Brush-like fibers on the human chromosome periphery

A brush-like border apparently composed of fibers protruding from metaphase chromosomes of human lymphocytes was observed for the first time using transmission (TEM) and scanning electron microscopy (SEM). On the basis of size and sensitivity to colcemid, the fibers may be related to microtubules and spindle organization. The brush-like fibers were observed when chemically fixed metaphase chromosome spreads were placed on glass slides and maintained under "wet" conditions (not allowed to air dry after fixation for conventional cytogenetic protocols) until postfixation protocols for TEM and SEM were performed. The purpose of this study was to establish the occurrence of the brush-like fibers and to determine the effects of colcemid on these fibers.     

Atomic force microscopy applied to study macromolecular content of embedded biological material

We demonstrate that atomic force microscopy represents a powerful tool for the estimation of structural preservation of biological samples embedded in epoxy resin, in terms of their macromolecular distribution and architecture. The comparison of atomic force microscopy (AFM) and transmission electron microscopy (TEM) images of a biosample (Caenorhabditis elegans) prepared following to different types of freeze-substitution protocols (conventional OsO4 fixation, epoxy fixation) led to the conclusion that high TEM stainability of the sample results from a low macromolecular density of the cellular matrix. We propose a novel procedure aimed to obtain AFM and TEM images of the same particular organelle, which strongly facilitates AFM image interpretation and reveals new ultrastructural aspects (mainly protein arrangement) of a biosample in addition to TEM data.     

Irregular Shaped,Assumably Semi‐Crystalline Calciumphosphate Platelet Deposition at the Mineralization Front of Rabbit Femur Osteotomy: A HR‐TEM Study

Although bone minerals have been widely studied by various techniques in previous studies, crystal structures, morphology of bone minerals and its building pathway remained still controversy. In this work, the ultrastructure of the mineralization front of rabbit femur has been studied by conventional and high‐resolution (HR) transmission electron microscopy (TEM). In order to induce a healing and demineralization process the animals were subjected to a standardized osteotomy stabilized with titan screws and sonic pins. After 84 days follow‐up time the newly build bone was investigated. The mineralization front of rabbit femur osteotomy contains partly mineralized collagen fibrils with a pronounced striped pattern together with a large number of agglomerated apatite platelets. The striation is caused by mineralization in the hole zones of the collagen fibrils, corresponding to the early stage of mineralization. In the TEM micrographs, the mineralization zone appears denser and compact when compared with fully mineralized bone, although most of the collagen fibrils are completely mineralized in the latter (higher concentration of interfibrillar apatite platelets within the mineralization zone). In bone some partly mineralized collagen fibrils are also observed, revealing the same arrangement, regular shape, and size of apatite platelets as collagen fibrils in the mineralization zone. Apatite platelets with irregular shapes are observed at the vortex‐shaped outer boundary of the mineralization zone, i.e. at the interfaces with nonmineralized collagen or osteoblasts. HR TEM micrographs reveal that the platelets are assumably semicrystalline and that within the platelet nanocrystalline domains of apatite are embedded in an amorphous calciumphosphate matrix. SCANNING 35: 169‐182, 2013. © 2012 Wiley Periodicals, Inc.     

Effect of medicaments used in endodontic regeneration on the morphological characteristics of bovine radicular dentin: Experimental immature tooth model

The purpose of this study was to evaluate the effect of different combinations of irrigating solutions and intracanal dressings in the pretreatment of bovine radicular dentin, using an experimental immature tooth model. Eighty healthy bovine teeth, simulated with incomplete rhizogenesis, were randomly distributed according to the protocols of root canal dentin pretreatment for a regenerative endodontic procedure (n = 10): Control (irrigation with distilled water); SH (irrigation with 1.5% Sodium Hypochlorite); EDTA (irrigation with 17% EDTA); SH/EDTA (irrigation with 1.5% SH + 17% EDTA); SH/CH/EDTA (irrigation with 1.5% SH + calcium hydroxide paste +17% EDTA); SH/MTAP/EDTA (irrigation with 1.5% SH + modified triple antibiotic paste + EDTA 17%); SH/TAP/EDTA (irrigation with 1.5% SH + triple antibiotic paste +17% EDTA) and SH/DAP/EDTA (irrigation with 1.5% SH + double antibiotic paste + EDTA 17%). After the completion of the protocol, the demineralization, the exposure of collagen fibers, and the dentin erosion was evaluated under scanning electron microscopy (SEM), by applying a score system (1–3) to classify the observed features. Statistical analysis was performed (Kruskal‐Wallis and Dunn Multiple Comparison tests—p < .05). SH/TAP/EDTA and SH/DAP/EDTA groups presented the highest rates of demineralization in both the coronal and middle thirds of the root (p < .05). In the SH/MTAP/EDTA group, the samples presented moderate demineralization. The samples from the SH/CH/EDTA group presented similar findings to the control group (p < .05). Conventional triple antibiotic (TAP) and double antibiotic (DAP) pastes promoted more pronounced morphological changes on the dentin surface.     

Side draw control design for a high purity multi-component distillation column

Industrial methanol production involves a multi component feed containing methanol, water and trace levels of ethanol being refined to produce AA grade methanol at high product recovery. Due to practical constraints, the bottoms discharge of the column is primarily water with only trace of methanol impurities. As a result of these constraints, ethanol, which is a non-key middle boiling component gets “trapped” near the side draw of the column forming an ethanol bulge, which in turn results in non-linear, inverse, time and state varying behaviour of the side draw ethanol composition. In this work, we established that the existence of the ethanol bulge creates the complex process behaviour of the side draw ethanol composition and that this bulge needs to be explicitly controlled. This type of explicit composition bulge analysis and subsequent control has not been attempted on methanol distillation columns before. For this purpose a novel, robust and practical side draw control scheme to detect and remedy the excess ethanol bulge movement using override control is presented. The side draw controller, together with other regulatory controllers is shown to maintain on-specification operations of the column. Disturbance rejection tests carried out illustrate that the side draw control scheme will keep the column operating within commercial specification. It is also shown that a traditional DV control structure is unable to achieve this objective.     

Glycoproteins and skin-core structure in Nephila clavipes spider silk observed by light and electron microscopy

Microscopical imaging of natural, unstressed draglines or of untreated bulk samples showed two types or threads with diameters of either approximately 1-2 microm or 4-5 microm, which could be identified as products of the minor or major ampullate glands. The threads had a circular profile in serial cross sections and are surrounded by a thin outer layer of a different material within the section. Such fibrillar configurations were also found in untreated threads or in the same serial sections of transmission electron microscopy (TEM) samples by means of the special technique of laser scanning microscopy. In TEM slides, numerous cavities with the same circular profile were detectable, and the length of these cavities is variable from 40-300 nm. The threads are oriented parallel and twisted around themselves to construct a double thread. In the interface between the two single threads, bridge-like structures are prominent. The single untreated thread consists of cylindrical fibers with a diameter of approximately 1-1.5 microm. Apparently more than eight fibers are within a thread and each fiber is composed of a great number of fibrils with a diameter of about 150 nm. The surface of threads is coated with a characteristic layer approximately 150-250 nm thick that contains glycoproteins. These were demonstrated for the first time by labeling with concanavalin A lectin-gold complex and are dependent on the diameter and length of the thread. The same substances could also be detected inside the single thread. The skin can be removed completely or partially by mechanical treatment, or by washing with phosphate-buffered saline or trypsin.     

Ultrastructure of motor nerve terminals in the anterior third of wistar rat tongue

The nerve terminals of intrinsic muscular fibers of the tongue of adult wistar rats was studied by using silver impregnation techniques, transmission electron microscopy (TEM), and high resolution scanning electron microscopy (HRSEM) to observe the nerve fibers and their terminals. Silver impregnation was done according to Winkelman and Schmit, 1957 . For TEM, small blocks were fixed in modified Karnovsky solution, postfixed in 1% buffered osmium tetroxide solution, and embedded in Spurr resin. For HRSEM, the parts were fixed in 2% osmium tetroxide solution with 1/15 M sodium phosphate buffer (pH 7.4) at 4°C for 2 h, according to the technique described by Tanaka, 1989 . Thick myelinated nerve bundles were histologically observed among the muscular fibers. The intrafusal nerve fiber presented a tortuous pathway with punctiform terminal axons in clusters contacting the surface of sarcolemma. Several myelinated nerve fibers involved by collagen fibers of the endoneurium were observed in HRSEM in three-dimensional aspects. The concentric lamellae of the myelin sheath and the axoplasm containing neurofilaments interspersed among the mitochondria were also noted. In TEM, myofibrils, mitochondria, rough endoplasmic reticulum, Golgi's apparatus, and glycogen granules were observed in sarcoplasm. It is also noted that the sarcomeres constituted by myofilaments with their A, I, and H bands and the electron dense Z lines. In areas adjacent to muscular fibers, there were myelinated and unmyelinated nerve fibers involved by endoneurium and perineurium. In the region of the neuromuscular junction, the contact with the sarcolemma of the muscular cell occurs forming several terminal buttons and showing numerous evaginations of the cell membrane. In the terminal button, mitochondria and numerous synaptic vesicles were observed. Microsc. Res. Tech., 2009. © 2009 Wiley-Liss, Inc.     

Electron microscopic study of novel threadlike structures on the surfaces of mammalian organs

The ultrastructures of novel threadlike structures (NTSs) and corpuscles on the surfaces of internal organs of rats were investigated using electron microscopy. The samples were studied in situ by using a stereomicroscope and were taken for further morphological analysis. Scanning electron microscope (SEM) images revealed a bundle structure of threadlike tissue, which was composed of several 10-micro m-thick subducts. The surfaces of the corpuscles were rather coarse and fenestrated. The corpuscles had cucumber-like shapes with an average length of about 2 mm and a thickness of about 400 micro m. Transmission electron microscope (TEM) images disclosed disordered collagen fibers, which formed the extracellular matrix of the threadlike tissue, and immune-function cells, like macrophages, mast cells, and eosinophils. Sinuses of various diameters, which were thought to be cross-sections of the lumens of the subducts, were observed in the TEM, cryo-SEM and focused-ion-beam SEM images. These SEM images were obtained for the first time to reveal the detailed structure of the NTSs that were only recently discovered.     

Comparing different preparation methods to study human fibrin fibers and platelets using TEM

For the study of cellular ultrastructure, the sample needs to be stabilized by fixation, with the ultimate aim to preserve the native tissue organization and to protect the tissue against later stages of preparation. Chemical and freezing fixation are most used, and chemical fixation employs agents that permeate tissues and cells by diffusion and covalently bind with their major biochemical constituents to fix them. Most widely used chemical fixatives are aldehydes, e.g., formaldehyde and glutaraldehyde, which are noncoagulating, crosslinking agents. Cryofixation methods for ultrastructural studies are also popular, and high-pressure freezing immobilizes all cell constituents and arrests biological activity by removing the thermal energy from the system. In the current research, we used platelet-rich plasma (PRP) to study expansive fibrin fibers and platelet ultrastructure to compare the two fixation techniques. We also used thrombin and calcium chloride as a clotting agent to determine the technique most suitable for the formation of extensive fibrin networks. Chemically fixated fibrin fibers were more compact and condensed and also showed a banding pattern on longitudinal sections. High-pressure frozen samples were more dispersed while platelets fixated showed better preserved cellular membranes and organelle structure. PRP coagulated by addition of CaCl(2) showed blood platelets that are noticeably more activated compared with PRP; however, with thrombin, a sharp ultrastructure was seen. We conclude that PRP mixed with thrombin, and freeze substituted, is the most suitable method for the study of extensive fibrin fibers as well as platelets.     

Optimizing the ethanol content of shear thickening fluid/fabric composites under impact loading

New dampers are provided by reinforced fibers in a full density network. Their mechanism is on their network structures that disperse the projectile energy in a wide area. Shear thickening fluid (STF) is made from the combination of nanoparticles with a specified weight fraction in polyethylene glycol to increase damping in network structures. To immerse fibers, STF is diluted in ethanol, and the fibers are placed therein for a specified period to impregnate all fibers in the fluid. To eliminate the sample ethanol, specimens are heated at 60°C to 80°C. Combining ethanol with fluid and eliminating ethanol reduce the sensitivity of the composite to the impact. Energy dissipation is highest in the 3:1 portion (ethanol and STF) because of the insignificant influence on the nanoparticle interface and polymer. However, decreasing the amount of ethanol makes the dispersion of STF unsuitable.     

Scanning and transmission electron microscopy for evaluation of order/disorder in bone structure

A comparative characterization of the structure of normal and abnormal (osteoporotic) human lumbar and thoracic vertebrae samples was carried out to reveal the type of possible disorder. Samples from the bone fragments extracted during the surgery due to vertebra fractures were examined by scanning electron microscopy (SEM), conventional and high resolution transmission electron microscopy (TEM and HRTEM), and X-ray energy dispersive spectroscopy (EDS). Contrary to what might be expected in accordance with possible processes of dissolution, formation and remineralization of hard tissues, no changes in phase composition of mineral part, crystal sizes (length, width, and thickness), and arrangement of crystals on collagen fibers were detected in abnormal bones compared to the normal ones. The following sizes were determined by HRTEM for all bone samples: 相似文献   

Freeze-substitution of chemically stabilized samples for biological field emission scanning electron microscopy.

The high resolution imaging capabilities of modern field emission scanning electron microscopes require adequately improved tissue preparation procedures to prevent the collapse of macromolecular structures and the extraction of molecules. A routine cryo-stabilization technique is described which utilizes chemical crosslinking and cryo-dehydration for mechanical and chemical stabilization of protein and lipid structures and increase of electrical conductivity of the sample. Thiocarbohydrazide (TCH) serves as a general mordant for osmium tetroxide crosslinking. However, extensive washing after all impregnation steps is necessary to dissolve unspecific osmium black precipitations at the sample surface. Collagen I aggregates showed increased stability against collapse after TCH osmification alone, whereas pulmonary surfactant liposomes require additional freeze-substitution in methanol and Freon 113 for stabilization during critical point drying. Environmental scanning electron microscopy (at water vapor pressures of 5-10 torr within the specimen chamber) was used to control, in the wet phase, the stabilization procedure at the level of chemical crosslinkage. It could be confirmed that tannic acid, often used to stabilize lipids, leads to artificial rearrangement of bilayered liposomes into compact presumable multilayered bodies, whereas the TCH osmification preserved liposome structures and their aggregates. The increase of electrical conductivity of sliced tissue was demonstrated on kidney. Support technologies for the cryo-stabilization procedures are described in detail, as well as simple routines for first stabilization trials with new samples. On pulmonary tissue, the excellent preservation of alveolar shape and fine structures of intermediate forms of surfactant are described.     

Chemical preparation of dielectrics for studying their microtopography by the SEM

A procedure for the chemical preparation of dielectrics to make their surfaces conductive is proposed. Its simplicity and rapidity, as well as the possibility of using it to study etching kinetics of specimen surface morphology are noteworthy. The procedure is demonstrated by the interaction of F-apatite with acids. The procedural steps include careful attention to the washing of the specimen following its interaction with an acid and to its drying thereafter. The composition of the washing solution and the washing time must be carefully controlled. Positive results are obtained for the interaction between F-apatite and phosphoric, hydrochloric, nitric acids and between glass and hydrofluoric acid. Acetone was employed as the washing solution and in all cases sharp images of the specimen surface were obtained in the SEM. The subsequent washing of the specimen in water restored the dielectric properties of its surface. By sequentially etching, drying, and observing the specimen in the SEM, a series of images was obtained of an area of the surface as it was being etched. This series of images permitted determination of the growth rate of etched pits.     

快速测定含水溶液中乙醇的氧同位素比值

近年来,乙醇的氧同位素比值（δ¹⁸O）在果汁和饮料酒真实性鉴别中起着重要作用,利用该指标可检测产品中的外源水、追溯产品的产地。本工作采用多孔聚合物气相色谱柱实现了水与乙醇的在线快速分离,建立了溶剂稀释后直接用气相色谱-裂解-稳定同位素比值质谱（GC-TC-IRMS）测定溶液中乙醇δ¹⁸O值的方法。实验结果表明,该方法可排除水对乙醇δ¹⁸O分析的干扰,乙醇浓度在1%~100%时测定稳定性良好,在不同水溶液中乙醇δ¹⁸O的测定值保持一致;乙醇δ¹⁸O值重复性和再现性的标准偏差均优于0.5‰,并通过欧盟实验室间能力验证（FIT-PTS）证明了方法的准确性。该方法具有样品用量少（仅需70～200 μL）、分析速度快（约18 min）、操作简单方便等特点,可为乙醇δ¹⁸O在果汁和饮料酒真实性领域的研究与应用提供方法参考。     

面向异构电力设备状态监测的低功耗OPC UA协议转换最小系统

电力系统中不同厂家的现场设备接口各异,使得实际使用中现场设备很难被集成,形成一个信息孤岛。面向电力设备运行状态监测需求,结合当前电力装备普遍采用的Modbus RTU协议和各类PLC协议等为主要现场数据采集接口的现状,进行现有接口协议的OPC UA转换单元设计,研制异构电力设备OPC UA协议转换的嵌入式最小系统硬件平台,建立基于OPC UA协议标准化的统一输出接口。该硬件平台利用软件配置的方式实现现场数据采集智能设备统一接入,可高效支撑以OPC UA协议为输出的异构标准化跨平台运行,有利于同时支持更多不同种类采集终端云边协同参数采集的实现。     

Oolong tea and LR‐White resin: a new method of plant sample preparation for transmission electron microscopy

Simplifying sample processing, shortening the sample preparation time, and adjusting procedures to suitable for new health and safety regulations, these issues are the current challenges which electron microscopic examinations need to face. In order to resolve these problems, new plant tissue sample processing protocols for transmission electron microscopy should be developed. In the present study, we chose the LR‐White resin‐assisted processing protocol for the ultrastructural observation of different types of plant tissues. Moreover, we explored Oolong tea extract (OTE) as a substitute for UA in staining ultrathin sections of plant samples. The results revealed that there was no significant difference between the OTE double staining method and the traditional double staining method. Furthermore, in some organelles, such as mitochondria in root cells of tomatoes and chloroplast in leaf cells of watermelons, the OTE double staining method achieved little better results than the traditional double staining method. Therefore, OTE demonstrated good potentials in replacing UA as a counterstain on ultrathin sections. In addition, sample preparation time was significantly shortened and simplified using LR‐White resin. This novel protocol reduced the time for preparing plant samples, and hazardous reagents in traditional method (acetone and UA) were also replaced by less toxic ones (ethanol and OTE).     

Preparation of TEM foils from Nb-10 a/o Si.

Ductile phase toughened composites contain phases with significantly different physical properties. Consequently, these phases thin at different rates depending on the sample preparation procedure. A new TEM foil preparation method for the ductile phase toughened Nb-10 a/o Si material has been developed. The method involves chemical thinning in a 70% nitric acid/30% hydrofluoric acid solution followed by electropolishing in a 12.5% sulfuric acid/87.5% methanol electrolyte at -40 degrees C. This procedure for making TEM foils results in large thin areas with the minimum of artifacts. Mechanical grinding of a sample followed by either ion milling, dimpling, or electropolishing produced foils with large electron transparent areas, but with uncharacteristic features of the original Nb-10 a/o Si alloy microstructure. These artifacts were identified as dislocations, surface mottling, and antiphase domains.     

X-ray microanalysis of cultured keratinocytes: methodological aspects and effects of the irritant sodium lauryl sulphate on elemental composition

Irritant substances have been shown to induce elemental changes in human and animal epidermal cells in situ . However, skin biopsies are a complicated experimental system and artefacts can be introduced by the anaesthesia necessary to take the biopsy. We therefore attempted to set up an experimental system for X-ray microanalysis (XRMA) consisting of cultured human keratinocytes. A number of methodological aspects were studied: different cell types, washing methods and different culture periods for the keratinocytes. It was also investigated whether the keratinocytes responded to exposure to sodium lauryl sulphate (SLS) with changes in their elemental composition. The concentrations of biologically important elements such as Na, Mg, P and K were different in HaCaT cells (a spontaneously immortalized non-tumorigenic cell line derived from adult human keratinocytes) compared to natural human epidermal keratinocytes. The washing procedure and time of culture influenced the intracellular elemental content, and rinsing with distilled water was preferred for further experiments. Changes in the elemental content in the HaCaT cells compatible with a pattern of cell injury followed by repair by cell proliferation were seen after treatment with 3.33 µ m and 33 µ m SLS. We conclude that XRMA is a useful tool for the study of functional changes in cultured keratinocytes, even though the preparation methods have to be strictly controlled. The method can conceivably be used for predicting effects of different chemicals on human skin.     

Ethanol sensing properties of PVP electrospun membranes studied by quartz crystal microbalance

In this study, ethanol gas sensing of PVP fiber membranes based on Quartz Crystal Microbalance (QCM) was investigated. The fibers were deposited on the QCM’s electrodes by electrospinning the viscous blend solutions of PVP. The effects of PVP concentration on morphology of the fibers and their permeability when exposed to ethanol were investigated. Membranes which were prepared using low concentration solutions, contained beads and high packing fibers and showed low permeability. Increase of the PVP concentration to 12 wt% resulted in continuous fine fibers with good permeability. Furthermore, higher PVP concentrations were found to decrease ethanol permeability due to larger fiber radius and lower surface area. Moreover, effects of fibrous layer thickness on gas response was examined. Consequently, optimum amount of PVP concentration and fibrous layer thickness for the best gas response was found.     

A silver impregnation utilizing only reagent-grade chemicals for visualization of peripheral axons and fibroblasts

A silver impregnation procedure is described that enables the representation of numerous tissue components. It especially visualizes nerves and fibroblasts, which may be clearly distinguished from other tissue elements. Since it can be performed on thick sections, three-dimensional analysis of nerve terminations and fibroblasts in the tissues can be performed. The results are illustrated with the innervation of the rat snout and human labial sweat glands for nerves, and with bovine and pathological human material for fibroblasts. Axons are visualized as thin, sinuous black structures, sometimes, as in the case of autonomic efferents, with varicosities. Fibroblasts are revealed in their total extent by the darker staining of their nuclei and cytoplasm compared with that of the surrounding collagen. Cell processes can thus be followed for long distances, and may be seen to approach other cells.
Previously published methods for the visualization of nerves and fibroblasts depended upon the use of commercial formalin, which is subject to the manufacturers' modifications. The method presented here uses exclusively analytical-grade reagents and distilled water. It is also less dependent than other methods on the fixation protocol.