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.

Amorphous chalcogenides films coated with certain metals are known to possess a remarkable sensitivity to radiation. Based on these effects they have found several important applications in the production of microcircuits, e.g. as resists in photo- and electron beam lithography. A variety of chalcogenides in combination with a range of metals has been extensively investigated with regard to potential applications in the fabrication of semiconductor devices.

The objectives of the present project are to extend knowledge of the behaviour of amorphous chalcogenides in contact with metals. This includes studying the metal diffusion process, understanding the film-electron beam interaction process and evaluating the potential of the metal lines formed by the electron beam in fabrication of photomasks for the production of higher densities of silicon microcircuits.

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.

Dihydrohpoamide acetyl transferase (E2), a catalytic and structural component of a multienzyme complex that catalyzes the oxidative decarboxylation of pyruvate, forms the central core to which the other components are bound. We have utilized protein engineering and 3-D electron microscopy to study the structural organization of the largest multienzyme complex known (Mr ∼ 107). The structures of the truncated 60-mer core (tE2) and complexes of the tE2 associated with a binding protein (BP), and the BP associated with its dihydrohpoamide dehydrogenase (BP'E3) and the intact E2 associated with BP and the pyruvate dehydrogenase (E1) were determined (Figs. 1 and 2). The tE2 core is a pentagonal dodecahedron consisting of 20 cone-shaped trimers interconnected by 30 bridges.

Previous studies have given rise to the generally accepted belief that BP and BP'E3 components are bound on the outside of the E2 scaffold and that E1 is similarly bound to the core in variable positions by flexible tethers.

This report summarizes our experiences gained with deconvolving 3-d images of various types of biological samples - animal and plant cells; live and fixed. We compared raw, unmodified images with images processed with standard filtering methods and images improved by using deconvolution. The blind decovolution software package from AutoQuant, Inc. was used. All images were acquired using a BioRad MRC 1024 confocal microscope (attached to a Nikon Diaphot microscope), using fluorescence or reflected light. For thin (up to 25 μm) flat specimens a 60x PlanApo NA1.4 lens was used while for thick (25-200 μm) water containing samples a 40x Fluor NA1.15 water immersion lens was used.

In an attempt to test the usefulness and validity of deconvolution for various types of biological samples we adopted the following approaches:

1. collect images of structures that are known to be both fluorescent and reflective - compare the results of deconvolution of fluorescence images with reflected light images that are not deconvolved.

We are developing a new family of heterogeneous catalysts for hydrogenation catalysis. Catalyst synthesis is accomplished using colloidal polymerization chemistry which produce high surface area xerogel catalysts. These xerogels have been synthesized by one-step sol gel chemistry. These catalysts contain ruthenium and modifiers such as gold occluded or incorporated in a titanium oxide matrix. The materials, especially the modified systems exhibit favorable performance in microreactor evaluations for hydrogenation reactions and exhibit high activities. Nanostructural studies have revealed that the materials contain dispersed catalyst clusters which are desirable microstructures for the catalysis since the majority of the atoms are exposed to catalysis and are potentially active sites.

The composition and atomic structure of the xerogel catalysts containing ruthenium and other metals have been examined using our in-house developments of environmental high resolution electron microscopy (EHREM) the atomic scale [1-3] and low voltage high resolution SEM (LVSEM)[4] methods.

Poly-[(trans-1,2-diaminocyclohexane) platinumj-carboxyamylose (“poly-plat”), 5-sulfosalicylato-trans -(1,2-diaminocyclohexane) platinum (SSP), and 4-hydroxy-∝-sulfonylphenylacetato (trans 1,2-diaminocyclohexane) platinum (II) (SAP) are second generation analogs of cisplatin (CDDP) with higher efficacy and potency than cisplatin. This is particularly true of “poly-plat” which contains 1/5 the platinum of CDDP. In order to understand the mechanism of action of these compounds, isolated murine peritoneal macrophages in culture medium were treated with “poly-plat”, SSP, or SAP (5 μg/ml) for 2 h. Drug containing medium was then replaced with fresh medium and the cells were allowed to incubate at 37° C (5% CO2) for 24 h. Supernatants were collected at 0.5, 1, 2, and 24 h post-treatment for immunocytochemical analysis. Confocal microscopy studies demonstrated an increase in the number of lysosomes in the treated macrophages, but only “poly-plat” and SSP treated macrophages were stimulated to form cytoplasmic extensions at 2 h and 24 h.

Optical tweezers, or the single-beam optical gradient force trap, is becoming a major tool in biology for noninvasive micromanipulation on an optical microscope. The principles and practical aspects that influence construction are presented in an introductory primer. Quantitative theories are also reviewed but have yet to supplant user calibration. Various biological applications are summarized, including recent quantitative force and displacement measurements. Finally, tantalizing developments for new, nonimaging microscopy techniques based on optical tweezers are included.

A new technique, referred to as rolling-assisted-biaxially-textured-substrates (RABiTS) has recently been proposed by Goyal et al., to fabricate long-range, biaxially textured substrates with suitable metal/ceramic surfaces for epitaxial growth of electronic devices [1]. Using standard thermomechanical processing, long lengths of flexible, biaxially oriented substrates with smooth surfaces (rms∼50nm) are obtained [1]. Epitaxial metal and/or oxide layers which can serve both as a chemical as well as a structural buffer are deposited on the biaxially textured metal. The metal with a suitable set of epitaxial, multi-layers comprises the substrate which is expected to have many potential applications, including superconductivity, photovoltaics and ferroelectrics [1].

When high temperature superconductors are deposited on a set of epitaxial multilayers on biaxially textured RABiT substrates, extremely high critical current densities, approaching 106 A/cm2 at 77K have been obtained [2].

Experimental conditions and setting up of an electron microprobe is directly linked to the assumption or the knowledge that can make the operator on the results of the measurements. Consequently, there is an increasing interest in modelling the X-ray spectrum in order to predict accurately and rapidly the response of the microprobe when a sample of approximately known composition is submitted to specific excitation conditions. Even if the spectra simulation is a current tool proposed in EDS, it always remains a challenge to create a complete absolute WDS spectrum. Indeed, the definition of the efficiency function was a limit to the calculation of absolute intensities.

In this work, we propose a new generation software for EPMA which uses the absolute simulation spectra to assist the operator in the choices of experimental conditions to achieve accurate measurements in a short time as well as to create new functionalities.

A new infrared spectroscopic imaging technique has been described, combining step-scan Fourier transform (FT) Michelson interferometry with indium antimonide (InSb) focal-plane array (FPA) image detection [1-3], for use in the range 3950-1975 cm−1. The coupling of such detector to an interferometer provides an optimized method for infrared spectroscopic imaging by simultaneously realizing both a multiplex and multichannel advantage. Specifically, the multiple detector elements enable spectra at all pixels to be collected simultaneously, while the interferometer allows all the spectral frequencies to be measured concurrently. This technique can rapidly generate very high quality, chemically specific images from a wide variety of samples. Preliminary results from the use of a mid-infrared liquid-helium-cooled arsenic-doped-silicon array have also been reported recently [4]. Focal-plane arrays using mercury-cadmium-telluride (MCT) are now commercially available, giving access to the range 3950-800 cm−1, and greatly broadening the applicability of this technique [5].

FT-IR microscopy does not require exotic sample preparation -- samples can be examined in transmission and reflectance, do not need to be stained or coated, and a vacuum is not required.

Two-photon fluorescence microscopy has become an important research tool in both biological and material sciences. The technique uses long wavelength, typically in the near IR, as the excitation light to obtain shorter wavelength fluorescence (e.g. visible light). Because of the low linear absorption coefficient of most biological and polymeric specimens, this technique allows deeper penetration of the excitation beam, achieving optical sectioning to a depth of 250μm or more into the specimen. As a result of the quadratic dependency of the two-photon induced fluorescence to the excitation intensity, the fluorescent emission and photobleaching are limited to the vicinity of focal spot. This capability of addressing a specimen’s 3D space allows exciting possibilities in biological researches, such as 3D photobleaching recovery experiment.

Two-photon confocal fluorescence microscopy is ideal for the study of thick biological and material specimen in 3D. For example, Figure 1 shows a three-dimensional isosurface rendered image of a vascular bundle from a maize stem.

Rechargeable Lithium ion batteries are widely used as portable power source in communication and computer technology, prospective uses include medical implantable devices and electric vehicles. The safety and cycle life of Li ion batteries is improved over that of batteries containing metallic lithium anodes because the insertion of Li between the crystal layers of both electrodes was proved to be safer than the electroplating of Li onto a metallic Lithium anode. in Li-ion batteries, the charge transport is governed by the oscillation of Li ions between anode and cathode. They are sometimes called “rocking-chair“ batteries. The most common materials for these batteries are lithiated carbons for anodes, and transition metal oxides (LixCoO2) as cathodes.

LixCoO2 has an ordered rhombohedral Rm structure consisting of alternating layers of Co-O-Li-O-Co. The capacity and energy density of the batteries is limited by the amount of Li that can be stored in the anode and cathode materials.

Energy Dispersive Spectrometry (EDS) is an ubiquitous method of elemental analysis for SEM, TEM, and STEM applications. The elements of interest are generally quantified without standards using theoretical calculations or by using standards that are high purity specimens of the elements measured. However, EDS is often used to determine a small percentage of an element in a matrix. The accuracy and limit of detection of these low concentration measurements has not been established. An earlier report proved the concept that a cross section high dose BF2 implanted specimen could provide a standard for EDS measurement of F. This study extends this quantification approach to transition elements of importance to the semiconductor industry.

The Fe and Co standards were created by high dose ion implantation. For ions implanted into silicon, a dose of lxl016 atoms/cm2 results in a peak concentration of approximately lxl021 atoms/cm3 or 2% atomic. The exact concentration can be determined using methods such as Rutherford Backscattering Spectrometry (RBS) and Secondary Ion Mass Spectrometry (SIMS).

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

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 2008 in Albuquerque, New Mexico, USA, August 3 – August 7, 2008

Freeze-fracture techniques have contributed much to our understanding of membrane composition and macromolecular architecture. However, further substantive contributions from freeze fracture have seemed unlikely because: a) replicas typically fragmented, destroying histological orientation within samples; b) histochemical staining techniques were thought to be incompatible with bleach- or acid-cleaned freeze-fracture replicas; c) the limit of resolution was presumed to be 3-5 nm; and d) there were no methods for directing the fracture plane to specific components within a tissue. Recent developments in “grid-mapped freeze fracture” and freeze-fracture immunocytochemistry have addressed each of these limitations.

Using grid-mapped freeze fracture, samples are mapped in three dimensions using confocal microscopy before freeze-fracture (Fig. A), as well as after replication and stabilization in Lexan plastic on a gold “Finder” grid (Fig. B). After replica cleaning, areas of interest are correlated in confocal and TEM images —— for example, cilia of the ependymal layer of rat spinal cord (Fig. C).

The resolution of any microscope is usually taken as the benchmark of its quality. When thinking about the scanning electron microscope(SEM) a comparison of its resolution to the resolutions quot-ed for other comparable instruments such as the transmission EM or the scanning tunneling micro-scope has unfortunately led some users to the view that the SEM is only a second best choice. In fact the spatial resolution that can be anticipated for an optimized SEM is highly competitive with its peers making it clearly the microscopy of choice.

The resolution of the SEM depends on two classes of factors; those which are inherent to the use of electrons as an imaging tool, and those which relate to the performance of the instrumentation (and possibly of the operator) used to perform the microscopy. The use of electrons as the scanning probe, and the choice of secondary electrons as the usual imaging mode, potentially affects the achievable resolution in three ways.