It is well known that birds frequently attract mates by a ritual display of feathers or other ornaments. The colors for these displays are either derived from pigments, such as carotenoids, or photonic crystalline arrangements near the surface of the skin, feathers, or other ornaments. Add to this the fact that many birds can perceive light in the near-ultraviolet end of the visible spectrum and the range for these visual displays is extended. In a spectrophotometric and microscopic study by Birgitta Dresp and Keith Langley, it was demonstrated that a unique arrangement of crystal-like structures near the surface of the beak horn of the King penguin (Aptenodytes patagonicus) most likely accounts for a photonic effect that probably plays a role in their mating rituals.

At the August M&M-2006 meeting in Chicago, we were standing next to our poster titled A Different Kind of Microscopy: Analyzing Features with an Automated Electron Beam when an acquaintance with long experience in electron microscopy wandered by. After a glance at the poster title, he challenged: “What's different about that?” Upon hearing our summary he asked (with what we took to be an encouraging tone of voice): “Are you going to publish this?” We had enough similar reactions from others to make that seem a good idea, and this is the result.

This discussion should begin by noting that the operative word in the title is “different,” not “new.” In point of fact, the foundations for the technique were laid in the 1970s when some workers began putting scanning electron microscopes and microprobes under software control by interfacing them to the “minicomputers” that powered the computerized x-ray analyzer units then entering the market. Even prior to this, there were a few “hard-wired” image analyzers that mechanized the process of extracting information from microscope images. Thus, automated analysis of features via an electron beam instrument is hardly a new concept.

Microwave-assisted preparation of histological samples has been performed for decades; what began with a few pioneering researchers has now become a routine and accepted practice in many clinical and research laboratories. Reliable, reproducible microwave protocols have been developed for a variety of operations: LM and EM processing, decalcification, fixation, special stains, antigen retrieval and more. Laboratories employing microwave procedures often do so for several compelling reasons: in addition to the expected time savings (often on the scale of orders of magnitude), improved morphology, retained immunoreactivity, and the elimination of hazardous reagents are benefits typically realized as well.

Despite the increasing availability of laboratory microwaves, consumer-grade (“kitchen”) microwaves continue to be used, almost invariably due to cost considerations. (EBS has maintained since 1992 that a kitchen microwave has no place in the lab.) At any time in the US there are hundreds of microwave models to choose from: a dizzying array of sizes, wattages, options, and configurations await the shopper.

Recent developments in nano- and semiconductor technology have substantially increased the demand for accurate and efficient site specific cross-sectioning of specimens and preparation of TEM samples. Moreover, nano-research is facing new challenges for manipulation, observation, and modification of devices on a submicron scale. At the same time in materials science a new focus on analytical nanoscale investigations—not only of specimen surfaces and cross sections—but on sample volumes is emerging.

These demanding requirements can be met if a focused ion beam (FIB) column for nanoscale structuring is combined with a high resolution SEM that is used to monitor the FIB milling and deposition process on a nanometer scale. Such an integrated Cross-Beam® system enables the high resolution observation and direct control of the FIB milling process in real time. Using this concept it is possible to prepare site specific TEM samples and cross sections with nano-scale accuracy. Such a system can be complemented with a gas injection system (GIS), for deposition and enhanced etching of specific materials, as well as, in-situ micro manipulation systems, and analytical detectors such as EDX and EBSP systems.

Atomic Force Microscopy (AFM) is a key technique for the measurement and analysis of samples when nanoscale topography is of interest. It offers a number of complementary probing modes that extend an AFM's measurement capability to a wide range of material and transport properties of surfaces, including hardness, friction, conductivity and adhesion among others. Sample temperature controlled AFM extends the study of surface morphology and properties to include changes in the material phases.

Recently, silicon microfabricated AFM cantilevers that have integrated heaters, as shown in figure 1, have become commercially available. These cantilevers were initially developed for probe based data storage by researchers at IBM Zurich, Figure 1. With the availability of these cantilevers, AFM measurements can be performed where the tip is heated as opposed to the sample.

The semiconductor industry has unquestionably entered the realm of nanotechnology. Critical dimensions of many features are specified in nanometers. Gate oxides are only a few nanometers thick. Barrier and seed layers for copper processes are not much more. Gate lengths are forecast at less than 20 nm by the end of the decade. Additionally, the drive to increase device density is leading to the adoption of FinFET and other new transistor designs that include complex three-dimensional structure. Even conventional planar CMOS designs now incorporate processes such as damascene interconnects that are inherently three dimensional. The combined need for higher spatial resolution and cross-sectional imaging of complex structures has led to a significant increase in the demand for scanning transmission electron microscopy (STEM) and (TEM) in semiconductor manufacturing operations.

Serial sectioning of TEM blocks is a common technique. The more sections that are desired, the more complex the procedure becomes. Some of the problems faced are staining without losing or obscuring sections, appropriate block trimming, finding the area of interest in the next section, transferring sections to the grid. Here, I relate a set of techniques that permit the microtomist to regularly get over 100 consecutive serial sections on a single slot grid. Much of what is discussed below is not new, but by putting all of these techniques together I believe that a powerful tool for TEM is presented.

The first issue that must be addressed is how to stain the sections on the grid without losing sections or obscuring sections with staining artifacts (e.g. lead pepper). The simplest solution is to stain the sample with both uranium and lead prior to embedding and not stain the sections on the grid.

Recent advances in in-situ and tomographic methods using (scanning) transmission electron microscopy ((S)TEM) provide the unique possibility of obtaining 3-dimensional (3D) information and in-situ studies of gas-solid chemical reactions on the nanometer scale. In combination with the well-known analytical techniques, electron energy-loss spectroscopy (EELS) and energy dispersive X-ray spectroscopy (EDXS) we can obtain 3D, dynamic, analytical, and structural information down to the atomic level.

The JEOL JEM-2500SE STEM/TEM at UC Davis (CHMS) is equipped with a Schottky field-emission source operated at 200 kV, a post column Gatan imaging filter (863 GIF Tridiem) for EELS and a Thermo System Six for EDXS.

Microelectronics and semiconductor wafer manufacturing are among the fastest evolving technology industries today. Wafer sizes typically are 200 mm to 300 mm while critical dimensions are shrinking to 0.09 μm and smaller. As the size of discrete devices continues to be reduced while device density increases, the need for fast, accurate, flexible metrology and inspection tools in the microelectronics industry grows.

Back in the early 1980's, semiconductor inspection was performed primarily by brightfield optical microscopes and with automated detection tools. The adaptation of automated detection tools led to the systematic control of increasingly smaller defects. The smallest detectable defect using these automated tools fell to below the 0.30-micron mark during the 1990's.

I took over the facility in 1996. We had aging equipment, no assistance, there were huge budget cuts that year and I was told, “There was no money. The only money you're gonna get are your wages. Everything else has to come from user fees.” I had to find funds for service contracts and we had to increase our user base.

I figured I would have to find an assistant because there is no way one can run a TEM, an SEM and confocal and light microscope facilities alone. So I found a grant called “First Job in Science and Technology” which would give me a graduate student for one year, and we had a summer student who had just graduated and was looking for a job before she went on to graduate school. She learned really fast and by February she was excellent, but by May/June we were back into the same boat again.

In case you didn't realize it, one of the things that is important about a business plan is to have an enthusiastic champion. I don't know if I've seen a person that has represented what she does more enthusiastically than Elaine did just now.

One of the reasons why I asked Elaine to go first was there's a lot of preparatory work that goes in place before you put together a business plan. Elaine did a very nice job in planning, and one of the key things in her Strategic Plan was that she took planning and made it a part of the business plan. A “S.W.O.T.” Analysis is an exercise that you go through when you are dealing with an enterprise that talks about Strengths, Weaknesses, Opportunities and Threats. Elaine put together the S.W.O.T. Analysis in addition to the current facility description and wish lists and that was extremely helpful in putting together her business plan.

Colloid and adsorption physics are a very complicated business and often have a lot of surprises. In 1991 Kramarcy and Sealock published a paper in JHC Vol. 39, No. 1,pp. 37-39, 1991: “Commercial Preparations of Colloidal Gold-Antibody Complexes Frequently Contain Free Active Antibody”. Their data indicate that proteins adsorbed onto colloidal particles of 5nm and larger can dissociate from the particle surface with time and that, at times even shortly after manufacturing, colloidal gold reagents may contain free binding molecules. This is not necessarily the result of bad manufacturing practice, as adsorption and desorption are in equilibrium at all times. Some proteins (there are even variations between antibodies from different animal species) are more liable to become dissociated than others, and the conditions of coupling play a role as well.

Nanowires or porous films grown on a substrate normally lack mechanical strength, and may be subject to damage during specimen preparation. When we made cross-sectional TEM specimen for this type of sample, we modified the traditional method by covering the sample with epoxy to improve the film strength, and applying single-section ion milling to protect the film from over-milling.

The sample surface is first covered with G1 epoxy. We choose G1 for this application because it is relatively thick and cures at low temperature. For samples with a dense-growth of nanowires or a thick porous film, a brief moment in vacuum helps to get rid of the air bubbles in the epoxy. The glue is cured at 100 degree C for 10 minutes, until its color turns to a reddish brown. To remove the excess glue and flatten the surface, the sample is then ground and polished until the glue is less than 0.1 mm thick.