The protozoan parasite Giardia, an intestinal flagellate, has evolved an unique attachment organelle called the ventral adhesive disk (VAD). This attachment organelle mediates attachment of the trophozbite to the microviUous border (MVB) of intestinal absorptive cells, and production of lesions via attachment are thought to lead to diarrheal disorder characteristic of giardiasis. The VAD has contractile proteins arranged around it's circumference and it has been suggested that contraction of this area might function like a purse-string suture. The resemblance of the VAD to a suction cup has led to the hypothesis that suction or a negative pressure produced under the VAD by a grasping action might produce the adhesive force regulating attachment. To test whether or not the generation of a negative pressure is necessary for attachment, experiments were designed in which the substratum was fabricated to have an uneven surface, or the substratum was composed of different arrangement of pores.

Numerous studies have demonstrated that fungi are capable of colonizing a range of indoor construction and finishing materials as well as a number of air handling system components. Many of the species found are known to produce mycotoxins and demonstration of the presence of ‘toxic mould’ in both the workplace and in dwellings has become a major issue. Mould issues now are the focus of litigation or are currently in contention in a number of states and hundreds of millions of dollars are at stake.

The indoor environment harbors a variety of microorganisms, including bacteria and fungi. Under normal indoor conditions most of these organisms are present as dormant forms. The fungi persist either as conidia (asexual spores), sexual spores, or hyphal fragments. The presence of water in an environment may lead to the growth and proliferation of fungal species, sometimes resulting in deterioration of the colonized substrates with possible negative effects on IAQ.

Nearly five years ago, Representative Vernon Ehlers, in his report to Congress on a House of Representatives study entitled “Unlocking Our Future: Toward a New National Science Policy,” noted that the American public does not understand science and its practice. A major recommendation that emerged trom this study was the need to “make scientists socially responsible.“ This sentiment was echoed in a National Research Council's report, “Materials in a New Era,”, where Neal Lane, former Director of the National Science Foundation, was quoted as saying, “It is necessary to involve material scientists in a new role, undoubtedly an awkward one for many, that might be called the ‘civic scientist’.” Why the concern? The answer is clear. “Our prosperity, security, and health depend directly on the educational achievement of all students, not only those who will become scientists and engineers, but all workers, voters, parents, and consumers.”

A silicon multi-cathode detector (SMCD) has been developed for microanalysis and x-ray mapping applications. The SMCD has a large active area (∼0.5 cm2), excellent energy resolution, and high count rate capability. The detector utilizes novel structures that have produced very low dark current, high electric field, uniform charge collection, low noise and high sensitivity to low energy x-rays. The detector's spectral response was evaluated using a 55Fe radioisotope source, as well as by fluorescing materials with an x-ray generator. Figure 1 shows a 55Fe spectrum with an energy resolution of 125 eV FWHM at 5.9 keV collected at 12 μs peaking time. This energy resolution has been repeatably measured on many different detectors. To evaluate the high count rate x-ray performance, which is very important for fast x-ray mapping, a Cu sample was fluoresced using a Rh-anode x-ray tube.

The confocal laser-scanning microscope (CLSM) has enormous potential in many biological fields. When tests are made to evaluate the performance of a CLSM, the usual subjective assessment is accomplished by using a histological test slide to create a “pretty picture.” Without the use of functional tests many of the machines may be working at sub-optimal performance levels, delivering sub optimum performance, and possibly misleading data. In order to replace the subjectivity in evaluating a confocal microscope, tests were derived or perfected that measure field illumination, lens clarity, laser power, laser stability, dkhroic functionality, spectral registration, axial resolution, scanning stability, PMT quality, overall machine stability, and system noise (1-3). It is anticipated by using this type of test data, performance standards for confocal microscopes will be obtained and the current subjectivity in evaluating CLSM performance will be eliminated.

Promoting and maintaining a facility is a major concern to many of us. I had the pleasure last spring of participating in the 2nd International Cryo-workshop held in the Bio-Imaging Facility at the University of British Columbia. I talked to the facility director, Elaine Humphrey, and saw what she has done in a relatively short number of years in building that facility. I was incredibly impressed so I asked Elaine if she would address mis topic.

Elaine started out as a marine biologist in England and came across a problem that required electron microscopy. The first facility she went to was very discouraging saying that it was an extremely difficult technique. She managed to find another facility that made it easy.

Principal components analysis of multivariate data sets is a standard statistical method that was developed in the early halt or the 20th century. It provides researchers with a method for transforming their source data axes into a set of orthogonal principal axes and ranks. The rank for each axis in the principal set represents the significance of that axis as defined by the variance in the data along that axis. Thus, the first principal axis is the one with the greatest amount of scatter in the data and consequently the greatest amount of contrast and information, while the last principal axis represents the least amount of information.

Renishaw's structural and chemical analyser (SCA) provides new analytical techniques for scanning electron microscopes (SEM). For many years energy dispersive X-ray analysis (EDS) has been the default in-SEM analytical technique, and whilst it can provide fast and accurate data from any samples, it can only provide elemental information about the sample being analysed. For materials applications, assumptions about stoichiometry may enable accurate characterisation, but in most Instances “chemistry by guesswork” does not provide an acceptable solution. Furthermore, EDS is rather insensitive to light elements - atomic number 11 (sodium) and below, which severely limits its applicability to organic chemistry and life sciences.

A major challenge in optical microscopy is to develop techniques with high spatial resolution, sensitivity, and chemical specificity. The latter, chemical specificity, is typically achieved through some form of labeling, which has potential to alter the nature of the sample under investigation. Raman or infrared (IR) microscopy can be utilized to image samples in their natural form using molecular vibrations as a contrast mechanism. IR microscopy suffers from spatial resolution issues, and spontaneous Raman microscopy suffers from low scattering cross-sections, so that high laser power is often required, introducing the possibility of sample photo-damage. Scattering cross-sections for Coherent Anti-Stokes Raman Scattering (CARS) are typically several orders of magnitude greater than those of spontaneous Raman Scattering. This, in addition to the high spatial resolution inherent in nonlinear optical microscopy, has led CARS microscopy to begin emerging as a powerful, noninvasive technique for biological and material imaging.

At the Microscopy and Microanalysis 2003 meeting, a member of the audience in one talk sharply criticized a speaker of a talk for his use of a Savitsky-Golay (S-G) digital filter for smoothing Electron Energy Loss Spectroscopy (EELS) data. The statement was made that the digital filter would decrease the resolution of the peak, i.e. widen the width, and that all digital filters do that, including the Top Hat filter. In support of this statement, he said to consider what would happen if the filter were applied to a Dirac delta function. I knew that his argument was not correct, but I didn't think of the reason until later. This interchange was on my mind because I was familiar with both of these filters due to implementing both the Top Hat and the Savitsky-Golay filters into my “Beerware” program, EELS-PLOT.

The cell is the fundamental unit of all living organisms, ranging from the unicellular archaea and bacteria (prokaryotes) to higher multicellular plants and animals (eukaryotes). All cells are bounded by a complex and dynamic plasma membrane, which functions principally to maintain cellular and organismal steady state by performing complex energy transformations and regulating the flow of information for the cell. The cell membrane also performs a number of vital housekeeping functions, which include control of the transport of substances between extracellular and intracellular environments, participation in cell signaling cascades by hosting receptors of extracellular ligands, and facilitating critical cell-to-cell communications in multicellular organisms (Karp, 2005).

Considerable research over the last fifty years has significantly increased our understanding of cell membranes and their structural organization. Every membrane is fundamentally comprised of a dynamic lipid bilayer that supports a variety of transmembrane and membraneassociated proteins.

One of the methods of preparing samples for analysis by Transmission Electron Microscopy (TEM) is the well-known procedure using a Tripod polisher and the wedge technique (1-3), developed in our laboratory. Though developed explicitly for preparing samples of integrated circuit structures built on silicon, the technique has been used in our lab for a wide variety of other materials, including metals and ceramics. In general when working with silicon samples, we have the luxury of starting with large wafer pieces or chips that are generally at least a couple of millimeters square or larger. Recently, however, we needed to work for an extended period on GaAs based lasers, where the devices requiring analysis were individual lasers extracted from individual packages. These small samples, measuring 100x100x50 microns, are too small for the manual handling involved in routine mechanical cross sectioning methods.

Let's face it—uranyl acetate is the perennial bad boy of chemicals, at least from the standpoint of disposal. A necessary evil in the electron microscopy laboratory, it presents a quandary for those trying to comply with hazardous materials regulalions, an effort complicated by its nominal radioactivity. Disposal costs of radioactive materials can be daunting. A past posting to the Microscopy Society of America listserver gave an estimate of US$7000 per liter and local follow up indicated that, although probably high, that may not be an unreasonable figure. In these times of diminished subsidies and expectations of increased cost recovery by academic service facilities like ours, this is of paramount concern.

The Electron Microscopy Core Facility at the University of Missouri-Columbia follows guidelines issued by our campus Environmental Health and Safety (EHS) office regarding hazardous materials and their disposal.

Modern microelectronics have rapidly decreased in geometry to enhance the speed and processing power of computers. Advanced devices are approaching design rules of sub 0.13 micron in size, and the trend continues at the rate dictated by Moore's Law, Coupled with this reduction in device size, is a change in materials used for producing these devices. Traditional aluminum interconnect metallurgy and oxide dielectric materials are being replaced with copper and low-k materials in an effort to continue the trend of shrinking device sizes and higher processing capacities.

These changes in materials and device sizes have provided the impetus for alternative methods for producing cross sections. Although focused ion beam instrumentation has been successfully used for preparing cross sections, a combinatorial approach using polishing and argon ion milling has been found to dramatically enhance the ability to produce high quality cross sectional samples in a reasonably short amount of time.

Insect tissue is often difficult to prepare for electron microscopy because of the impenetrable barrier surrounding the body tissues. Drosophila salivary glands have been used for numerous studies because of the large size of the cells and their large polytene chromosomes. Early TEM studies of salivary glands used a protocol that took several days. We were able to achieve excellent preservation and good ultrastructure in Drosophila salivary glands and imaginal discs from Stage L3 larvae using microwave processing in a protocol requiring less than 2 hours.

We used a Pelco Laboratory microwave (model #3451) equipped with a Cold Spot, Steadytemp chiller/recirculator run at 15° C, and vacuum chamber (Ted Pella, Mountain Lakes, CA). The heads and attached salivary glands were removed from the animals and placed in PBS. The tissue was transferred to Pelco prep-eze specimen holders (#36157) for ease of handling. Our goal was to use the microwave effect, not the heating effect, to prepare the tissue.

If we look back at the year 2003 it becomes quite clear that the South African microscopy year was quite unique. …Of course, I can't speak for all the companies and laboratories in the country, but from my personal view it was definitely interesting.

FEI definitely had a great year in that they managed to sell 5 systems. Then LEO added to that another 3, and we have heard that there is also a Cameca probe being bought. This may not sound like a lot, but if you take into consideration that in the last few years an average of 4-5 EM columns per year would register as a banner year. Given that we have seen our currency improve over the past 2 years, it does imply that buying imported equipment is getting cheaper. However as this effect has also negatively affected exports, we can't say that we have become richer, and so could afford more systems.

Flicker stereo alternates two pictures (∼ 1 frame/sec.) of the same structure with views tilted 6° ± 1° about a vertical axis—Natural appearing stereo visualization results. A flicker stereo program to be used for viewing with a personal computer (PC) was developed by the authors. The two images are installed, adjusted for superposition and then alternated on the computer monitor or computer projection system. Flicker stereo can be applied to micrographs from the optical microscope, SEM, TEM, pictures or line drawings. Flicker stereo provides simultaneous three dimensional viewing for several individuals without special viewers, projection systems and/or glasses. Different flicker images may be retrieved and viewed on a PC in rapid succession. Flicker stereo facilitates technical discussion between collaborators and provides a convenient approach for 3D visualization by audiences using a digital LCD projection. The program is available to readers.

Laser scanning confocal microscopy has been used to analyze cytological specimens for research purposes. However, for many pathologists, its application in diagnostic cytology still is terra incognita. In our laboratory, confocal microscopy has been used for seven years, mainly to solve problems in cervical cytology, in breast aspirates, and for endometrium cytology.

In contrast to histology where we can cut new sections from the block, the cytologic sample is often limited to one smear. Accordingly, when we wish to expand our diagnostic possibilities, we have to extract the desired additional information from that single smear. This is possible by first de-staining the original smear and subsequently re-staining it. In this article, we describe how we restain old slides with the fluorescent dyes Ethidinm Bromide and Eosin, a DNA/protein stain, resulting in staining patterns comparable to the diagnostic Papanicolaou and Giemsa methods in that mere is an optimal distinction between nucleus and cytoplasm.

Scanned Probe Microscopy (SPM) techniques are attractive because they provide easily acquired micrographs that map specimen properties with nanometer scale resolution. SPM micrographs can be collected without the use of high vacuum (as opposed to many electron microscopies). Moreover, SPM methods allow in-situ imaging of specimens in a variety of environments, including under liquids and at higher temperatures. However, from a metrology perspective, the great promise of SPM must be balanced by the fact that SPM techniques generally provide qualitative data unless supplementary actions are taken. That is, quantification of SPM image contrast is difficult since it depends highly upon probe characteristics, probe/ sample interactions (both of which can be difficult Eo gauge), and instrument calibration.

There is no substitute for good microscope care. A unit that is stored and covered, with its lens protected in a constant temperature/ humidity environment, which is dust free (in the laboratory away from acid wash areas) speaks for itself. During cleaning, oculars in particular, microscope rubes should be covered to prevent entry of dust. A lens should be inspected with a view to cleaning if there is a lack of contrast, cloudiness or poor definition in the images it produces.

While extolling the virtue of cleaning microscope components, it should be emphasized that cleaning can cause considerable damage if it is done incorrectly.