LAS Tour Speakers
To help support local society meetings, MSA provides reimbursement for travel expenses for a national speaker to attend LAS meetings. Speakers may be booked as available. Visit the LAS Programs Available page for more information.
Mike Marko, MSA President 2016
- Historical perspectives on biological EM and cryo-EM
- Cryo-FIB technology
- TEM phase-plate imaging
P. O. Box 509
Empire State Plaza
Albany, NY 12201-0509
John Mansfield, MSA Past-President
- ESEM Applications
- FIB Applications
- Electron Diffraction
- Cultural Heritage work with Microscopy and Microanalysis
John Mansfield PhD Cphys MInstP
North Campus Electron Microbeam Analysis Laboratory
Building 22, Room G010, University of Michigan
2800 Plymouth Road, Ann Arbor MI 48109-2800 USA
Phone: 734-936-3352 FAX: 734-763-2282
Ian Anderson MSA President-Elect
Independent Research Physicist
6856 SE Twin Oaks Circle
Stuart, FL 34997
LAS Sponsored Speakers
3-D and In-situ Characterization of Nanomaterials in the Scanning Transmission Electron Microscope
All nanomaterials are three-dimensional (3-D) in nature whether they are used for catalysis, energy storage, semiconductors, or medicine. While (scanning) transmission electron microscopes ((S)TEMs) are typically used to analyze these materials, the images are 2-D projections of 3-D objects. In order to understand the true nature of the nanomaterial, a 3-D tomogram is necessary on the nano- or atomic scale. Traditionally, this involves taking a series of images of the sample at different tilt angles, normally ranging between -70º to +70º every 1 to 2 degrees, and using these two dimensional images to reconstruct a three dimensional volume of the sample. This tilt range may increase depending on the sample geometry and the holder used, but there is a constant battle against an artifact in the reconstruction called the missing wedge. This effect may be reduced greatly by performing dual axis tomography, or overcome completely using new holder technologies, but each technique has its pros and cons. Another approach that has been taken in the last 3-5 years is the development of novel algorithms that greatly reduce the effects of the missing wedge and even provide atomic resolution 3-D tomograms from just a few projection images.
With recent advances in in-situ microscopy, a new era in microscopy has arrived that allows for the dynamic imaging of materials under reaction conditions. It is no longer sufficient to image materials in vacuum conditions, but to get closer to the conditions in which the material will be used, such as high temperature, liquid environments, gas environments or a combination thereof. Combining an in-situ or ex-situ experiment with STEM tomography is a very powerful method for materials characterization. The benefits and limitations of all these methods will be discussed through examples of different inorganic materials.
Ilke Arslan received her Ph.D. in Physics from the University of California at Davis, and is currently a Senior Scientist at the Pacific Northwest National Laboratory in Richland, Washington. She was recently honored by President Obama with the Presidential Early Career Award for Scientists and Engineers. Before joining PNNL, Ilke was on the faculty of the Chemical Engineering and Materials Science Department at the University of California, Davis, and still holds an Adjunct Professor position there. She has held fellowships from the National Science Foundation, the Royal Society, and the Truman Fellowship at Sandia National Labs. Her interests include catalysis, energy storage materials, and technique development in tomography, in-situ liquid and gas microscopy, and EELS.
Ilke Arslan, Ph.D.
Pacific Northwest National Laboratory
Physical Sciences Division, Institute for Integrated Catalysis
902 Battelle Blvd, K2-57, Richland, WA 99352
Tel: (509) 372-4603
C. Barry Carter MSA President 1997 IFSM Vice President, immediate Past-President
The Future of TEM and why we must Remember the Past
The subject of this talk concerns the future of TEM. TEM is facing many challenges including the fact that the top-of-the-line microscopes are becoming more expensive and more complex even when they seem simpler because of the increasing use of computers and a clear affordable textbook. The techniques used by the different communities (physical sciences and life sciences) are also often converging especially for those specializing in 3D imaging, spectral imaging, low-dose imaging (we all should be) and aberration-corrected imaging. (Who is specializing in non-aberration-corrected imaging?)
I'll illustrate the talk with some examples of work from my group and from friends. My field of research is Ceramic Materials so I'll use my crystal ball to suggest some potential directions that TEM as a whole might follow in the next few years, and in so doing explain the title.
Lucille A. Giannuzzi
Theory and Applications of ex situ Lift Out FIB Specimen Preparation
The focused ion beam (FIB) ex situ lift out (EXLO) technique for scanning/transmission electron microscopy (S/TEM) specimen preparation was historically the first lift out technique developed. Over the years, EXLO and manipulation of electron transparent specimens for S/TEM has been applied to numerous materials systems and examples will be presented. EXLO is also perfectly suited for manipulation of electron transparent specimens to MEMS carrier devices used for recently developed in situ TEM holders. Examples of EXLO of thick specimens for surface science analysis or analytical scanning electron microscopy will also be shown. Adhesion forces control the micrometer sized FIB specimens making the EXLO process feasible. The EXLO manipulation capabilities also aid in specimen preparation for particles and may be used for preparing fibers or other particulate materials. A disadvantage to conventional EXLO is the requirement of a support film for adhesion. Hence, the recent development of a new grid design and technique called EXpressLO™ allows EXLO and manipulation without a thin film support. The specimen is lifted out and manipulated directly to a slotted grid surface that provides the necessary adhesion forces. In this configuration, the specimen may be directly analyzed and/or further FIB milled, broad beam ion milled or plasma cleaned. Using this new grid design, a specimen can also be easily manipulated into a "backside" orientation which avoids curtaining artifacts after further FIB milling. Large Xe+ plasma FIB specimens can be lifted out via the EXpressLO™ technique. Lift out applications will be presented and various lift out techniques for FIB prepared specimens will be shown.
Lucille A. Giannuzzi received her B.E. and M.S. from SUNY Stony Brook and her Ph.D. from The Pennsylvania State University. She joined the University of Central Florida in 1994, ending her career at UCF as Professor, Department of Mechanical Materials and Aerospace Engineering. She joined FEI Company in 2003 as a product marketing engineer for FIB/SEM products. In 2010 she started her own consulting company and in 2013 spun off a product company which offers the EXpressLO™ FIB specimen preparation solution. Dr. Giannuzzi is a Fellow of AVS and MSA.
Lucille A. Giannuzzi
5483 Lee St, Unit 12
Lehigh Acres, FL 33971 USA
Jay Jerome MSA President 2006
The Role of Microscopy in Atherosclerosis (Hardening of the Arteries) Research
Microscopy has been critical to research in cardiovascular disease. This talk, aimed at the educated lay audience, reviews our current understanding of the causes of hardening of the arteries with particular emphasis on how microscopy has contributed to this understanding.
Quantitation of Biological Structure
A discussion of the various schemes for gaining quantitative structural information microscopically from biological samples using my own work as practical examples and how to avoid some common mistakes I have run across over the years.
Basic Digital Imaging and Image Formats
Most microscopy is now digital image based yet many microscopists do not fully understand basic digital image concepts. This talk covers, in a general, easy to follow manner, the basics of digital imaging in order to provide the microscopist with sufficient information to avoid common pitfalls. The field of "scientific" digital imaging is only a small subset of digital imaging. There are lots of things you can do in digital imaging that you should not do in scientific imaging because it destroys the integrity of your data; the image. Unfortunately, with modern digital imaging it is far too easy to inadvertently alter the image without even knowing that you have done so. In this talk, we review the basics of a digital image and discuss how to match the microscope parameters and image capture parameters in order to maximize image fidelity. We will also discuss post image processing and how these can affect the image data. Finally, the basics of image formats (JPEG, TIFF) are critical but not always understood, so we include a discussion of the appropriate uses of these formats. The image information is the data and not understanding basic "scientific" digital imaging can lead to accumulation of artefactual errors.
W. Gray (Jay) Jerome
Department of Pathology
Vanderbilt University Medical Center
1161 21st Ave, South
Nashville, TN 37232-2561
William J. Landis
The mechanism of mineral formation in vertebrates
Mineralization in vertebrates occurs normally in bone, calcifying cartilage and tendon, and dentin, cementum and enamel of the teeth. Vertebrate mineralization occurs through the interaction of inorganic calcium and phosphate ions with organic components synthesized and secreted to extracellular matrices by specialized cells of these various tissues. The principal organic constituent regulating mineral formation of all vertebrate tissues except enamel is collagen. The organic constituents of enamel mediating mineralization are amelogenin and enamelin. Largely through light and transmission electron microscopy, including high voltage microscopy and three-dimensional tomography, this presentation will describe the mechanism by which normal mineral formation occurs in association with collagen and amelogenin in the appropriate vertebrate tissues above. Reference to pathological mineralization, in muscle, skin, and other tissues, will also be made.
Tissue engineering of models of human digits and ears
Tissue engineering is a relatively new and potential powerful means of augmenting, repairing, and replacing various tissues that may be congenitally defective, injured, diseased, damaged or otherwise impaired in the human body. The approach of tissue engineering commonly involves seeding isolates of specific cells onto a biodegradable polymer scaffold to form a cell/scaffold construct. The construct is subsequently developed in vitro or in situ for ultimate use as a possible replacement tissue. Bone and cartilage structures, such as a human digit or ear, have now been modeled by tissue engineering methods. The presentation will describe by light and electron microscopy, correlated with laser capture microdissection and gene expression, the tissue engineering of current models of human phalanges and ears. Compared to bone and cartilage in vivo, these models demonstrate several similarities in structure, composition, and response to mechanical forces and they suggest great promise for further advances in clinical applications.
Effects of gravity on cultured bone cells
Many bones of vertebrates normally adapt to changes in mechanical forces, including the force of gravity. Typically the mass of responsive bones increases or decreases with increasing or decreasing force, respectively. This presentation will describe compositional and structural effects on cultured normal bone cells flown in microgravity during a series of NASA Shuttle spaceflights, retrieved on landing and analyzed by molecular biological techniques and light and electron microscopy. The data will be compared to results from the same cultured bone cells maintained at normal (1G) gravity. The information obtained from the NASA flights provides insight into the means by which bone mass is lost by astronauts in spaceflight and can be extrapolated to the observed bone mass loss in individuals confined to bed for long periods of time or lacking exercise and other forms of physical activity.
William J. Landis, Ph.D., is the G. Stafford Whitby Chair in Polymer Science at the University of Akron. He holds current joint faculty appointments at the Northeast Ohio Medical University (NEOMED), Kent State University, and Case Western Reserve University. Previously he held a joint faculty appointment at the University of Pennsylvania. He moved from Massachusetts to Ohio in 1998. At that time, he was an associate professor of Orthopedic Surgery and Anatomy and Cellular Biology at the Children's Hospital and the Harvard Medical School, Boston. In 1998 he took appointments as chairman of the Department of Biochemistry and Molecular Pathology and professor of Orthopaedic Surgery at the Northeastern Ohio Universities College of Medicine in Rootstown, OH (now NEOMED), and in 2010 he moved to the University of Akron in his present position. He has research interests in biomineralization, tissue engineering, and the effects of mechanical forces on mineralized tissues, and he has published more than 145 peer-reviewed journal articles, book chapters, and reviews in these areas. His long-standing research programs have been supported principally by the National Institutes of Health, the National Aeronautics and Space Administration, the Musculoskeletal Transplant Foundation, and the Austen BioInnovation Institute in Akron. He has been a Senior Fulbright Scholar at the Weizmann Institute of Science in Rehovot, Israel, and the recipient of several honors and awards for his research studies. He is a member of numerous scientific organizations, including MSA, MAS, the New England Society for Microscopy (NESM), and the Microscopy Society of Northeastern Ohio (MSNO).
William J. Landis, Ph.D.
Department of Polymer Science
Goodyear Polymer Center, Room 1201C
The University of Akron
170 University Avenue
Rapid Specimen Preparation Methods for Biological Electron Microscopy of Resin-embedded Samples
While many labs are moving toward cryo-methods for preparing biological samples for EM, the great majority of work involving sections of resin-embedded materials is still done by methods that are fundamentally unchanged from the 1960's. Consequently, the preservation of cell ultrastructure is compromised by the artifacts that result from this type of processing. In this presentation we will show how using cryofixation and low temperature dehydration we can greatly reduce the total processing time to a few hours, while producing results that have far fewer distortions compared to conventional preparation methods. High pressure freezing of living material is the preferred starting point, but we will discuss alternatives for samples that do not lend themselves to that approach. Dehydration and chemical stabilization of cell structure can be done by freeze substitution over a period of 3 hours or so, followed by infiltration with resin and subsequent polymerization in another 3 hours. Rapid methods can also apply to samples destined for on-section immunolabeling in either Lowicryl, or LR resins. Correlative light and electron microscopy techniques for preserving fluorescence in polymerized resin will also be discussed.
Kent McDonald received his Ph.D. in Botany from the University of California, Berkeley in 1972. Following a year teaching at UCLA, he took a Post-doctoral position at the University of Colorado, Boulder. In 1975, he re-focus his research on the cell biology of mitosis which led him back to Berkeley in 1979 to work on correlative light and electron microscopy of PtK cells and diatoms. In 1987 he returned to Boulder to join the High Voltage EM Lab and at this time realized the importance of cryofixation by high pressure. In 1993 he moved back to Berkeley to assume the Directorship of the campus Electron Microscope Laboratory. His current research interest is improving the instrumentation for correlative light and electron microscopy using high pressure freezing and tomographic 3-D imaging.
Electron Microscope Laboratory
26 Giannini Hall MC3330
University of California
Berkeley, CA 94720-3330
Sara Miller MSA President 2004
Emerging Diseases and Microscopy
Infectious diseases are the leading cause of death worldwide and the third leading cause in the US; many can be classified as emerging diseases. Some may be caused by truly novel pathogens, in other cases, the causative organisms have been present for many years, but have escaped detection until recently. Still others represent the re-emergence of known pathogenic organisms after a long period of quiescence. The mention of emerging pathogens brings to mind sensational, exotic and feared microorganisms such as Ebola virus, human immunodeficiency virus (HIV), hantavirus, West Nile virus, Yersinia pestis (plague), and prion diseases such as bovine spongiform encephalitis (BSE, mad cow disease) which have been associated with variant Creutzfeldt-Jakob disease (CJD) in humans. However, other organisms that have been known for some time can be classified as emerging pathogens as they continually mutate, recombine, and adapt, like influenzavirus, causing misery and death. A major category of emerging diseases is that of drug-resistant organisms. Changes in technology permit organism spread by contaminated water and air conditioning systems, by surgical and diagnostic instruments, or by transplantation. Geographic spread of disease organisms through more widespread human travel, transport of vectors in shipping containers, and increased mobility of insects and animals accounts for some emerging diseases. Food-borne illnesses are a world wide problem. In addition to agents that actually invade and cause disease, numerous organisms cause tremendous morbidity and some mortality through toxin production. Increased awareness due to better detection and identification methods has brought these organisms to the forefront. Microscopy techniques for detection of organisms are rapid and do not require specific probes. Electron microscopy is particularly useful for viral agents. It can visualize a wide variety of viruses at once, including non-cultivable and unexpected ones. It does not require antibodies or nucleic acid reagents for identification, and it is rapid; negative staining of fluid samples can be accomplished in a matter of minutes to a couple of hours. Thin sectioning of cells can be accomplished in one to two days. Finally, speed of microscopical methods and lack of requirements for probes are the reasons microscopy laboratories are being asked to participate in the surveillance for bioterrorism agents-horrible and feared examples of emerging diseases.
Sara Miller, Univ Georgia BS (Microbiol/Chem) 1968. Univ Georgia PhD (Microbiol) 1972. Director, EM Diagnostic Virology Lab; Director, Surgical Pathology EM Lab, Duke Hospital. Director, EM/IEM Cancer Ctr Shared Resource, Duke Univ Med Ctr. Assoc Res Prof, Microbiol 1983-; Assoc Clin Prof, Pathol 1994-.
Duke Medical Center
Pathology, Box 3712
Durham, NC 27710
David W. Piston MSA President 2010
Imaging the Molecular Mechanisms of Glucose-Stimulated Insulin Secretion
The islet of Langerhans is the functional unit responsible for glucose-regulate secretion of insulin and glucagon, and thus plays a key role in blood glucose homeostasis. Over the last 25 years, we have been interested in understanding the multicellular mechanisms of islet function, and their role in the regulation of blood glucose under normal and pathological conditions. Using our unique quantitative optical imaging methods and novel microfluidic devices, the dynamics of these molecular mechanisms can be followed quantitatively in living cells within intact islets.
Many of these experiments require hyperspectral imaging and live-cell Förster resonance energy transfer (FRET) with differently colored fluorescent proteins. We have exploited spectral confocal microscopy for quantitative multi-color imaging as well as FRET. Current spectral imaging devices are limited by a trade-off problem between image acquisition rate and signal throughput and dynamic range. To overcome this problem, we have used a snapshot hyperspectral imager — Image Mapping Spectrometer (IMS). The IMS utilizes an image mapper to remap a sample's x,y,λ datacube to a 2D detector array for parallel measurement. Since no scanning is employed, the whole datacube can be captured in a single snapshot, and optical throughput is maximized. FRET measurements can also be facilitated using photoswitchable FPs to achieve a lock-in type detection. This allows FRET to detect the interactions of a few fluorophores even in a sea of non-interacting molecules.
As an alternative to FRET, we have leveraged two-color Fluorescence Fluctuation Spectroscopy (FFS), which can be used to directly measure diffusion and binding rates of proteins within the cell. Using FFS to measure protein binding and diffusion allows quantification at the molecular level of protein interactions, and also overcomes many of the difficulties found in FRET imaging experiments. FFS can also provide information on protein molecular concentrations. Quantifying the molecular concentrations and interactions within islet cell pathways illuminates signaling mechanisms, and thus provides essential information for developing therapeutic treatments for patients with diabetes.
Many aspects of these studies can be covered in a single lecture with a focus on imaging technology, quantitative data analysis, or biomedical applications.
David W. Piston is the Edward J. Mallinckrodt Jr. Professor and Head of the Department of Cell Biology & Physiology at Washington University in St. Louis. Dr. Piston was trained in physics at the University of Illinois, completed a postdoctoral research fellowship in applied physics at Cornell University, and joined the faculty at Vanderbilt University in 1992 where he remained until 2014. He served as President of the Microscopy Society of America in 2010, and has been recognized as a Beckman Young Investigator Award (1993), NIH Study Section Chair (2004-2006), Searle Scholars Advisory Board (2006-20112) and election as a Fellow of the American Physical Society (2000), the Microscopy Society of America (2014), and the American Association for the Advancement of Science (2016). His research group focuses on the understanding the molecular mechanisms that underlie hormone secretion from islets of Langerhans in the pancreas. Driven by this biomedical focus, the lab develops, optimizes, and applies novel fluorescence microscopies and probes, largely based on the Green Fluorescent Protein and its relatives.
David W. Piston
Professor & Chair
Department of Cell Biology & Physiology
660 S. Euclid Ave.
Campus Box 8228
St. Louis, MO 63110
Some Basics of Confocal Imaging: How Deep is Deep and are They Really Colocalized?
Questions asked by operators of confocal microscopes often include how deep can I image into a tissue, are the labeled structures co-localized, and can I quantify the fluorescence in the image? This presentation will discuss the role of optics, selection of appropriate fluorochromes, and the proper set up of imaging parameters on a confocal microscope to correctly address these types of common questions.
Bob Price received his graduate degrees from Southern Illinois University and has directed core imaging facilities at Southern Illinois University, Case Western Reserve University and The University of South Carolina School of Medicine. He has participated in and led a number of confocal microscopy workshops. He is currently Editor-in-Chief of Microscopy and Microanalysis.
USC School of Medicine
6439 Garner's Ferry Road
Columbia, SC 29209
Fungi in the Human Environment
Fungi are fundamentally recyclers. Their main function in the environment is to break down complex materials, which allows the components to be re-used by other organisms. These complex materials include dead plants, dead animals, building materials, valued artifacts of civilization and any number of other things. Problems arise when these organisms invade the built environment, either work or living spaces. Various methods, such as air sampling, have been commonly used to estimate the density of fungi in a structure. Volumetric sampling may indicate high levels of fungi or one particular fungus in a building compared to the outdoor environment or some predetermined standard. This method may indicate the presence of viable fungal conidia or hyphal fragments in the air column but it cannot identify sites of colonization. Surface cultures may indicate the presence of viable fungal propagules but do not prove colonization. Surface sampling for light microscopy using clear adhesive tape mounts may demonstrate the presence of colonizing fungi. The methodology, such as types of tape and optics employed may affect the results obtained. Examination of tape samples from environmental surfaces may show the level of colonization and, in many cases, allow for identification of colonizing species. Scanning electron microscopy studies of suspect materials may determine the nature of surface features and contamination not readily identifiable in the light microscope. Suspect materials may be shown to be biological in nature or non-biological surface. Microanalysis of materials may yield clues to the origin of non-biological contamination. Rapid and accurate analysis of suspect materials on indoor surfaces is vital to the identification of potential fungal colonization sites. These data may be used as an aid to determining an appropriate course of action.
Microbial Ecology of Extreme Environments: Automobile Air Conditioning Systems
Automobile air conditioning systems might be considered an extreme environment for many microorganisms. Organisms surviving and proliferating in these systems may be presented with temperature changes ranging from sub-zero to over 140oF, water activity from saturation to dryness and a nutrient complexity including varying levels of hydrocarbons. Microbial communities may develop in these systems and sometimes proliferate to the extent of massive colonization and production of objectionable odors. In a few instances microorganisms emanating from ACS have been associated with hypersensitivity pneumonitis and other allergic reactions. We have demonstrated that foam insulation and glues, in particular, on system insulations may be colonized by fungi such as Aspergillus, Aureobasidium, Cladosporium, and Penicillium. Such fungi often are implicated in colonization of similar substrates in buildings categorized with the sick building syndrome. Combined light microscopy, scanning and transmission electron microscopy and culture techniques have provided profiles of the microbial communities which inhabit some automobile air conditioning systems.
Microscopy into Art: Adaptive Interpretation (NEW!)
Beautiful images captured through microscopes are often considered works of art in themselves. Artists have been taking these images as inspiration for creative adventures using media other than photography to convey the complex and often breathtaking sights previously seen by only a select few. In the hands of artist such as Salvador Dali, Fuco Verdura and Luke Jerram the (mostly) unseen world comes alive through the use of glass, paper, paint and other media bringing microscopic worlds to light. The ever-present influence of microscopy on the art world is enlightening, inspiring and often quite surprising.
Robert Simmons is a native of Atlanta, Georgia. He earned his Bachelor of Science (Hons) degree in biological sciences at the University of Ulster, and continued with MS and Ph.D. degrees at Georgia State University. He joined the Biology Department at Georgia State University in 1983 and is the Program Director for Biological Imaging. His main research involves the interaction of microorganisms with the human environment, with an emphasis on fungi and air handling systems. Recent work includes investigation of colonization of hydrogel contact lenses by Fusarium and other fungi.
Department of Biology
College of Arts and Sciences
P.O. 4010, Georgia State University
Atlanta, GA 30302-4010
Janet H. Woodward MSA President 2012
Real World Microscopy: Problem Solving in the Paper Industry
In the paper industry, paper breaks and the production of off-quality paper must be addressed quickly. The most basic microscopic tools and techniques often provide rapid and reliable solutions to these problems. Tools include the stereomicroscope and bright field light microscope. Basic techniques include a variety of staining methods: simple and differential staining of deposits, staining for fiber differentiation, and staining for starch. Cross sections of paper or board can be made with a razor blade for microscopic examination. All of these techniques can be done on-site. Other problems, such as coating defects or poor ink receptivity, require tools and techniques that are not available at a mill site. Conventional SEM, ESEM, BSE, EDX, and FTIR are routinely employed to address these and other "real world" problems in the paper industry.
Janet H. Woodward
1256 North McLean Blvd
Memphis, TN 38108
Nestor Zaluzec MSA President 2011
The Evolving Prospects for Microanalysis in the AEM
797 Bonnie Brae Ct
Bolingbrook, IL 60440