning using the Metasystems Metafer3/Metafer4 Scanning System (200x magnification), duration of scanning, number of gallery images generated, duration of manual review of gallery images, and number of confirmed fetal cells were recorded. RESULTS: From all slides the number of target fetal cells located by manual and automated microscopy was highly correlated (r = 0.90). However, automated scanning required on average 4-fold more time than manual scoring (P < 0.0001), with an average automated scanning time of 9.7 h per slide compared with 2.4 h per slide when scored manually. CONCLUSIONS: In general, the accuracy of automated and manual microscopy is comparable, although manual scoring is more efficient because of the level of magnification necessary for automated scanning of cells, and a large number of gallery images generated by automated scanning that must then be reviewed manually. This suggests that when rapid analysis is required (i.e., clinical situations), manual microscopy is preferable. In contrast, automated scanning may have advantages over manual microscopy when time constraints are less imposed (i.e., research situations).     

X-ray microanalytical resolution in frozen-hydrated biological bulk samples

In coated frozen-hydrated gelatin gels the backscattered electron yield does not increase during electron irradiation as it does in uncoated samples. Neither is the backscattered electron yield greater from coated frozen-hydrated gels than that from more conductive organic samples. This is interpreted as indicating that a significant distortion of the electron interaction volume, due to the development of a space charge, does not occur in electron irradiated frozen-hydrated gelatine gels when they are coated with a conducting coat. The depth resolution as estimated from models of biological samples in the form of frozen-hydrated photographic film and frozen-hydrated sections of gelatine gel is consistent with that computed from X-ray depth distribution curves, i.e. close to 2.0 μm at 15 kV. Lateral resolution was estimated from photographic film to be close to 2.0 μm also, at 15 kV.     

Unstained and in vivo fluorescently stained bacterial nucleoids and plasmolysis observed by a new specimen preparation method for high-power light microscopy of metabolically active cells

Microscope slides were coated with a layer of gelatin, the thickness of the gelatin increasing linearly along the long axis. The bacterial suspension is applied to the dried gelatin and covered by a coverslip. The medium is absorbed by the gelatin and thus the cells applied against the coverslip. By this method, cultures of concentrations below 10⁸ cells/ml provide statistically relevant numbers for observation without prior concentration steps. It is easier to apply than the existing methods for the observation of bacterial nucleoids by phase contrast imaging. Because the cells are maintained in growing conditions the method is useful for the vital fluorescence DAPI-staining of various bacterial species and for observations of plasmolysis and its reversal at different physiological conditions and extracellular osmolalities. The previously generally assumed view that the plasmolytic changes of the cell morphology are immediate upon the hyperosmotic shock and are rapidly repaired when the cell is able to metabolize actively was confirmed; this is in contrast to some recent claims.     

实现微米级灭菌范围控制的微细等离子体射流装置

为了实现微细等离子体的精准灭菌,设计了新型微细等离子体射流装置,并对该装置产生的含氧活性粒子(Reac-tive Oxygen Species,ROS)和含氮活性粒子(Reactive Nitrogen Species,RNS)分布范围及其灭菌范围进行研究.淀粉碘化钾混合溶液里的碘离子可以被微细等离子体射流产生的ROS...     

The preparation and X-ray microanalysis of bulk frozen hydrated vacuolate plant tissue

A series of modifications have been devised which allow the peak to background ratio X-ray analytical method to be used more effectively to measure elemental concentrations in large vacuolate plant cells. Planar, frozen-hydrated fracture faces of bulk plant tissue are coated with a thin film of evaporated chromium, which prevents surface charging. Provided the film is sufficiently thin, c. 5–10 nm, there is no attenuation of the electron beam and only a small absorption of soft X-rays. The chromium makes a small but measurable contribution to the spectral background and suitable corrections may be made to the quantitative results. An improved back-scattered imaging system is described, which helps to overcome the problem of spurious X-ray signals from rough surfaces. The microscope column has been modified to permit a continuous readout of beam current, sensu stricta, during X-ray microanalysis and to allow rapid exchange of the electron gun assembly during low temperature operation. Calculations are given relating the size of the X-ray interactive volume to electron penetration and X-ray emission in both frozen hydrated and frozen dried cells. The problems of X-ray microanalysis are discussed in relation to the highly vacuolate cells found in most mature plant tissues and an example given of the distribution of four major cations in tobacco leaves.     

基于PROE曲柄滑块机构的运动仿真及分析

在各类机械传动结构中,曲柄滑块有着广泛的应用,根据曲柄滑块机构设计的原理,提出在Pro/E中实现曲柄滑块设计及实体造型的方法,并利用Pro/E的运动分析模块（Mechanism）对曲柄滑块进行运动分析和仿真,提供对曲柄滑块的优化设计。     

Perceptual clustering for automatic hotspot detection from Ki‐67‐stained neuroendocrine tumour images

Hotspot detection plays a crucial role in grading of neuroendocrine tumours of the digestive system. Hotspots are often detected manually from Ki‐67‐stained images, a practice which is tedious, irreproducible and error prone. We report a new method to segment Ki‐67‐positive nuclei from Ki‐67‐stained slides of neuroendocrine tumours. The method combines minimal graph cuts along with the multistate difference of Gaussians to detect the individual cells from images of Ki‐67‐stained slides. It, then, automatically defines the composite function, which is used to determine hotspots in neuroendocrine tumour slide images. We combine modified particle swarm optimization with message passing clustering to mimic the thought process of the pathologist during hotspot detection in neuroendocrine tumour slide images. The proposed method was tested on 55 images of size 10 × 5 K and resulted in an accuracy of 94.60%. The developed methodology can also be part of the workflow for other diseases such as breast cancer and glioblastomas.     

Low-friction wear-resistant coatings for high-temperature foil bearings

Compliant foil bearings offer many advantages over rolling element bearings in high-speed and high-temperature applications. However, implementation of foil bearings in these applications requires development of solid lubricant coatings that can survive the severe operating conditions encountered at high speeds and high temperatures. The objective of this paper is to present results on development of an advanced coating system for use with compliant foil bearings that permits higher operating speeds and temperatures. In order to evaluate the coating performance and to select the best coating combination for implementation, tests were conducted using a high-temperature, high-speed tribometer. In these tests, Inconel test substrates, representative of a portion of a foil bearing, were coated with several different Korolon^TM coatings. The counterface disks were coated with a dense chrome, plasma sprayed PS304, hard chrome and Korolon^TM 1350B. Each test was conducted for 500 start–stop cycles up to 810 °C foil pad temperature under 13.8 kPa normal loading.The test results confirmed the excellent tribological behavior of Korolon^TM coatings for high-speed, high-temperature foil bearing applications. While the tribological behavior of Korolon^TM coatings were determined to be a function of temperature, in most cases a minimum coefficient of friction less than 0.1 was observed during startup/shutdown periods. Based on the measured coefficient of friction and post-test visual inspection of the mating surfaces, the hard chrome coating proved unacceptable for high-temperature applications due to extensive surface cracking. The other disk coatings exhibited excellent tribological performance.Following these tests, a foil journal bearing was designed and a composite coating consisting of Korolon^TM 1350A with an overcoat of Korolon^TM 800 was applied to the bearing top foil; and a dense chrome coating was applied to the journal surface. The foil bearing and journal were installed in a 240-lb thrust turbojet engine and operated successfully to 54,000 rpm for over 70 start–stop cycles and 14 h.     

Correlated light optical and scanning electron microscopy of Gram smears of bacteria and paraffin sections of cardiac muscle

Light-microscope slides (3 in. × 1 in.) bearing Gram smears of Erysipelothrix rhusiopathiæ, or Staphylococcus aureus, after preliminary examination under the light-optical microscope (LM), were cut down in size, glued onto specimen stubs, coated with gold and examined in the scanning electron microscope (SEM). These preparations served as a control for investigations into bacteria-cell junctions in tissue. Cover-slips from stained sections of staphylococcal or swine erysipelas endocarditis mounted on 3 in. × 1 in. microscope slides (which had been intensively studied previously with conventional light microscopy) were floated off by immersing the slides in xylol. After dehydration of the tissues on the slides, the preparations were treated similarly to the Gram smears, and were examined with the SEM. Lesions of endocarditis were thus examined, and the information gained from these preliminary examinations shed new light on the pathogenesis of the disease. This information had not previously been available by any other technique. Because of this, and in view of the simplicity of preparing sections for scanning electron microscopy, it is suggested that the SEM might be a useful tool to be applied to routine histological sections.     

Capsule-supporting ring: a new device for resin embedding of glass-mounted specimens

The goal of specimen preparation for transmission electron microscopy is to obtain high-quality ultra-thin sections with which we can correlate cellular structure to physiological function. In this study, we newly developed a capsule-supporting ring that can be useful for resin embedding of glass-mounted specimens. The present device allowed us to re-embed a semi-thin section on a microscope slide into a resin block not only for efficient ultra-thin sectioning but also for a correlative light and electron microscopy. Similar to epoxy resins for morphological observations, semi-thin sections of low-viscosity hydrophilic resins, such as Lowicryl series, can be re-embedded into the resin, which can be useful for cytochemical gold labelling. A further application of the present device improved flat embedding of cultured cells on glass cover slips for electron microscopy, preserving in situ sub-cellular structures close to their native state. We practically describe the use of capsule-supporting ring and demonstrate representative micrographs as results.     

A novel consistent photomicrography technique using a reference slide made of neutral density filter

Obtaining consistent photomicrographic images of pathology slides is not always easy because of many different types and settings of the equipment such as the microscopes and digital cameras. In this study, we developed a photomicrography technique that could acquire consistent images of pathology slides. The neutral density (ND) filter was attached to a transparent glass slide as a reference slide, photographed using consistent settings, and acquired images that harbored all of the areas of gray, white, and black. In the same way, the slide was replaced by the actual pathology slide, and photomicrographed. To simulate different light environment, the above photographic session was repeated using two different light intensities and microscopes. A graphic program was used to adjust levels of the reference slide images and this leveling, or calibration, was saved as a file for each. This file for leveling process was applied to actual subsequent photomicrographic images. The same sites of noncalibrated and calibrated images of the pathology slide were calculated into CIELAB or CIE L*a*b* coordinates. Then, the color differences (ΔE*ab) were calculated. As results, in the study using a 50% transmittance ND filter, two original different images were made nearly identical to the unaided eye, especially in two‐point (white and gray) and three‐point (black, white, and gray) leveling. In comparison of different light intensities, the ΔE*ab of the selected area was 0.9 in two‐point leveling. Between different microscopes, 10.7 of ΔE*ab was the smallest value in three‐point leveling. This method would be helpful for acquiring consistent photomicrographic images of pathology slides. Microsc. Res. Tech., 2011. © 2010 Wiley‐Liss, Inc.     

Backscattered electron SEM imaging of cells and determination of the information depth

Backscattered electron imaging of HT29 colon carcinoma cells in a scanning electron microscope was studied. Thin cell sections were placed on indium‐tin‐oxide‐coated glass slides, which is a promising substrate material for correlative light and electron microscopy. The ultrastructure of HT29 colon carcinoma cells was imaged without poststaining by exploiting the high chemical sensitivity of backscattered electrons. Optimum primary electron energies for backscattered electron imaging were determined which depend on the section thickness. Charging effects in the vicinity of the SiO₂ nanoparticles contained in cell sections could be clarified by placing cell sections on different substrates. Moreover, a method is presented for information depth determination of backscattered electrons which is based on the imaging of subsurface nanoparticles embedded by the cells.     

AutoCAD下幻灯片文件的应用及其批量制作幻灯片文件的方法

AutoCAD环境下的幻灯片文件是一个屏幕显示的快照,目前主要应用在AutoCAD环境下进行二次开发所涉及的用户化界面设计方面.本文尝试了AutoCAD环境下幻灯片文件在零件资料库和用脚本文件播放幻灯片两方面的应用,研究了使用VisualLISP批量制作幻灯片的方法.     

光学显微镜载玻片防断裂装置

在生物学实验中 ,尤其在暗室操作时 ,显微镜 (以下所称显微镜均指光学显微镜 )载玻片常被油镜镜头压碎。而一个简单的装置解决了这个问题 ,弥补了许多显微镜设计上的不足。     

Thermal problem of friction when the plane-parallel layer-base tribosystem brakes

The boundary problem of heat conductivity in the plane-parallel layer and semi-infinite base tribosystem is solved analytically. The layer slides over the base surface with speed that recedes linearly in time. The thermal contact is nonfull and the bodies exchange heat with the environment by convection. The effect of the heat exchange and thermal conductivity factors on the temperature distribution in the cermet layer-cast-iron base tribocouple is analyzed.     

Fine Structure of Bacterial Adhesion to the Epithelial Cell Membranes of the Filiform Papillae of Tongue and Palatine Mucosa of Rodents: A Morphometric,TEM, and HRSEM Study

The palatine mucosa and filiform papillae of the dorsal tongue mucosae of rodents were examined using transmission electron microscopy (TEM) and high resolution scanning electron microscopy (HRSEM). In the HRSEM method, the samples were fixed in 2% osmium tetroxide, dehydrated in alcohol, critical point‐dried, and coated with gold‐palladium. In addition, the HRSEM technique was used for morphometric analysis (length, width, and length/width ratio of cocci and bacilli). For the TEM method, the tissues were fixed in modified Karnovsky solution (2.5% glutaraldehyde, 2% formalin in 0.1M sodium phosphate buffer, pH 7.4) and embedded in Spurr resin. The results demonstrated that there are thick polygonal keratinized epithelial cells where groups of bacteria are revealed in three‐dimensional images on the surface of filiform papillae in these animals. The bacterial membranes are randomly attached to the microplicae surface of epithelial cells. Morphometrics showed higher values of length and width of cocci in newborn (0 day) as compared to newborn (7 days) and adults animals, the bacilli showed no differences in these measurements. At high magnification, the TEM images revealed the presence of glycocalyx microfilaments that constitute a fine adhesion area between bacterial membranes and the membranes of epithelial microplicae cells. In conclusion, the present data revealed the fine fibrillar structures of bacteria that facilitate adhesion to the epithelial cell membranes of the oral cavity and morphometric changes in newborn (0 day) rats as compared with other periods. Microsc. Res. Tech. 76:1226–1233, 2013. © 2013 Wiley Periodicals, Inc.     

Visualization of the nucleoid in living bacteria on poly-lysine coated surfaces by the immersion technique

A method for visualizing the nucleoid in living bacteria is described. The method involves immobilization of the cells by attachment to a poly-l-lysine coated slide and subsequent immersion in bovine serum albumin solutions of defined refractive index and osmolality. Dependence of the visibility of the nucleoid on the refractive index of the medium is investigated.