Extended abstract of a paper presented at Microscopy and Microanalysis 2008 in Albuquerque, New Mexico, USA, August 3 – August 7, 2008

A variety of porous materials are used in a number of tissue-engineering applications such as cell transplantation and drug delivery. They provide a temporary scaffolding for a large cell mass, however they must have a fully interconnected pore structure. One such material is sodium alginate, a naturally derived polysaccharide which is isolated from seaweed. Sodium alginate can de dissolved in water to form a highly viscous solution that can be cross-linked in the presence of divalent cations (e.g., Ca2+) to form what is known a as hydrogels. We have developed a method for the fabrication of porous beads of sodium alginate and have attempted to measure the pore size, density and distribution in the beads. Characterization of the pore size and distribution is necessary to determine if the material is appropriate to promote cellular in-growth. The characterization procedures have been limited to conventional scanning electron microscopy (SEM) and mercury intrusion porosimetry. There has been some concern as to the potential introduction of artifacts (spatial distortion) during dehydration and sample preparation during these characterizations.

Extended abstract of a paper presented at Microscopy and Microanalysis 2008 in Albuquerque, New Mexico, USA, August 3 – August 7, 2008

Extended junctional sarcoplasmic reticulum (EJSR) is an invariant differentiation of the sarcoplasmic reticulum (SR) in bird cardiac myocytes (CM) and central to excitation-contraction coupling (ECC). EJSR occurs as both continuous and discontinuous extensions of junctional sarcoplasmic reticulum (JSR), and surrounds and pervades the Z/I band as the “ EJSR Z-rete” whose geometry has mechanistic implications for the function of “couplings” in ECC, in general. “Peripheral coupling(s)” (PC) in birds, and the additional “interior coupling(s)” (IC) at transverse tubules (TT) in mammals, are formed by tight apposition to plasmalemma of JSR, a specialized calcium (Ca) store of the SR. Free SR (FSR; i.e. free of JSR/EJSR specializations) is the rest of the smooth, tubular SR network, which connects intercalated patches of EJSR forming the EJSR Z-retes and, elsewhere, displays both longitudinal and transverse geometries in surrounding the contractile material for the purpose of sequestering Ca after each muscle contraction. Except for EJSR having no plasmalemmal contact, morphologically, EJSR and JSR are homologues:1 both have similar sizes; are studded (approx. 32 nm center-to-center) with junctional processes (JP; ryanodine receptor (RyR)/-Ca-release channels);

Analytical transmission electron microscopy (TEM) and scanning electron microscopy (SEM) have been applied for the characterization of evolution, lateral arrangements, orientations, and the microscopic nature of nanostructures formed during the early stages of ultrahigh vacuum electron beam evaporation of Cu onto surfaces of VSe2 layered crystals. Linear nanostructure of relatively large lateral dimension (100–500 nm) and networks of smaller nanostructures (lateral dimension: 15–30 nm; mesh sizes: 500–2000 nm) are subsequently formed on the substrate surfaces. Both types of nanostructures are not Cu nanowires but are composed of two strands of crystalline substrate material elevating above the substrate surface. For the large nanostructures a symmetric roof structure with an inclination angle of ∼30° with respect to the substrate surface could be deduced from detailed diffraction contrast experiments. In addition to the nanostructure networks a thin layer of a Cu-VSe2 intercalation phase of 3R polytype is observed at the substrate surface. A dense network of interface dislocations indicates that the phase formation is accompanied by in-plane strain. We present a model that explains the formation of large and small nanostructures as consequences of compressive layer strains that are relaxed by the formation of rooflike nanostructures, finally evolving into the observed networks with increasing deposition time. The dominating contributions to the compressive layer strains are considered to be an electronic charge transfer from the Cu adsorbate to the substrate and the formation of a Cu-VSe2 intercalation compound in a thin surface layer.

Quantitative analysis of a structure means to determine which atoms are where. The where is restricted by the lateral resolution, and the which is related to the sensitivity against differences of the atomic species. Electron holography is on the move to tackle both questions.

• where - Lateral Resolution Electron holography has surmounted conventional electron microscopy in that it has reached atomic resolution both in the amplitude and phase images: with our CM30FEG/UT Special Tuebingen electron microscope, the information limit of 0.09nm and the pixel number of the 2048_ - CCD camera allow to take holograms with image details smaller than 0.095nm; the quality of correction of the aberrations allows an interpretable resolution approaching 0. lnm both in amplitude and phase [1].

• which - Phase Detection Limit (PDL) The phase detection limit describes the minimum phase difference detectable well above a given noise level.

The environmental scanning electron microscope (ESEM) first marketed by ElectroScan and now by FEI/Philips has been recognized as the only scanning electron microscope which allows liquid water to be present in the sample chamber at temperatures above freezing. As most biological material contains a high percentage of water it would seem essential to use this technology to examine specimens in their natural state. The purpose of this presentation is to describe in detail the way in which the microscope should be used to maximize the information obtainable from a variety of biological samples. This tutorial will also consider methods which can be applied to “difficult” samples whose structure becomes altered during what would be considered a normal imaging schedule.

The main features of the microscope are the use of the Peltier effect cooling stage and control of chamber water vapor pressure to enable wet specimens to be imaged.

Extended abstract of a paper presented at Microscopy and Microanalysis 2006 in Chicago, Illinois, USA, July 30 – August 3, 2006

Extended abstract of a paper presented at Microscopy and Microanalysis 2006 in Chicago, Illinois, USA, July 30 – August 3, 2005

Human α2 macroglobulin (α2M) has structural and functional properties that contribute to its uniqueness as proteinase inhibitor. It is the largest known (Mr=720,000) and the only natural proteinase inhibitor which has a broad range of reactivity and for which the reaction is irreversible. It has a vital role in the clearance of proteinases from the circulation and in regulating their activity in fibrinolysis, coagulation and complement activation.

An α2M molecule can entrap two proteinase molecules such as chymotrypsin and trypsin and can therefore be considered to contain two functional domains. Each subunit in the homotetramer has a bait region with cleavage sites for nearly all known endoproteinases and an internal thiol ester bond. A proteinase cleaves the two bait regions within both functional units leading to an activation and cleavage of the thiol ester bonds. Consequently, α2M undergoes a major structural change resulting in the irreversible entrapment of the proteinase.

Interest in electron beam x-ray microanalysis with low incident beam energies, defined arbitrarily as 5 keV and below, has been greatly stimulated in recent years by the development of the high performance field emission gun scanning electron microscope (FEG-SEM), which can produce a nanometer-scale probe with sufficient current to operate with both energy dispersive (EDS) and wavelength dispersive (WDS) spectrometers. Microanalysis in this regime requires the analyst to confront new spectrometry problems that are not typically encountered, or that can be safely ignored, when operating with conventional beam energies, 10 keV or greater. With low energy operation, the choice of atomic shells that can be accessed is restricted, forcing the analyst to make use of shells that have low fluorescence yields for intermediate and high atomic number elements, and possibly strong chemical effects, which are evident with high resolution x-ray spectrometry.

In 1992, researchers at Mobil Research and Development created a new class of porous silicates, most notably mesoporous MCM-41.2-3 This material features a hexagonal arrangement of linear pores and surface areas in excess of 1000 m2 g-1. MCM-41 exhibits narrow pore size distributions in the nanometer range. The walls of MCM-41 are essentially amorphous silica, but its porous nature makes it about 3 kJ mol-1 less stable than the collapsed form. Particles of MCM-41 are beam sensitive and it appears that they cannot withstand the large current densities required to obtain reliable analytical data at the nanometer scale in the transmission electron microscope (TEM). For example, low beam currents were used to preserve the pore structure but resulted in energy-filtered TEM elemental maps of oxygen K and Ti L2,3 edge intensities that were too noisy to reveal structure at 5 nm resolution.

In flowering plants, pollination and the process of fertilization are complex processes involving a series of cell-to-cell communication events. Though details of the progression of the pollen tube through the pistil, beginning with germination of the pollen grain on the stigma and culminating with delivery of the sperm cells to the embryo sac, are well established for several higher plant species, the mechanisms involved have yet to be elucidated. It has been shown that the transmitting tissue, which coincides with the path of pollen tubes in the gynoecium, is composed of highly secretory cells characterized by an extensive extracellular matrix (ECM). The actual roles that this ECM plays in pollination are currently unknown, although functions proposed include mechanical and/or chemotropic pollen tube guidance as well as pollen tube nutrition.

Precipitation-hardened stainless steels are important structural materials in a variety of industrial applications in aircraft, chemical and power plants. These steels have a combination of good mechanical properties, corrosion resistance and simple fabrication process. Precipitation reaction in a PH 17-4 stainless steel which contains approximately 3 wt.% of Cu begins with formation of coherent copper-rich precipitate, which subsequently transform to incoherent fee Cu by further aging. By prolonged aging at 400 °C, embrittlement occurs as hardening progresses. Both the coarsening of Cu and the decomposition of the martensite matrix could be the reasons for this embrittlement. Thus, this study attempted to clarify the phase separation and the precipitation processes in a PH 17-4 stainless steel on prolonged aging by atom probe field ion microscopy (APFIM), in order to understand the mechanism of the degradation of mechanical properties by long term medium temperature aging.

Each field of electron microprobe analysis has its own specific analytical difficulties. Problems relating to the analysis of geologic materials can be primarily attributed to the fact that the samples are nonconductive, some are unstable under the beam, and they have a complex matrix. There are various techniques to solve these analytical problems, but the selection of appropriate standards is one of most important factors.

In study of geologic materials a convention prevails that the summation approximates 100%. For most analyses summation is an indication of the quality of analysis, although some elements could be over or under corrected. Significant progress has been made in developing correction procedures, yet some fundamental parameters still are not well known, and the results will vary depending on which procedures or fundamental parameters are used. One of the most practical solutions to eliminate significant matrix effects is to select reference materials with a close composition to the unknown.

A knowledge of grain boundaries (GBs) and interfaces is essential in establishing the relationship between the structural details of the boundaries and the strength of superconducting coupling and critical current densities at the boundaries in textured Bi cuprates. As a step towards such understanding, we studied GB crystallography and lattice mismatch using the constrained Coincidence-Site-Lattice (cCSL) theory. We note that to characterize arbitrary GBs in a complex system such as Bi2212, it would be desirable to find some simple analytical solutions for the cCSL approach.

We report our new concept of a “two-mirrors” model (2M-model), which allow us, in principle, to find sufficient conditions to generate corresponding analytical solutions for CSL (or cCSL), when it is applied to crystals with cubic, hexagonal, or orthorhombic symmetry. The 2M-model is based on the fact that any boundary can be described by rotating one crystal relative to the other along a common axis.

Sample preparation for transmission electron microscopy (TEM) has been a source of speculation with regards to potential for the creation of artifacts which may confound data gleaned from TEM analysis. For semiconductor integrated circuit (IC) materials characterization, the most common sample preparatory methods are based on final thinning by ion beam milling. The latest shift towards Copper / low dielectric constant (k) composite systems in the semiconductor IC industry provides several challenges for TEM sample preparation resulting from differences in milling rates and materials properties for neighboring features.

In conjunction with process development for integration of Cu / low-k materials, conducted at SEMATECH, we have systematically studied the effects of TEM sample preparation by ion milling in order to search for artifacts that could result from sample thinning procedures. For this purpose we have studied wafers with patterned copper lines isolated by a low-k polymer. One sample was stressed by thermal and electronic bias, while a second was subjected to only thermal stress.

One of the most challenging tasks for high-resolution electron microscopy (HREM) is the atomistic investigation of defects and interfaces in thin films of semiconductors and ceramics. In particular, the application of these materials in electronic devices requires the understanding of the atomistic structure responsible for electronic and optical properties. The imaging of these structures requires a point resolution down to 0.1 nm. For example the smallest atomic spacing of GaAs in the (110) projection is 0.14 nm. According to Scherzer the point resolution of the TEM is proportional to spherical-aberration coefficient Cs1/4 and to the wave length λ3/4. Commercial medium-voltage microscopes up to 400 kV offer only a point resolution of 0.16 nm due to the high spherical-aberration constant Cs of the electromagnetic objective lens. Decreasing the wave length λ by increasing the electron energy improves the point resolution, but has the drawback of severe radiation damage and high costs.