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We have investigated the role of plasminogen activator inhibitor 1 (PAI-1) in the regulation of fibrinolysis using a model thrombus composed of thrombin-stimulated platelets, fibrin(ogen), plasminogen, and recombinant tissue-type plasminogen activator. Laser light scattering kinetic measurements showed that clot lysis was significantly delayed both by thrombin-stimulated platelets and their cell-free releasate. This delay in lysis was almost fully reversed by the addition of a PAI-1-specific monoclonal antibody that blocks the ability of PAI-1 to inhibit plasminogen activators. Lysis half-times exhibited a linear dependence on the concentration of PAI-1 antigen present, as determined by enzyme-linked immunosorbent assay (ELISA). Sodium dodecylsulfate-polyacrylamide gel electrophoresis (SDS-PAGE) followed by immunoblotting confirmed the presence of PAI-1 antigen in the platelet releasates. Scanning electron micrographs of the model thrombus components sampled late in lysis showed considerable unproteolyzed fibrin still attached to platelets. Immunogold cytochemistry detected large amounts of PAI-1 antigen in the partially lysed platelet-fibrin thrombi. This PAI-1 appeared to be bound to the fibrin network rather than to the platelet surface itself. We conclude that the residual clots observed late in lysis represent platelet-associated fibrin to which platelet-released PAI-1 has bound, rendering it less susceptible to degradation.

Time-dependent thrombolytic resistance is a critical problem in thrombolytic therapy for acute myocardial infarction. Platelets have been regarded as the main source of plasminogen activator inhibitor-1 (PAI-1) found in occlusive platelet-rich clots. However, endothelial cells are also known to influence the fibrinolytic capacity of blood vessels, but their ability to actively mediate time-dependent thrombolytic resistance has not been fully established. We will show that, in vitro, tumor necrosis factor-α–stimulated endothelial cells secrete large amounts of PAI-1 over a period of hours, which then binds to fibrin and protects the clot from tissue plasminogen activator–induced fibrinolysis. In vivo, endothelial cells covering atherosclerotic plaques are influenced by cytokines synthesized by plaque cells. Therefore, we propose that continuous activation of endothelial cells in atherosclerotic blood vessels, followed by elevated PAI-1 secretion and storage of active PAI-1 in the fibrin matrix, leads to clot stabilization. This scenario makes endothelial cells a major factor in time-dependent thrombolytic resistance.

Phloem proteins are specifically presented in phloem tissues of monocotyledons and dicotyledons. Ultrastructural studies showed that they were filamentous, spherical, tubular, crystalline and amorphous in shaped They were present in the sieve elements and sometimes are also found in companion cells and phloem parenchyma cells. The diversities in shape and biogenesis of plant p-protein in relation to plant species and developmental stage of phloem tissues were reported. In the present study, a 42.5 kDA protein obtained from phloem exudate of Luffa cylindricafruits were examined with amino acid composition, immnoblotting, and immunolocalization.Phloem exudates leaking from Luffa cylindricafruits by a simple razor blade cutting were mixed with the extraction medium containing antioxidant and rapidly separated by 2-dimensional gel of native and sodium dodecylsulfate (SDS) polyacrylamide gel electrophoresis (PAGE).

Recent initiatives have resulted in a considerable increase in our understanding of the structure of the nuclear pore complex (NPC). The biochemical factors involved in both import and export have been rapidly characterised, with steady progress in the molecular dissection of the structural elements of the NPC, which is a unit of considerable molecular architecture (MW 125 kD), comprising an estimated 50- 100 different proteins. Despite this progress, the crucial molecular interactions involved in the mechanics of transport through the central transporter of the NPC remain unclear. NPC structure in Diptera, fish, (Fig 1) amphibians, birds and mammals shows a high degree of evolutionary conservation. 3D reconstructions of isolated yeast NPCs, show that the core structure is very similar to ‘higher’ organisms, but peripheral structures may be considerably reduced in structural complexity (1).Individual NPC components have been accessed in FEISEM by a variety of methods, including proteolysis,

The Electron Microscope Unit at the University has a good selection of microscopes: two tungsten SEMs, two FESEMs, a microprobe analysis system, a 125 kY TEM, a 200 kV TEM, an AFM, an FIB miller, a Zeiss Photomicroscope and a Leica Macroscope. We service around 300 clients’ projects a year in every field of experimental science and engineering, logging over 8,000 hrs of beam time annually.But funding constraints have always kept us short staffed and our laboratory has been working towards complete digital image capture for the past ten years to enhance our productivity. The perceived benefits of digitisation for us are:Photographic processing of negatives eliminated.Archiving of (bulky) photographic negatives eliminated.Need for special darkroom & graphics skills eliminatedResponsibility for archiving and indexing images devolved to individual usersResponsibility for image processing devolved to individual users.Rapid turnaround of images.Rapid sharing of results.

X-ray absorption spectroscopy (XANES and EXAFS) applied with x-ray microprobe instrumentation (figures 1) can be used for studying the electronic structure of specific elements in complex materials in a spatially-resolved manner (figure 2). Such techniques are valuable in a wide range of studies including hydrothermal fluid processes, migration and encapsulation of toxic and radioactive wastes, and redox evolution of solar system bodies.One of the major technical challenges in this work is the production of high flux microbeams from high power, hard x-ray synchrotron sources. Some of the microbeam technologies under development include tapered glass capillaries, zone plates, and elliptical mirrors. Each approach has advantages and disadvantages and the optimum microfocusing device depends on the particular experiment.One of the most versatile of these devices is the dynamically bent, elliptical mirror, especially when a pair of mirrors are arranged in a Kirkpatrick-Baez (KB) geometry to provide two-dimensional focusing. This versatility derives mainly from four attributes.

Nucleic acid polymerase structure has been studied by both X-ray and electron crystallography. To date, only the smaller, single subunit polymerases have been subjected to X-ray analysis, including the bacteriophage T7 RNA polymerase, which is the only RNA polymerase whose structure is known to atomic resolution. Lower resolution structures of several multisubunit polymerases have been determined by electron crystallography, including a mutant form of yeast RNA polymerase II which lacks subunits Rpb4 and Rpb7 (denoted A4/7 polymerase). All polymerase structures obtained by both X-ray and electron crystallography show a large cleft appropriate in size for binding duplex DNA, and further appear to contain a mobile arm allowing open and closed conformations of the cleft, presumably permitting entry and retention of DNA. Subunits Rpb4 and Rpb7 of RNA polymerase II form a dissociable subcomplex that has been implicated in the stress response and in the initiation of transcription. Human homologs of Rpb4 and Rpb7 have been identified.

Scanning Probe Microscopy (SPM) is being developed as a possible solution to the problems inherent with analyzing the nanometer scale electronic properties of ULSI integrated circuits. Scanning Kelvin Probe Microscopy (SKPM) and Scanning Capacitance Microscopy (SCM) are both being developed to provide two dimensional dopant profiles of semiconductor devices. SKPM can also determine surface potentials, work functions, dielectric properties, and capacitance.SKPM is based on the concept of Kelvin probe oscillating capacitor work function measurements. The small capacitance area of the SKPM tip and the high resistance of the system produce difficulties in monitoring and minimizing the current in the system. SKPM solves this problem by utilizing the force monitoring capability of the SPM to minimize the Kelvin force instead of the current. An AC voltage applied to the cantilever produces a DC force and AC forces at the AC frequency and the first harmonic of the AC frequency.

The intermetallic, stoichiometric compound NiAl is a candidate for aggressive environments such as those encountered in aircraft engines due to its excellent oxidation resistance and high melting point. However, NiAl is limited in application by its insufficient ambient temperature ductility. This may potentially be overcome via synthesis of NiAl with a nanocrystalline microstructure. Since microstructure can also affect oxidation resistance [1], the focus of this investigation is to examine the oxidation characteristics of nanocrystalline NiAl.Nanophased NiAl specimens were prepared by mechanical alloying followed by shock wave consolidation, which yielded consolidated material with an average grain size of 11 ±8 nm [2]. Thermogravimetric analysis (TGA) of nanosized NiAl was used to characterize the material's oxidation resistance, and performed for three different times (0.5 h, 5 h, 48 h) in compressed air at 1373 K. It was found that the thickness of the scale increased with increasing oxidation time, in accordance with parabolic oxidation kinetics.

Meteorites are remnants of the primordial material from which the solar system condensed. Most meteorites originated in the asteroid belt between Mars and Jupiter and fell to earth when their orbits were disturbed by collisions. Metal phases are present in all types of meteorites and are alloys of Fe and Ni containing S and P. The study of metal meteorites has yielded valuable information about the early thermal history of the solar system, since their heat treatment has been preserved in the microstructure and microchemistry of the meteorites and can be discerned by electron microscopy and microanalysis. A full understanding of the structure and chemistry of meteorites requires detailed knowledge of the Fe-Ni, Fe-Ni-S and Fe-Ni-P phase diagrams and determination of these diagrams has been carried out over more than three decades of electron-beam analysis by the authors.

The evaluation of the two dimensional projected atom column positions around a defect or an interface in an electronic ceramic, as it has been performed in numerous examples by (quantitative) conventional high-resolution electron microscopy (HRTEM), is often not sufficient to relate the electronic properties of the material to the structure of the defect. Information about point defects (vacancies, impurity atoms), and chemistry or bonding changes associated with the defect or interface is also required. Such complete characterization is a necessity for atomic scale interfacial or defect engineering to be attained.One instructive example where more than an image is required to understand the structure property relationships, is that of grain boundaries in Fe-doped SrTi03. Here, the different formation energies of point defects cause a charged barrier at the boundary, and a compensating space charge region around it. The sign and magnitude of the barrier depend very sensitively on the atomic scale composition and chemistry of the boundary plane.

Electrical phenomena such as positive temperature coefficient of resistance (PTCR), grain boundary layer capacitance, and varistors are controlled by Schottky barriers to current transport formed in grain boundary regions. These grain boundary potential barriers are a result of charged defect accumulation, either through equilibrium segregation or kinetic diffusion processes. Recent work on Nb and Mn doped BaTiO3 highlights the importance of substitutional defects which activate grain boundary trap states and give rise to the PTCR effect. The dynamic interaction of charge carriers with such trap states also produces the rapid electrical breakdown characteristic of varistors. This study explores the issues of defect species, space charge, and electrical transport in donor doped SrTi03 by electron holography and electron energy loss spectroscopy.The specific defect species and mechanisms of current transport through donor doped SrTiO3remain unresolved because of the difficulty in connecting defect distributions (structural information) and resultant Schottky barrier effects (chemical information).

Electron microscope tomography is a useful method for deriving three-dimensional (3-D) structures from tilt series. We used an intermediate, high-voltage electron microscope operated at 400 kV to study the 3-D structure of selectively stained or labeled mitochondria from diverse sources. We have examined the structure of neuronal mitochondria and mitochondria from brown adipose tissue (BAT) employing either single-axis or double-axis techniques over a tilt range of ±60° with 1° or 2° angular increments (Fig. 1). Although BAT mitochondria contains only lamellar cristae with no tubular cristae as we have observed in neuronal mitochondria, we found that the inner membrane of BAT mitochondria invaginates to form cristae onlythrough narrow, tubular openings or crista junctions(Fig. 2).Recent interest in the possibility of structural artifacts in chemically fixed mitochondria imaged in situmotivated us to perform electron tomography on cryofixed brain and brown fat tissue.

The Center for X-ray Optics (CXRO) built and operates a high-resolution soft x-ray microscope (XM-1) at the Advanced Light Source in Berkeley. We report on the use of this instrument in a variety of scientific fields, including biology, civil engineering and environmental sciences.The microscope is a conventional (full field) x-ray microscope, which uses zone plate lenses to provide high resolution transmission images. The optical setup is similar to the Göttingen x-ray microscope, operated at the BESSY synchrotron radiation facility in Berlin, Germany. A condenser zone plate, fabricated by the Göttingen group, is illuminating the sample and an objective zone plate, fabricated by Erik Anderson (CXRO), is forming an enlarged image on an x-ray CCD camera. While the optical path of the microscope is in vacuum, the sample is at atmospheric pressure, flushed by helium. The spatial resolution of our microscope is 43 nm, measured as the distance from 10%-90% intensity in the image of a knife-edge.

The Need for Improvements.Though atom probe techniques have impressive atomic-scale analytical capabilities, there are some definite needs for improvement. The needle shape of APFIM specimens is a difficult geometry to make for many types of materials, especially technologically important thin-film materials. Even though 3DAPs collect more data than traditional APs, the analysis volumes are still small by analytical standards: typically there are fewer than 1 million atoms in an image which is about 15nm x 15nm x 50nm. The data collection rate (up to tens of atoms per second) limits the practical size of analyzed volumes to about 1 million atoms. Poor mass resolution makes it difficult to separate ions of similar mass-to-charge ratio in spectra of approximately 20% of samples. Thus, there are three principal areas where improvements are desirable: 1) specimen geometry, 2) data collection rates, and 3) mass resolving power.

The ESEM is able to image liquids in their natural state, unlike conventional SEM, and offers significantly better resolution than optical microscopy. The ability to image binary liquid mixtures, such as water-oil emulsions, depends upon sufficient contrast for differentiation of the dispersed and continuous phases. We have made observations of such water-oil emulsions in an Electroscan 2010 Environmental SEM, and have duly obtained images with good contrast. Experiments were carried out on emulsions of vegetable oil-in-water, polybutene-in-water and water-in-polybutene, all emulsified with nonionic polysaccharide surfactants. Each sample was imaged at a temperature of approximately 3°C, in water vapour at a pressure of 4 torr, in order to maintain the liquid state.From the ESEM micrographs in Figures 1, 2 and 3, it is clear that water exhibits a greater secondary electron emission than an unsaturated hydrocarbon.

The small size of many parasitic organisms requires the use of election microscopy for adequate elucidation of their structure. While both transmission and scanning electron microscopy can provide complementary results which allow considerable degree of structural correlation, each technique has its inherent limitations. Since previous studies have demonstrated that soft X-ray microscopy could be used to study parasitic protozoa and provide new information, the objective of this study was to determine whether soft X-ray microscopy could also be used to elucidate the morphology of small metazoa to complement the data obtained by other microscopy techniques.Newborn larvae, approximately 7 μm × 110 μm in size, of parasitic nematode Trichinella spiraliswere used as a model system. Some of the larvae, deposited by adult females maintained in vitro,were isolated and processed for examination by transmission and scanning electron microscopy as described in our previous studies, others were fixed in 4% glutaraldehyde (Millonig's buffer) and examined in the X-ray microscope XM-1, and in the BioRad MRC 1024 confocal laser (krypton/argon) microscope of the Advanced Light Source, Berkeley National Laboratory.

Atomic force microscopy (AFM) provides unique opportunities to study cell-surface and molecular scale interactions in three dimensions under aqueous conditions. Two plasma proteins, von Willebrand Factor (vWf) and fibrinogen, play central roles in the regulation of hemostasis and thrombosis by participating in coagulation and facilitating adhesion and aggregation of activated platelets. vWf and fibrinogen are believed to facilitate platelet adhesion under regions of relatively high and low vascular wall shear stress, respectively. Consequently, elucidating vWf and fibrinogen structure-function relations under shear is of considerable importance in developing a comprehensive understanding of the pathophysiology of thrombogenesis. Previously, we reported on molecular level AFM images of human vWf during shear-induced structural transition and human fibrinogen under aqueous conditions when both proteins were adsorbed on a hydrophobic surface. This presentation will report on the shear dependent interactions of dimeric and multimeric vWf with a hydrophobic surface and the substrate-dependent interactions of fibrinogen.

Compositional mapping with energy dispersive x-ray spectrometry (EDS) in the scanning electron microscope (SEM) is one of the most widely used techniques for elemental imaging of microstructures. SEM/EDS maps are usually qualitative presentations of elemental distributions in x-y space. In older implementations of mapping, the x-ray intensity from a single spectral region of interest (ROI) above a threshold was recorded on film as a white “dot” during analog scanning. With modem digital control and data recording, x-ray intensities from many ROIs can be recorded in parallel and true counts stored in memory. For display purposes a gray level or color value is used to represent the stored x-ray intensity at each pixel. For major constituents (arbitrarily, major >10%), it is generally possible to make valid, robust comparisons between regions within an individual compositional map, but comparisons may be problematic among several maps of different constituents within the same field of view, depending on the sophistication of the mapping software.

Modern high-performance computers are now capable of calculations that can be used to determine the preferred atomic arrangements in complex systems. Both first-principles and semiempirical approaches have been developed with complementary capabilities. At the same time, powerful transmission electron microscopes have been developed that yield direct atomic-scale images of crystals and extended defects such as dislocations, grain boundaries and buried interfaces. This paper presents several examples where a synergistic approach combining theoretical results, Z-contrast scanning transmission electron microscopy, and spatially resolved electron energy loss spectroscopy (EELS) have led to the elucidation of complex atomic structures. In some cases, theory predicts, experiment confirms and expands and theory revisits; in other cases, observations come first and theory helps put together a comprehensive picture or goes beyond the original observations with new predictions. In both cases, equilibrium structures, impurity or stressinduced structural transformations, and dynamical processes such as diffusion, segregation or precipitation are elucidated in great detail.

Progress towards the large scale integration of active devices within electronics packages has imposed stringent heat dissipation requirements necessitating the development of innovative materials solutions. One possibility being considered is the use of chemically vapor deposited diamond (CVDD) thin films as heat sinks. However, there are technological challenges which must be overcome before these materials become commercially viable. For example, the CVDD substrate must be metallized to provide interconnections between the various devices in the package. Conventional metallizations, such as Au, Cu or Al, display poor adhesion to the diamond causing problems associated with the reliability and stability of metallized diamond packages. A novel solution has been proposed involving the consecutive deposition of a thin layer of Cr and an electrically insulating layer of alumina on the diamond substrate such that the thermal conductivity of the treated substrate is not degraded significantly.

Strained epitaxial films are often unstable to morphological transformations. The classic example of this phenomenon is the SiGe system. A nonplanar morphology results in non-uniform strains in the system (Figure 1). These strain fields can modify the local chemical potential of a uniform alloy driving Ge enrichment in the regions of low compression.Recent work has shown that annealed films of SiGe on Si{001} exhibit both vertical and lateral segregation during the transition from a metastable planar film to an array of ﹛100﹜ directed island. Vertical segregation is evidenced by an enrichment of Ge in the last few monolayers of the film while lateral segregation is seen as an enhancement of this surface layer in the region near the island peaks. This segregation is due to surface diffusion as the bulk diffusion lengths are too short. The case of annealed, metastable films can be compared to growing films.

The Materials Microcharacterization Collaboratory (MMC) was created last year as a pilot project within the US Department of Energy's DOE2000 program. The DOE2000 program has three main goals: improved ability to solve DOE's complex scientific problems, increased R&D productivity and efficiency, and enhanced access to DOE resources by R&D partners. One of the strategies to meet these goals is the construction of national collaboratories. These provide integration on the Internet of unique or expensive DOE research facilities and expertise for remote collaboration, experimentation, production, software development, modeling, or measurement. In addition, collaboratories will benefit researchers by providing tools for video conferencing, shared data-viewing, and collaborative analysis. Cooperative pilots projects, jointly funded by DOE2000 and a scientific program area, are expected to lead to clear success in the scientific environment: they are to provide new capabilities and to increase the efficiency of doing the work (e.g., reduce travel, increase communication, provide for sharing of data among formerly disparate environments).

The addition of the Si-Li detector to the electron probe, the scanning electron microscope, and more recently the transmission electron microscope (resulting in the analytical electron microscope) has made it possible to obtain elemental analysis on individual “particles” with dimensions less than 1 nm using EDS. Although some initial particle studies on micrometer-sized particles were done on the electron probe using wavelength dispersive spectrometers, WDS, the variability and complexity of many particle compositions coupled with the high currents necessary for WDS made elemental analysis of particles by WDS difficult at best. In addition, the use of multiple spectrometers, each with a different view of the particle and therefore different particle geometry as shown in Fig. 1, limited the quantitative capabilities of the technique. With the introduction of the Si-Li detector, there was only one spectrometer with a single geometry resulting in the development of various procedures for obtaining quantitative elemental analysis of the individual particles.

The technologies of nanofabrication as applied to inorganic materials and substrates are advanced and continue to develop. These sophisticated processes enable the formation of complex electronic, optical and mechanical devices with feature sizes down to tens of nanometers. Adaptation of these types of processes to surface chemical patterning and topographical patterning provides a new set of experimental tools for investigating biological systems and realizing sensors and devices that require the interaction of biological systems and fluids with inorganic materials and surfaces.In this talk we discuss methods of pattering self assembled monolayers, proteins and antibodies on silicon and glass surfaces by lithography and microcontact printing. This is of utility in a variety of experiments in cell-surface interactions and in sensor devices. In certain types of devices the manipulation of fluids and sieving of molecules is a critical function such as in DNA sequencing6 by electrophoretic separation.

The nuclear envelope (NE) physically separates the genetic machinery residing in the nucleus from protein synthesis taking place in the cytoplasm. Bidirectional transport of proteins, RNAs and RNP particles between these two compartments is mediated by the nuclear pore complexes (NPCs), large (-120 MDa) supramolecular assemblies embedded in the NE and being built of -100 different polypeptides, called nucleoporins. Extensive structural studies have revealed the 3D architecture of NPCs, and epitopes of several nucleoporins have been localized within their 3D structure.In an attempt to go beyond the current consensus model of the NPC (Fig. lc, inset), we have embarked on a more systematic structural analysis of the molecular architecture of native NPCs. This is achieved by field-emission (FETEM; Fig. la) or energy-filtering (EFTEM; Fig. lb) TEM of completely unfixed and unstained Xenopusoocyte NEs embedded in thick (i.e. -200 nm) amorphous ice so that the 3D organization of the cytoplasmic and nuclear periphery of the NPCs (i.e. the cytoplasmic fibrils and nuclear baskets) is fully preserved (Fig. lb).

Introduction:This is a personal account of the author's attempts to solve some of the problems associated with the fixation and embedding of difficult biological materials such as plant tissues, almost all insect tissues, and large pieces of tissues including whole organs.Historically, the search began in the 1950's with the study of the maize root tip using the relatively new technique of electron microscopy. This system of cells, separated by thick and partially impermeable walls, did not respond well to the procedures most suitable for animal tissues. New approaches were needed and, in time, some were found that were at least useful if not ideal. A brief account of several of these is given in the following paragraphs.Fixation of plant cells:Potassium permanganate was chosen very early in the study primarily because it worked better for plant cells than did any of the fixation procedures currently available.

Light microscopic imaging has experienced a renaissance in the past decade or so, as new techniques for high resolution 3D light microscopy have become readily available. Light microscopic (LM) analysis of cellular details is desirable in many cases because of the flexibility of staining protocols, the ease of specimen preparation and the relatively large sample size that can be obtained compared to electron microscopic (EM) analysis. Despite these advantages, many light microscopic investigations require additional analysis at the electron microscopic level to resolve fine structural features.High voltage electron microscopy allows the use of relatively thick sections compared to conventional EM and provides the basis for excellent new methods to bridge the gap between microanatomical details revealed by LM and EM methods. When combined with electron tomography, investigators can derive accurate 3D data from these thicker specimens. Through the use of correlated light and electron microscopy, 3D reconstructions of large cellular or subcellular structures can be obtained with the confocal microscope,

Ex-situ and in-situ studies were performed for self-assembled monolayers (SAMs) formation on mica. The behavior of two surfactants [octadecylphosphonic acid (OPA) and dodecylphosphonic acid (DPA)] in two different solutions [ethanol or tetrahydrofuran (THF)] were studied at different concentrations. A cross section analysis of the monolayer images at partial coverage showed thickness of 1.8 nm, 0.7 nm, and 1.0 nm for OPA, DPA and mixed OPA/DPA, respectively. These values are similar to the ones presented by Xiao and those expected for a monolayer of mixed surfactants as proposed by Whitesides. Since the topography of a monolayer resembles the substrate, the presence of the monolayer at full coverage was determinated by force-distance curve measurements. It was observed that the solvent had the ability to re-dissolve the OPA monolayer, suggesting that the bonding is of a physisorption nature.

The cytoskeleton plays an important role in cell structure, polarity, locomotion and division. Individual elements of the cytoskeleton are composed of subunit proteins which assemble and disassemble in specific places and times within the cell. Knowledge of the temporal and spatial regulation of subunit assembly and disassembly is essential to understanding how the cytoskeleton contributes to cellular activities. The assembly and dynamics of two cytoskeletal structures, namely adhesion plaques (APs) and intermediate filaments (IFs), have been difficult to study by traditional methods. We have generated GFP-chimeras to label these structures and to study their dynamics in motile fibroblasts.To study the dynamics of APs, we prepared stable 3T3 cell lines expressing a GFP-β1 integrin chimera. The chimera was prepared by fusing GFP to the transmembrane and cytoplasmic portions of β1 intergrin, since previous studies had shown that the cytoplasmic tail of β integrins is sufficient to direct integrins to APs.

In many cases in experimental science, it is true that the instrument interface becomes a limiting factor in the efficacy of carrying out unusual experiments, or prevents the complete understanding of the acquired data. We are developing an advanced interface for Scanning Probe Microscopy (SPM) which allows intuitive rendering of datasets and natural instrument control, all in real-time. The interface, called the nanoManipulator, combines a high performance graphics engine for real-time data rendering with a haptic interface which places the human operator directly into the “feedback loop” that controls surface manipulations. In practice, the user holds a stylus in hand. By moving the stylus laterally, the user directs the movement of the SPM tip across the sample. The haptic interface enables the user to “feel” the surface by forcing the stylus to move up and down in response to the surface topography.

Due to environmental concerns, there is a trend to avoid the use of chemicals needed to develop negatives and to process photographic paper, and to use digital technologies instead. Digital technology also offers the advantages that it is convenient, as it enables quick access to the endresult, allows image storage and processing on computer, allows rapid hard copy output, and simplifies electronic publishing. Recently significant improvements have been made to the performance and cost of camera-sensors and printers. In this paper, field ion images recorded with two digital cameras of different resolution are compared to images recorded on standard 35 mm negative film. It should be noted that field ion images exhibit low light intensity and high contrast.Field ion images were recorded from a standard microchannel plate and a phosphor screen and had acceptance angles of ∼60°.

Sea urchins have been used for over a century as a remarkable animal model system in which to study cell, molecular, and developmental biology. The studies presented here have used sea urchin eggs and embryos for pioneering experiments to explore the effects of microgravity on the cytoskeleton during a space flight on the space shuttle Endeavor. The culture conditions followed those described previously utilizing the Aquatic Research Facility (ARF) to fertilize and culture eggs and embryos up to the pluteus stage under controlled temperature (12°C) and fixation conditions. To achieve a final fixation with 0.5% glutaraldehyde and 4μM taxol, concentrated fixation fluid was injected at preselected time points to preserve microtubules, centrioles, centrosomes, microfilaments, mitochondria, and cell membranes.The analysis of the results revealed that the centriole-centrosome complex during cell division and cilia formation showed alterations in samples that had been exposed to microgravity while control cells cultured in a centrifuge at lg in space and those cultured on ground appeared normal.

A thorough microstructural study of common ceramics requires analysis at various length scales. Yttrium doped, polycrystalline α-Al203 was studied using a broad selection of microscopy techniques over a scale range covering seven-orders of magnitude (i.e. from mm to Å).Yttrium doped, (1000 (Y/Al) atomic ppm) polycrystalline alumina samples were prepared by hot pressing at 1450°C at 50 MPa. Some were subsequently crept in tension. Microstructure of these samples was studied by optical microscopy, scanning electron/orientation imaging microscopy (OIM™), conventional and high resolution transmission electron microscopy. Analytical electron microscopy was performed on a VG HB501 FEG-STEM equipped with an EDS and PEELS.On a large scale, the microstructure often consisted of bimodal sized alumina grains and second phase YAG (Y3A15012) precipitates. YAG precipitates at grain boundaries (GB) and triple points caused clusters (50μm to 1mm) of small equi-axed alumina grains (0.5-2 μm) randomly distributed over 40 area% of the sample.

The focused ion beam (FIB) has become an indispensable tool for a variety of applications in materials science, including that of specimen preparation for the transmission electron microscope (TEM). Several FIB specimen preparation techniques have been developed, but some problems result when FIB specimens are analyzed in the TEM. One of these is X-ray fluorescence from bulk material surrounding the thin membrane in FIB-prepared samples. This paper reports on a new FIB specimen preparation method which was devised for the reduction of X-ray fluorescence during energy dispersive X-ray spectroscopy (EDS) in the TEM.Figure 1 shows three membrane geometries that were investigated in this study on a single-crystal Si substrate with a RF sputter-deposited 50 nm Ni film. Membrane 1 is the most commonly reported geometry in the literature, with an approximately 20 urn wide trench and a membrane having a single wedge with a 1.5° incline.

HK97 is a tailed temperate bacteriophage of E. coli that builds an icosahedral capsid using steps that include regulated assembly, proteolysis, radical conformational changes and the formation of novel covalent bonds (Fig. 1). This pathway is being exploited as a model system to explore how the formation of multiprotein complexes can be regulated by each of these mechanisms. We have identified and purified at least four intermediates (Prohead I, Prohead II, Head I and Head II) and examined them by cryo-electron microscopy and three dimensional reconstruction procedures (Fig. 2). Comparison of particle reconstructions at resolution of about 25 - 30 A have lead to major insights into the causes and purposes of the regulated changes that we have also characterized biochemically and genetically.Prohead I consists of 420 copies of the 42 kDa gp5 capsid protein arranged as 72 blister-shaped morphological capsomers in a thick walled hollow T=7 icosahedral particle with a diameter of -470 Å.