The MSNO website (www.msneo.org) is the place to check out the latest info about MSNO, and related local society meetings. There are live links to many of our corporate members and to the sponsoring parent societies. If you have something you would like submitted to the website, please submit it to any of the Board members for review. The Board corresponds electronically and can reply quite quickly to your submission.

Each spring, Case Western Reserve University’s Macromolecular Science Department hosts the annual meeting for its Center for Applied Polymer Research, Inc. CAPRI consists of corporate member companies and the Macromolecular Science department professors and students; the corporate members sponsor directed research that is done by the department professors and students. The meeting is a combination of extensive talks in the morning, student poster session competition in the afternoon, tour of the Macromolecular Science labs and a mixer and dinner in the evening. In 2002, Ed Boyd was one of Noveon’s representatives to CAPRI, and graciously agreed to write this review of some of the talks and posters that featured microscopy or microanalysis methods.

The Twenty-First Annual Symposium of the CAPRI was held on Wednesday, May 29^th, 2002 at the Kent Hale Smith building on the Case Western Reserve University Campus. Representatives from twenty-four companies were in attendance to view presentations of research conducted by students of the Department of Macromolecular Science and Engineering. The event was hosted by Professors Anne Hiltner (Director of CAPRI) and Eric Baer.

A variety of topics were represented covering many areas of the structure-property-processing relationships of both polymer and polymer composite materials. The diversity of research interests in the department was emphasized in the twenty-two separate poster presentations on display. Registered attendees were also presented with summaries of all posters on display as well as Volumes VI and VII of the published Collected Papers of CAPRI scientists and engineers. Also distributed was a spiral-bound notebook showcasing the instrumentation available at CAPRI. Noted was the presence of a variety of optical microscopes, a JEOL 840A SEM, an RMC MT6000XL microtome with a CR-X cryo-attachment, and a recently purchased DI Nanoscope III AFM.

The day began with registration at 8:30 AM followed by 6 twenty-five minute, detailed presentations by some of the graduate students on the work that complemented their poster presentations to be viewed later in the day. Lunch was served at 12:15 PM at “The Restaurant” at Severance Hall where students, faculty and attendees were given an opportunity to interact. At 1:45 PM the remaining students, who were also presenting posters, were allowed to give a brief, three-minute synopsis of their poster presentations. From 2:45 PM to 4:45 PM corporate attendees were encouraged to view the twenty-two posters and select their personal choice for “best poster”. At 4:45 PM the votes were tallied and the top three poster winners were held in confidence until dinner that evening. At 5:15 PM attendees were treated to a “Facilities and Art Tour” of the Kent Hale Smith building. At 6:30 PM the social hour and dinner were held at the University College Club where delicious fare was enjoyed by all and the top three poster presenters were given a monetary award.

In general the posters were rich with optical, AFM, SEM and even TEM images, examples of which are given as follows: Optical images were used to discern the morphologies and layer thickness of co-extruded polymers and crack propagation. Polarized light images were used to denote the presence of crystallinity in specific stoichiometries of copolymer systems. AFM images were used to characterize polymer interfaces, to investigate adhesion properties, and to identify regions of immiscibility in co-monomer systems. SEM was used to show regions of microporosity and filler content in polymer samples.

Overall, the day was quite enjoyable and rich with examples of applied microscopy. I encourage all of those interested to attend next year’s symposium.

Denise Wade Rafferty, long-time MSNO member and former Board member, presented the following talk at the 2002 Cleveland Society of Applied Spectroscopy (SAS) May Conference. Denise’s work was done with Professor Jack Koenig of the Macromolecular Science Department at CWRU.

Diffusion of Binary Non-Solvent Mixtures in Polymers: Aqueous Ethanol Solutions in Polyvinyl Acetate by Denise Wade Rafferty and *Jack L. Koenig, Macromolecular Science Department, Case Western Reserve University, Cleveland, Ohio 44106 USA

The diffusion of binary non-solvent mixtures of ethanol and deuterium oxide (D₂O) into polyvinyl acetate (PVAc) was studied by FT-IR imaging. Initial diffusion rates were calculated for all mixtures from the concentration profiles, and a positive deviation from ideal behavior was observed; higher initial diffusion rates and greater degrees of swelling were observed for the solvent mixtures than for the pure liquids. PVAc swells to various extents in pure deuterium oxide, 20, 30, 40 weight percent (wt %) ethanol, and pure ethanol. Dissolution of PVAc occurs for 50, 60, 70, 80, 90, and 95 wt % ethanol-water mixtures. The lowest initial diffusion rate occurred for pure deuterium oxide, and the highest initial rate for 80 wt % ethanol. The spectral data suggests that ethanol and water form a complex that solvates PVAc more effectively than either pure liquid. Furthermore, the data indicates that sufficient quantities of this complex for effective dissolution are formed above 40 wt % ethanol, while optimum quantities occur between 70 and 90 wt % (50 to 80 mole%) ethanol.

Microscopy and Microanalysis 2002 was held in picturesque Quebec City August 5-8. The city is a lovely blend of old and new. The convention center and conference hotels were only a few blocks away from the walled old town containing many shops, cafes, bistros and historical sites. The weather was delightful – in the 70s for a high – and a real escape from the sultry heat that gripped most of the country this summer. The continental ambience made one feel as if he or she were in Paris.

Several workshops and short courses were offered on Sunday, August 4, including one by MSNO’s Valerie Woodward. Her short course was “What the Heck Happened to This?,” a compilation of tips, tricks, approaches and alternate methods for failure analysis and problem solving.

Performers from the Cirque du Soliel greeted people attending the Sunday evening reception at the Museum of Civilization. The performers mingled with the guests, who were enjoying continental beverages and hors d’oeuvres. The galleries had exhibits dedicated to the history of the region.

Monday morning the meeting began in earnest. The MSA Technical Forum (TF) Vice Chair Cathy Johnson and Valerie Woodward co-chaired a well-attended early session on polymer characterization. MSNO’s Ron Smith was one of the invited speakers at this session, and he spoke on “The Total Microscopy of a Tire.” Topics such as FIB (focused ion beam preparation), scanned probe, nanoscale technology, metallography and vascular corrosion casting were also offered later in the meeting. In the afternoon, the TF sponsored a Special Topics presentation by MSA Council member W. Gray (Jay) Jerome on Immunology 101: the Basics of Immunoglobulins and Immunostaining. A large group listened as Jay introduced immunologic concepts and presented paradigms for antigen localization and antibody staining.

The Monday sessions were capped by MSA’s Presidential Happenings. President Stan Erlandsen handed out several awards. The Distinguished Scientist Award, Biological, went to Marc Adrian for his work on cryoelectron microscopy. Ryiuchi Shimizu received the Distinguished Scientist Award, Physical, for his studies on the interaction of charged particles with solids. The Burton Medal went to Nigel Browning, who analyzes the structure-property relationships at material interfaces and defects. Tech Forum Past Chair Bev Maleeff of GlaxoSmithKline was given the Morton D. Maser Distinguished Service Award in recognition of all the work she has done on behalf of MSA. Former TF chair José Mascorro was the latest recipient of the Hildegard H. Crowley Outstanding Technologist Award in Biological Sciences for his comparison studies on epoxy resins. This year’s Professional Technical Staff Awards (PTSA), biological went to Marilyn Wadsworth, University of Vermont, and Richard Mount, Hospital for Sick Children, Toronto. Donald Robertson, University of Wisconsin, and Don Steele, Alcan International, won the PTSA, physical sciences. Congratulations to all award winners!

After the MSA awards, Colin Smith of Adobe Systems, Inc., demonstrated the use of new tools and features in PhotoShop 7.0. It was a very informative talk and gave everyone a glimpse of this major upgrade in the popular imaging program.

Tuesday featured a full day of sessions and posters, including a Problem Solving with the Experts session on Issues in Digital Imaging for the Microscopist. The afternoon included a session on Microscopy, Microanalysis and Image Analysis in the Pharmaceutical Industry. Of special interest in this session was the impact of 29 CFR Part 11 on digital asset management and data handling.

Wednesday featured the TF’s day-long symposium on Special Staining Techniques for Biological and Materials Samples. MSNO’s Ron Smith started the session with a discussion on the Staining of Polymers. Ron presented several different polymer blends and showed various tricks to visualize the structures with either osmium, THF, silver nitrate, ruthenium tetroxide, PTA and cryo-sectioning. Ron made his staining database available on diskette at the Cyber Café in the MSA Mega Booth. Eileen Boylston of USDA New Orleans discussed how to stain textiles for light and electron microscopy. Also speaking that morning were Ralph Albrecht of the University of Wisconsin, on different labeling approaches for viewing cellular antigens. Janet Woodward, from Buckman Industries of Memphis, TN, presented various ways of looking at paper composition and/or defects using stains and light microscopy to elucidate certain paper ingredients. The afternoon session featured MSNO’s Judy Drazba, Cleveland Clinic Foundation, who discussed Fluorescent Specimen Techniques for Confocal Microscopy. Judy’s Power Point presentation was jam-packed with information, and can be viewed at www.lerner.ccf.org/services/imaging.

Between the Wednesday sessions was the 20^th Anniversary TF luncheon. Each attended was presented with a luggage tag to commemorate the TF’s 20 years as the original focused interest group (FIG) within MSA. MSA past president Ron Anderson was one of the speakers, and Bev Maleeff shared a 5-minute Power point presentation summarizing the history and activities of the TF in pictures and song. TF president, MSNO’s own Jeanette Killius, was the mistress of ceremonies, and did a wonderful job. The food and décor were top-notch.

The TF business meeting was that afternoon after the Symposium, and Jeanette reported that the TF is very well-respected within MSA and is regarded as a template for how the

other FIGs should operate. Jeanette thanked everyone for his or her support in making her term a productive one. She then introduced Cathy Johnson as the 2003-2005 TF chair, and Cathy asked for input into the M&M 2003 TF program. MSNO’s Valerie Woodward is Cathy Johnson’s vice-chair.

Thursday, the TF held its annual Roundtable discussion, covering the topic of Legal and Ethical Issues of Data Ownership. Debra Sherman, Purdue University, chaired the distinguished panel that included Bertha Knoppers, an ethics attorney from Montreal, Jeff Hanson from Eli Lilly Research, Louis Sherman from Purdue University and Henry Shaffer, MSA legal counsel. The audience participated in an interactive Q&A sessions with covered such issues as micrograph ownership and copyrights, research patents, ownership of genetic databases and correct storage procedures for digital images.

MSNO member and president Bill Landis presented a platform session on Wednesday morning entitled “Vascularity of a Tissue-Engineered Model of Human Phalanges;” the work was done in collaboration with MSNO members Robin Jacquet, Jennifer Hillyer and Jeanette Killius of NEOUCOM, and others. MSNO member and trustee Lorraine Siperko presented a poster session on Monday entitled “Human Recombinant Bone Morphogenetic Protein OP-1;” her work was done in collaboration with MSNO member Bill Landis, and others.

Yes, even retirees can have a summer vacation and in my case it was a welcome respite from the August heat of low desert Arizona. In addition to experiencing Canada, I had the opportunity to make two presentations at the Quebec MSA meeting thanks to the separate invitations from Jeanette Killius (Polymer Staining) and Valerie Woodward (Tire Microscopy). The last national meeting I attended was in Cincinnati (1993) and in comparison to Quebec 2002 I see similarities and differences. Some similarities: excellent technical program, standout exhibits, smooth registration, and long lines (bordering on mayhem) for food and drink at socials. Some differences: more poster papers, a preponderance of computer presentations*, an Internet lounge (with long lines), and fewer old timers. For me, the two most lasting impressions of M&M 2002 will be the spacious and ultramodern facilities of the Quebec Convention Centre and the dramatic impact that digital processes have had in microscopy. A bonus attraction in Quebec City was the annual “Plein Art” festival that was held coincidentally with M&M – a great place to support the local economy (and I did). For my wife, Donna, the most lasting impression will be exiting Canada at the airport security checkpoint where a battery operated toy (baby shower gift) could not pass through security because there were no batteries installed to demonstrate that it was, indeed, a workable toy and not a disguised bomb.

*Observation: Not glitch free however. The counterpart glitch to upside down slides is the oft seen troublesome startup of computer presentations. That will improve with time.

MSNO TECHNICAL MEETINGS
FALL 2002 AND WINTER 2003

by Jeanette Killius and Valerie Woodward

The Fall 2002 meeting was held October 23 at Noveon, Inc. in Brecksville, OH. Ed Boyd and Valerie Woodward were the hosts for the evening. The afternoon began with an “Imaging Appetizer,” with short informational presentations about image capture devices, image processing and image output devices, by Michael Savage of Fryer Co., Luigi Armogida of B&B Microscopes Ltd., and Craig Laube of Laube Photo, respectively. Each speaker presented the high points of his topic for 20 minutes then entertained 5 minutes of questions. Michael discussed camera types, prices, and enhancers, as well as the chip logic and pros and cons of individual camera and chip designs. Luigi talked about why one needs to process images, the common image enhancement techniques (brightness, contrast, and gamma), special filters (background subtraction, max-median-min, sharpening) and protection of original images. He also supplied a handout of his presentation. Craig discussed printer types (dye sublimation, inkjet and laser), costs, resolutions needed for different applications (e.g., scanned TEM negatives vs. digital camera JPEGs), papers, and color calibration of printers. All three presenters incorporated some theory into their presentations, and provided the audience with a “grabber” for learning more about each facet of digital photomicrography.

After the imaging session, the meeting moved to the Noveon dining room for a wine and cheese social (sponsored by MSNO) and dinner. The Noveon catering service decorated the dining room in an autumn theme, and given the nearness to Halloween, added some whimsical jack-o-lanterns, ghosts, bats and spider webs to the décor. The dinner was a buffet of spring greens with various vinaigrettes, lemon chicken piccata, pasta primavera, rice pilaf, green beans and assorted pies. A high point of the evening was the drawing for two tote bags from the 2002 M&M meeting in Quebec. The winners were Bill Butler and Ed Boyd. Hmmm, two Board members – the fix must be in!

After dinner, MSNO president Bill Landis presented a very informative and illustrative talk on Laser Capture Microdissection (LCM). Bill and his group at NEOUCOM are quite active in this area, and have developed the technique for a number of applications. MSNO’s Robin Jacquet has become an expert in the technique, and many of the illustrations in the presentation were from her work. Bill told the audience about a number of uses for the method, including understanding gene expression and protosynthesis, identification of discrete cells, regions or matrices, and oncology and pathology applications. A non-cover slipped slide with a biological thin section of interest is placed on an inverted light microscope (LM) and the feature of interest (cell, region, etc.) is aligned with the laser source. A cap made from a thin proprietary polymer film is placed over the area and the laser is triggered to melt the cap to the area of interest. All features/areas outside of the apertured laser excitation area remain in the section. The cap is lifted and the cells are adhered to it. The cap is fitted on an Eppendorf tube with the appropriate reagents and the cells are removed for further analysis. Prior to fitting to the Eppendorf, the bottom of the film can be photographed to show plucked cells. This method can also be used directly on the cut face of a sample block, and then a section can be cut with the cryomicrotome. The film/thin section buildup can be transferred to a glass slide, placed on the LM, and the area/feature of interest can be plucked out. A specific example of the group’s work was the role of osteopontin in bone formation. Whether its role is facilitative or inhibitory is unclear. The osteopontin binds to hydroxyapatite crystals, which is relatively easy to locate in prepared sections. Cells can be specifically plucked from different cartilage regions of the epiphysical plate – resting, proliferative, hypertrophic, calcified, etc., and the amount of osteopontin in those cells can be quantified. This technique can be used to capture a single cell or successively “laser stick” up to a 6-mm diameter circle filled with cells. The group found that between 1 and 5 days of development, there was very little mineralization, and after 5 days and up the mineralization increased; as mineralization increased, the hypertrophic cells had less osteopontin. Another use of the method in Bill’s group is to isolate cells for quantitative gene expression. Bill’s images were spectacular, and generated a large amount of interest from the group. Bill noted that there are also systems available for the same type of feature isolation for materials applications – we decided that materials LCM would be an excellent topic for a future meeting.

The Winter 2003 meeting was held February 18 at the Lerner Research Institute of the Cleveland Clinic Foundation, and was hosted by Judy Drazba and her colleagues and staff of the Core Imaging Facility. Judy and Jay Mansbach, MSNO trustee, were the organizers of the meeting and handled all of the arrangements. The topic of the meeting was “Imaging – the Main Course” and was a more in-depth continuation of the Fall meeting “appetizer.” David Smith of Fryer Co. was the first presenter of the meeting, and his topic was Image Acquisition. David stated emphatically that his only text reference was John Russ’ “Image Analysis Handbook.” Of note, John Russ is the developer and primary instructor of the very popular and highly respected quantitative image analysis classes held at North Carolina State University’s College of Continuing Education. David opened with why we do image acquisition (for better definition of contrasting areas, increase precision and accuracy in measurement, reproducibility of results, higher throughput than manual methods), the hierarchy (eyes, simple microscopes, compound optical scopes, EMs, AFMs), and the steps involved (acquisition, processing, measuring and results). He emphasized that sample preparation is the most important step in obtaining a good image, period. Microscope adjustments such as alignments, objective n.a., etc., and illumination adjustments are the next most important steps, camera adjustments the next important, and post-acquisition processing the least important. He discussed the topic of spatial resolution, and offered that many users do not truly understand the concept. He used the example of a 10X objective with 0.45 n.a.; the resolution is 0.677 um, the field is 1000 um and the scope resolution is 1477 units that one can observe, so why do you need 3 megapixels? David then discussed in great detail the types of cameras - color for qual work, monochrome (more appropriate for quant work), video and digital – and the types of chips in each. For quantitative use, the B&W chip specifications of importance are pixel size, well depth (how many electrons fit into a pixel on the chip), quantum efficiency, linearity of response, noise, chip size, and speed of readout. He pointed out that there are three main attributes of chips, resolution, speed and sensitivity, and you can’t have all three. Sort of reminded us of the mantra of the analytical service lab – results can be cheap, fast and good – which two do you want? He also discussed how CCD chips work and their types (full frame, frame transfer and interline transfer) advantages/disadvantages, noise sources, speed optimization, and how cooling can greatly facilitate noise reduction and efficient low light image capture. Finally, he recommended that potential users try to match the camera to the application by considering the following camera parameters: spatial resolution, temporal resolution, dynamic range and linearity, low light sensitivity and adaptation to instruments. This presentation was loaded with useful, practical information, and I couldn’t write fast enough to capture it all!

After Dr. Smith’s talk, the group adjourned to the Lerner Pavilion atrium for a lovely social hour. The drinks and appetizers were provided by the Core Imaging Facility, and included a beautiful fresh fruit platter, vegetables and dip, crackers and cheese, hot hors d’oeuvres and Judy’s delicious home made cheese dip! The decorations were individual potted primroses – a welcome colorful sight for a cold winter evening. During the social hour, Judy and her colleagues led guided tours of the Core Imaging Facility. It was a lively social hour, and there were guests who were attending their first MSNO meeting, so the “veterans” were actively encouraging these guests to join us for future meetings and hopefully, membership! Dinner followed, a catered buffet with garden salad, chicken Marsala, vegetable lasagna, garlic mashed potatoes, roasted vegetables, and a scrumptious assortment of pastries and brownies for dessert. The table service and decorations were a striking combination of teal green, black and white. The dinner conversations were lively and varied, and one of the main topics was how tired we all were of the very cold weather. Thanks go to Kate Mansbach, who helped with serving at the social hour and dinner. We think she should be an honorary member (hint, hint, Jay)!

After dinner, Luigi Armogida of B&B Microscopes, Ltd. reprised his October 2002 talk on image processing. He started out by relating image processing to capture devices – one can process in the preview window, at acquisition, and post-capture. He asked why do we need to process – and the answers included to accurately document samples, enhance trends found in images that are often overlooked, and to prepare images for other analyses. Luigi is an advocate of “power processing,” that is putting together combinations of many different processing steps to obtain a single well-focused, well-resolved, meaningful image. These combinations can involve the simple operations (brightness-contrast-gamma or sharpen-blur), more complicated ones (special filters) or very complicated ones (Fourier transforms, power spectra, Boolean operators). Many times, power processing will involve all levels of these operations. Luigi warned that it is important to preserve the original image (in case you have gotten so far gone from it that you can’t get back), so it is better to operate on copies of the image and record the processing steps (most software programs now keep track of the steps automatically). His warning also covered the ethics of changing or deleting original data, and how more and more agencies and facilities are making preservation of the original image mandatory. Color detection and discernment is still the domain of the human eye – it is better than any camera system. He described the accepted color spaces, CMYK (cyan-magenta-yellow-black for print processes), RGB (red-green-blue), HSI (hue-saturation-intensity), HSV (hue-saturation-value) and HSL (hue-saturation-luminance), and discussed the most accurate color space CIELAB, which is the best for textiles and printing. Luigi also gave us some practical take-home tips such as stretching your image histogram to enhance contrast, using Fourier transform filters to remove certain types of noise such as scan lines, and applying pseudocolor to an image to locate variations in gray levels due to hot spots or uneven illumination. As in David Smith’s talk, there was so much information that writing notes couldn’t do it justice. MSNO was truly fortunate to have two such dynamic and knowledgeable speakers for this meeting.