March 24 – 26 at OHSU, Portland, Oregon

2011 Northwest Regional Cytometry Meeting

Governor Tom McCall Waterfront Park from the Steel bridge

Cytometry Data: Mining, Modeling, and ManagementData analysis is the bottleneck in cytometry. Analytical approaches useful in low parameter phenotype experiments are too slow and biased for abstracting features in highly multivariate functional experiments, such as cytokine and phosphorylation assays. Automation and standardization are essential for valid measurements, especially for functional assays and across platforms. New bio-informatic tools for warehouse and mining, extracting features, modeling, simulating, and visualizing results, are needed if phenotypic and functional applications are to develop as expected, and if cytometric experiments done with a variety of platforms and approaches are to contribute to a unified systems view of cells and tissues. Please join us in the conversation.

J. Paul Robinson, “Coordinated parallel systems approach to high content/high throughput flow cytometry drug screening”

Garry Nolan, “Systems immunology in the post-fluorescence era”

Maria Jaimes, “Quality assurance and variability in intracellular staining”

Ger van den Engh, “Parsing cytometric noise” Ryan Brinkman, “Automated gating and analysis: State of the art and prospects”

Norman Dovichi, “Chemical and metabolic cytometry” Manfred Auer, “Imaging biological function across scales: From macro-molecules to cells, tissues, and microbial communities”

Bob Zigon, “Architectures for real time processing and mining of flow cytometry data”

C. Bruce Bagwell, “Probability State Modeling for automated high-dimensional cytometry”Dave Novo, “Consolidated software for flow and image cytometry”

Masanobu Yamamoto, “50 years of laser technology: Innovating flow cytometry with Blu-ray technology”

Holden Maecker, “Standardization and validation of fluorescence and mass cytometry panels”

ISAC is the premier society for development and application of hardware and software for flow and image cytometry (http://isac-net.org).

Gregory Kaduchak, “Acoustic focusing flow cytometry”

Bill Ortyn, “High parameter morphological classification replicates the sensitivity and specificity of immunofluorescence labeling in whole blood samples”

Joe Gray, “Spatial systems biology – the central role of cytometry in the interpretation of the genome”

Prospectus and talks

2011 Northwest Regional Cytometry Meeting

Downtown from across the Willamette river

The 2011 Flow Informatics and Computational Cytometry Society (FICCS7) meeting is consolidated with the 2011 NWRCM. FICCS7 sessions will be Thursday, March 24, from 8:30am – 3:00pm. For information on the FICCS meeting, contact Ryan Brinkman at info@FICCS.org. A FloCyte course, “Data Analysis”, taught by Dr. Tim Bushnell, is concurrent with the FICCS7 meeting. Contact Sue DeMaggio at FloCyte@FloCyte.com for more information. The first session of the Northwest Regional Cytometry Meeting (NWRCM) begins at 3:00pm on the 24th. Reception dinner is at 7:00pm in the Old Library. The Friday session of plenary talks begins at 8:00am and continues to 6:30pm, with a social night on the town following. The Saturday program includes a number of software practicums (details on page 5). In addition, a cytometry program for high school teachers begins at 10:00am, as does the FloCyte course on standardization. All talks are free and open to the public – the $25 registration fee, helping defray the cost of food, is the meal ticket for the Thursday evening buffet and Friday lunch. For registration, go to www.ohsu.edu/ohrc , 2011 meeting caption to the left. For more information, contact Allan Kachelmeier at kachelme@ohsu.edu or 503-494-2373.

Thursday, March 24FloCyte course, “Data Analysis for flow”, taught by Dr. Tim Bushnell, is from 8am – 3pm. A 10% discount is offered to timely registrants of the 2011 NWRCM (discounted before March 19). For FloCyte registration, go to www.FloCyte.com . The FICCS7 meeting (consolidated with the 2011 NWRCM) is from 9am – 3pm. Contact Ryan Brinkman at info@FICCS.org for more details.

Garry Nolan, Professor, Baxter Laboratory in Stem Cell Biology, Department of Microbiology and Immunology, Stanford University, “Systems immunology in the post-fluorescence era” Norman Dovichi, Grace-Rupley Professor of Chemistry and Biochemistry, University of Notre Dame, “Chemical and metabolic cytometry” Manfred Auer, Staff Scientist, Lawrence Berkeley National Lab, “Imaging biological function across scales: From macromolecules to cells, tissues, and microbial communities”

Reception dinner in the Old Library at 7:00pm. Dinner address by Joe Gray, Gordon Moore Professor of Biomedical Engineering, OHSU, “Spatial systems biology –the central role of cytometry in the interpretation of the genome”

Masanobu Yamamoto, Chief Technical Communications Advisor, Sony Life Electronics, “50 years of laser technology: Innovating flow cytometry with Blu-ray technology”Gregory Kaduchak, Director of Engineering, Life Technologies, “Acoustic focusing flow cytometry”

Schedule

FloCyte courses:Data analysis for flowStandardization

FICCS7 meeting

NWRCM itinerary:New directionsShowcased technologyReception dinnerApplicationsQuality controlData processingDiscussion

Cytometry for teachersPracticums

March 24 – 26, OHSU Old Library, Portland

Ger van den Engh, VP, BD Biosciences, “Parsing cytometric noise”

Maria Jaimes, MD, Senior Scientist, BD Biosciences ,“Quality assurance and variability in intracellular staining”Ryan Brinkman, Senior Scientist, Terry Fox Laboratory, and Associate Professor, Medical Genetics, University of British Columbia, “Automated gating and analysis: State of the art and prospects”

Friday, March 25 Schedule

Bob Zigon, Software Lead for Flow Cytometry Analysis, Beckman Coulter, “Architectures for real time processing and mining of flow cytometry data“

Coffee break and concurrent technology presentations (see the listing of vendor talks on page 6)

Lunch-time panel discussion, “Cytometry informatics in 2020”, chaired by Janet Siebert, with panelists including C. Bruce Bagwell, Adam Triester, Julie Hill, Nikesh Kotecha, and Ed Walker.

Coffee break and concurrent technology presentations (see page 11)

Dave Novo, PhD, President , De Novo Software, “Consolidated software for flow and image cytometry”

Holden Maecker, Phd, Director, Human Immune Monitoring Center (HIMC), Stanford University, “Standardization and validation of fluorescence and mass cytometrypanels”

Saturday, March 26 Software practicums and demos (see page 5)Cytometry for high school teachers led by Julie Hill The FloCyte course, “Standardization, validation, characterization, and controls”, taught by Dr. Tim Bushnell, is from 10am – 5pm. A 10% discount is offered to timely registrants of the 2011 NWRCM (discounted before March 19). For FloCyte registration, go to www.FloCyte.com .

Friday afternoon finger food with support from Life Technologies and Beckman Coulter.

C. Bruce Bagwell, MD, PhD, President, Verity Software House, Inc, “Probability State Modeling for automated high-dimensional cytometry”

Impressionistic image obtained by selective filtering

J. Paul Robinson, SVM Professor of Cytomics, School of Veterinary Medicine, and Professor, Weldon School of Biomedical Engineering, Purdue University, “Coordinated parallel systems approach to high content/high throughput flow cytometry drug screening”

William Ortyn, President, COO, and co-founder of Amnis Corporation, “High parameter morphological classification replicates the sensitivity and specificity of immunofluorescence labeling in whole blood samples”

Friday evening fun with support from Tree Star.

Software practicums

FloJo for Programmers

FlowJo for programmers

2011 NWRCM courses and practicumsData analysis for flowStandarization, validation, and controls

Analysis with Gemstone

Cytometry workshop for teachers

“FlowJo for Integrators”, taught by Adam Treister, on March 26, 10am – 3pm. FlowJo is widely used in research labs, but too rarely integrated with servers and other software to create an enterprise level analysis infrastructure. The next level of improvement comes from integrating FlowJo with other best-of-breed applications, and developers at Tree Star have been building in the hooks to support it. The curriculum, geared to the needs of participants, will include discussion of using FlowJo with server based repositories, building a multiple server environment, integration of FlowJo with R programs, turning external classification into gates, hands free compensation, and command line arguments. This class is Tree Star's first formal presentation of this information. For registration, contact Tree Star directly at flowjo@treestar.com .

“Data analysis for flow”, taught by Tim Bushnell, on March 24, 8am – 3pm. For FloCyte registration, go to www.FloCyte.com .

“Analysis of high dimensional data in Gemstone”, presented by C. Bruce Bagwell on March 26*. Large listmode files tax current analysis methods. Bruce is conducting an interactive session to demonstrate the utility of Probability State Modeling in Gemstone for analyzing phenotype/functional signatures in a 16 parameter dataset acquired by M. Inokuma at BD..

“Standardization, validation, characterization, and controls”, taught by Tim Bushnell, on March 26, 10am – 5pm. This FloCyte course is aimed at a more experienced user. Continuing education contact hours available for CLSs. For FloCyte registration, go to www.FloCyte.com . “Cytometry workshop for middle and high school teachers”, facilitated by Julie Hill, on March 26, 10am – 3pm. Attendees will learn some of the basics of flow cytometry, and why flow cytometry is so important in research and clinical diagnosis. The program will include lecture, hands-on activities, small groups break out session, and take-home materials. To request a registration form or for questions, email nwrcytometrym@gmail.com.

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“Management and analysis of flow cytometry based signaling assays using Cytobank”, presented by Nikesh Kotecha, on March 26*. Cytobank is a cloud-computing platform for managing, sharing, and analyzing flow cytometry data from anywhere. The approach grew out of work in Dr. Garry Nolan’s lab at Stanford in handling large flow experiments such as from phosphoproteinprofiling. The session will include discussion of PermKey, a new BD resource powered by Cytobankto facilitate the design of multi-parameter staining panels for analysis of intracellular and cell surface proteins. “Innovative approaches to visualizing and analyzing multivariate data using InCyte software“, presented by Ray Lefebvre on March 26*. Though automated flow instruments make it easy to acquire large datasets, data analysis is a weak link. Needed are tools to interactively explore and visualize large datasets. EMD Millipore's new InCyte software provides interactive tools in an easy-to-use drag-and-drop format. InCyte can apply the analysis automatically to a large set of runs without the need for complicated batch files or scripts. The visualization tools include provisions for multi-plate heatmaps and IC-50 curve analysis.

*Unspecified times will be determined on the basis of interest shown in registration preferences. Every effort will be made to maximize access and minimize overlap. Most of these sessions will begin at 8am. The registration packet will have the finalized details.

Analysis of signaling assays with Cytobank

FloCyte courses

Vendor practicums and demos

“Interactive session on image cytometry analysis”, facilitated by Dave Novo on March 26*. Extra computers and demo datasets to download on your own labtop provided.

Image cytometry analysis in FCS ExpressInteractive tools in InCyteStandardizing fluorescence on an Accuri

“Standardizing fluorescence on an Accuri: Theory and practical applications”, presented by Clare Rogers and Maria Dinkelmann on March 26*. What does it mean to “standardize fluorescence”? Why is it useful? Standardization simplifies experiment to experiment comparison, antibody titration, and quantification of epitope density.“Kaluza: Exploring new dimensions in flow cytometry analysis”, presented by Bob Zigon on March 26*.

Analysis with Kaluza

Stratedigm, Inc.: Shervin Javadi, CEO, “Performing multi-color analysis using Stratedigm’s S1000Ex analyzer with ‘Smart Detect’ technology”

BD Biosciences: Joe Trotter, Director of Cytometry, Advanced Technology Group, “New Influx software: Fundamentally changing how software supports cell sorting” and presenter TBA, “BD PermKey: A compendium of results for guiding the design of multiplexed intracellular protein and surface marker staining for flow cytometry”

Union Biometrica, Inc.: Nathan Portier, Applications Scientist, “BioSorter™: Flow cytometry for large objects”

Accuri Cytometers: Clare Rogers, Marketing Manager, “Simplifying fluorescence quantification by standardizing fluorescence detection: A new approach using the Accuri C6 flow cytometer”

BioLegend: Lani Duenas, “A new dye from BioLegend”

Amnis, Inc.: Bill Ortyn President, COO, and co-founder, “Advances in applications for imaging flow cytometry”

Bay Bioscience: Jeff Harvey, “New from Bay Bioscience, US: JSAN Cell Sorter update and advances in platelet analysis with the Apogee A50 Analyzer”

Automation Lab Technologies: see table display

Beckman Coulter: Chris Novak, Strategic Marketing Manager, “Astrios: A new frontier in cell sorting”

DeNovo Software: Dave Novo, President, “What’s new in FCS Express 4?”eBioscience: Colin Fitzpatrick, Account Manager, “FlowCytomix™ - Multiple analyte detection on the flow cytometer”

Cytobank: Nikesh Kotecha, President, “Cytobank: A Web-based approach for analysis and publication of flow cytometry experiments”

Intellicyt: Kim Luu, PhD, Application Sciences, “Validation of a 96-well high-throughput flow cytometry (HTFC) screening assay for simultaneously measuring five indicators of apoptosis using the HTFC™ screening system”

Labkey Software: Kevin Krouse, Developer, “Reagent database”Life Technologies: Presenter TBA, “Benchtop instruments from Life Technologies to complement your work-FLOW”Millipore: Ray Lefebvre, “Innovative approaches to analyzing and visualizing multivariate data using InCyte software”Miltenyi Biotec: TBAPartec: see table display

Sony Life Electronics: Matt Alexander, Senior Cytometry Systems Specialist,“4 lasers, plate loader, and electronic volume: The new Eclipse is an affordable cytometer for DNA, fluorescent proteins, and immunophenotyping”Stem Cell Technologies: see table display

Tree Star: Jay Almarode, “Meta-analysis: How good are your gates? A look at new FlowJo metrics for analyzing the quality of human or algorithmic gating operations”

Verity Software House: Mark Munson, Technical Sales, “An update on GemStone: A guided journey through multidimensional space"

Propel Labs: see table display

2011 Northwest Regional Cytometry Meeting

Vendor talks

Woodside Logic: TBA

Speakers

Garry Nolan, PhD, Professor, Baxter Laboratory in Stem Cell Biology, Department of Microbiology and Immunology, Stanford University. Dr. Nolan received his PhD under the mentorship of Leonard and Leonore Herzenberg, and later did postdoctoral work under David Baltimore at MIT and Rockefeller, work in which he cloned and characterized the p65 subunit of NF-kB. In recent work at Stanford the Nolan lab has spearheaded flow cytometric analysis of phosphoproteins in single cells and now application of mass spectrometry at the single cell level (CyTOF), which, coupled with high throughput proteomics and Bayesian statistics, has facilitated the understanding of signaling pathways in normal and diseased immune cells at a systems immunology level. Clinical development of diagnostics is one of the goals of the research.

J. Paul Robinson, PhD, SVM Professor of Cytomics, School of Veterinary Medicine, and Professor, Weldon School of Biomedical Engineering, Purdue University. Dr. Robinson is Director of the Purdue University Cytometry Laboratories and Deputy Director for Cytomicsand Imaging in the Bindley Biosciences Center. His research is focused on reactive oxygen species and in studying the biochemical pathways of apoptosis and ROS in mitochondria. His bioengineering group developed the first high speed multispectral cytometer, tools for quantitative fluorescence measurement, and advanced classification approaches for clinical diagnostics and bacterial classification. Dr. Robinson was one of the first scientists to engage the internet when he established the PUCL website in 1993, now at more than 10 million hits in a year, and also broke new ground in publishing web-CDROMs in 1996. He is past president of the International Society for Analytical Cytometry, Editor-in-Chief of Current Protocols in Cytometry, and a fellow of the American Institute for Medical and Biological Engineering.

Norm Dovichi, PhD, Grace-Rupley Professor of Chemistry and Biochemistry at the University of Notre Dame. Dr. Dovichi is the only chemist on a list of a dozen “Unsung Heroes of the Human Genome Project” compiled by the journal Science. His group was instrumental in developing the capillary array electrophoresis with sheath flow cuvette based laser induced fluorescence detection that was the primary analytical instrument used in sequencing the genome. More recent work focuses on developing tools to study the proteome and metabolome using similar chemical cytometrytechniques. The long term goal is study of protein expression in single cells concomitant with changes in physiology. His work has been honored with the Chemical Instrumentation Award and Spectrochemical Analysis Award from the American Chemical Society, McBryde, Noranda, and Fisher Awards from the Canadian Institute of Chemistry, and Heinrich Emanuel Merck Award for Analytical Chemistry. He serves as Associate Editor for Analytical Chemistry.

“In under a year, genomics technologies will enable individual laboratories to generate terabyte or even petabyte scales of data at a reasonable cost. However, the computational infrastructure that is required to maintain and process these large-scale data sets, and to integrate them with other large-scale sets, is typically beyond the reach of small laboratories and is increasingly posing challenges even for large institutes.” - Schadt, Linderman, Sorenson, Lee, and Nolan (2010), Nature Reviews | Genetics 11:647-657.

Manfred Auer, PhD, Staff Scientist, Lawrence Berkeley National Lab. The Auer lab focuses on visualizing molecular machines at molecular resolution, and identifying protein composition through novel labeling approaches. 3D electron tomography is a major tool used for the analysis of the machines in their native cellular environment, complemented by correlative biochemical and biophysical techniques, and sophisticated visualization, segmentation, and quantitative analysis.

Auer M., Scarborough G. & Kühlbrandt W. (1998) “Three-dimensional Map of the Plasma Membrane H+-ATPase in the Open Conformation”, Nature 392: 840-843. Bajaj C, Yu Z., Auer M. (2003) “Feature extraction, analysis and visualization of tomographic molecular imaging”, J Struct Biol144:132-143. Auer M, Koster AJ, Ziese U, Volkmann N, Bajaj C, Wang DN, Hudspeth AJ (2008) “3D architecture of hair cell stereociliar extracellular links as revealed by electron tomography”, JARO 9:215-224.

Masanobu Yamamoto, Chief Technical Communications Officer, Sony Life Electronics. Mr. Yamamoto has been involved in the development of optical disc storage from the beginning in the 1970s. He subsequently worked on CDs, and on higher density storage like WORM and MO. Since 1996 he has been leading the development of a recording technology based on shorter wavelength lasers and higher NA optics. The result of this development, Blu-ray technology, is now being applied to flow cytometry. Sony acquired iCyt Mission Technology in early 2010, and Mr. Yamamoto is now working in Champaign, IL on applications to flow.

Gregory Kaduchak, PhD, Director of Engineering, Life Technologies. Greg obtained his PhD in Physics from Washington State University where he studied elastic structural responses. He joined Los Alamos National Laboratory in the late 1990s as a staff scientist and project leader working on biological and chemical detection systems. While at Los Alamos he formed collaborations with the resident flow cytometry teams, including Dr. Gary Salzman, and subsequently developed devices to manipulate cells in solution using acoustic forces. Greg joined Life Technologies in 2008, where he has been instrumental in developing the Attune™ acoustic focusing cytometer. In the Attune, hydrodynamic focusing is replaced with forces from acoustic radiation pressure, with the advantage the flow can be varied to control interrogation time and volumetric rate. In his talk, acoustic focusing will be developed from fundamental principles, and will include specific details of implementation.

Goddard G, Martin J, Graves S, Kaduchak G (2006), “Ultrasonic particle-concentration for sheathless focusing of particles for analysis in a flow cytotometer”, Cytometry 69A: 66 – 74.

Joe W. Gray, Gordon Moore Professor of Biomedical Engineering, and, Director, Center for Spatial Systems Biomedicine, OHSU. Dr. Gray comes to OHSU from his position as Director of the Division of Life Sciences at Lawrence Berkeley National Lab. The Gray Laboratory explores mechanisms by which genomic, transcriptional, and proteomic abnormalities occur in selected cancers, with particular focus on how these abnormalities contribute to cancer pathology and influence response to gene targeted therapy. Aspects of his research include integrated analyses of the spectrum of recurrent abnormalities that determine cancer behavior, mathematical models that describe how cancer-associated molecular abnormalities influence individual responses to therapeutic inhibitors, novel therapeutic approaches to treat breast or ovarian cancer subpopulations that do not respond well to current aggressive chemotherapeutic strategies, proteomic strategies for early detection of breast cancer related proteins in blood, automated functional assessment of genes deregulated by genomic abnormalities in cancers, and molecular imaging for early detection of metastasis prone breast cancer.

Ger van den Engh, PhD, Vice President, Advanced Cytometry Cell Analysis, Becton Dickinson Biosciences. Ger has a broad background including Director of the Flow Cytometry Core, Lawrence Livermore National Laboratory, various professorships at the University of Washington including in Bioengineering, Molecular Biotechnology, Immunology, Genome Sciences, and Oceanography , founding member and professor in the Institute for Systems Biology, founder and president of Cytopeia, and his present position at BD. While best known for his development of a high-speed cell sorter for the analysis and purification of human cells and chromosomes, there is a strain of ‘numismatic gyrations’ in his interest, peregrinations in karyotyping and equipment informed by a technician’s love of electronics and a physicist’s love of random walks. He has more than 20 patents relating to flow cytometry and cell sorting. Awards include Fellow of the American Institute for Medical and Biological Engineering and Howe Fellow/Becton Dickinson.

van den Engh, G., Nelson, P., Roach, J. (2000), “Numismatic gyrations”, Nature 408:540

Ryan Brinkman, PhD, Senior Scientist, Terry Fox Laboratory, and Associate Professor, Medical Genetics, University of British Columbia. Dr. Brinkman's research group is focused on developing flow cytometry automated analysis methods and applying them to large datasets. As proponent s of open source software, the high throughput tools his group have developed for quality checking, data normalization, and automated clustering are freely available as both R/Bioconductor packages and GenePattern modules . He also chairs the International Society for Advancement of Cytometry Data Standard's Task Force, and, in his words, is largely to blame for FCS3.1.

Nima Aghaeepour, Radina Nikolic, Holger H. Hoos, Ryan R. Brinkman, “Rapid Cell Population Identication in Flow Cytometry Data”, Cytometry A (In Press)Habil Zare, Parisa Shooshtari, Arvind Gupta and Ryan R Brinkman (2010), “Data reduction for spectral clustering to analyze high throughput flow cytometry data”, BMC Bioinformatics 2010, 11:403

Maria Jaimes, MD, Senior Scientist, BD Biosciences. Dr. Jaimes, a Senior Scientist working at BD Biosciences in the Reagent Application Group, manages the NIH/NIAID/DAIDS Intra-cellular Cytokine Staining (ICS) Quality Assurance Program. The goal of the program is to assess inter-laboratory assay variation among multiple laboratories testing HIV vaccine candidates. Dr. Jaimes also has been a consultant for other ICS Proficiency Testing Programs including the ones developed by the Cancer Vaccine Consortium, the Foundation for the NIH, and the ANRS (National Agency for Research in AIDS, France).

Jaimes MC, Maecker HT, Yan M, Maino VC, Hanley MB, Greer A, Darden JM, D'Souza MP (2010), “Quality assurance of intracellular cytokine staining assays: Analysis of multiple rounds of proficiency testing”, J Immunol Methods (In Press).

Holden Maecker, PhD, Director, Human Immune Monitoring Center (HIMC), Stanford University, and a consultant in flow cytometry. His research focuses on cellular immune response to chronic pathogens and cancer, and the correlation of the response to immune protection. Dr. Maeckerreceived his Ph.D. from Stanford University, where he also did his postdoctoral work. He previously held positions at Loyola University, Chicago and with BD Biosciences, San Jose, CA. He participates in several international groups related to immune monitoring. The HIMC has developed standardized immunophenotyping panels, using lyophilized reagent cocktails, and phospho-epitope flow cytometry panels, using fluorescence barcoding. The center is now developing similar panels for mass cytometry using CyTOF (Cytometry by Time Of Flight mass spectrometry).

C. Bruce Bagwell, MD, PhD, President, Verity Software House, Inc. Cytometry data is traditionally displayed as a one or two dimensional histogram of intensity, a graphical format adequate in low dimension for revealing relationships, but one that doesn’t clarify relationships at high dimension. New graphical formats are needed to properly visualize multi-parameter data and automate data analysis. Probability State Modeling (PSM) is a parallel coordinate like format for visualizing and analyzing listmode data, which, because it shows intensity, percentage, and variance information on individual phenotypes, is particularly appropriate for displaying the interplay of phenotypic changes as cells differentiate. The discussion of this format will open the larger one of graphic display, and how the organization of the graphic medium can be used to wring more intuitive meaning from high dimension data. Bruce has a strong interest in promoting and teaching cytometry. He has been involved with the Annual Course in Cytometry at Bowdoin for over 30 years. Bruce’s contributions to cytometry include the software compensation now used in virtually all cytometers, broadened rectangle and trapezoid DNA models, and probability state modeling.

William Ortyn, President ,COO, and Co-founder of Amnis Corporation. In the US and Europe approximately 25% of all samples analyzed by mainline hematology systems are designated as “Not Within Normal Limits” (NWNL). These samples need further review and are subsequently analyzed by either flow cytometric immunofluorescent labeling or manually by human review after a slide preparation has been made. Human review is subject to the variability and skill of the operator, while immunofluorescent labeling requires expensive antibodies. Multiparametric Imaging Flow Cytometry (MIFC) provides the speed, imagery, and feature set in the context of a fluid sample to perform automated, highly parametric,morphological classification of whole blood samples without slide preparation, immunofluorescent labeling, or primary human review. In this study whole blood was immunofluorescently labeled to identify cell types to form a training set. An automated classification methodology was employed in 1000 dimensional feature space using IDEAS cell analysis software to train classifiers using features from brightfield, darkfield, and nuclear image channels to identify five major leukocyte populations in the blood. The trained classifiers were then compared to immunofluorescent labeling on numerous donor samples run on the ImageStreamX to determine the sensitivity and specificity performance of the non-antibody, morphological approach. In this talk he will discuss the methodology and results of this work.

Bob Zigon, Software Technical Lead for Flow Cytometry Analysis, Beckman Coulter. Bob is a graduate of Purdue with degrees in Computer Science and Mathematics. He is the technical lead responsible for developing Kaluza. His interests include high performance computing, numerical analysis, and information retrieval theory. He has been a member of the ISAC Data Standards Committee and Data Standards Task force for 4 years. Bob has a strong interest in developing data visualization and analysis techniques for flow cytometry.

Dave Novo, PhD, President , De Novo Software. David began selling his flow cytometry software, FCS Express, while still in graduate school at UCLA obtaining his PhD in Biophysics. He has recently expanded the scope of the FCS Express to include image analysis, responding to the explosion in image cytometry from improved, more cost effective imaging technology. Concurrent with the rise in automated imaging is the expanded use of mathematical analysis on cell images to generate multi-parameter data . The current state of image cytometry, however, mirrors the state of flow cytometry in the early 1980’s with many different manufacturers producing similar types of data but saving the data in proprietary and incompatible formats. The FCS Standard encouraged growth of a healthy 3rd party flow cytometry data analysis market, and was a critical element in the success of flow cytometry. David will discuss progress towards a new image cytometry data standard, as well as describe key elements and challenges in developing robust image cytometry data analysis software.

Janet Siebert, President, CytoAnalytics. Janet is a computer scientist who believes that good data analysts care about the stories in the data. Discovery happens when life scientists and computer scientists are able to have lively and informed conversations about possible stories in complex data sets. Janet’s work focuses on analyzing and presenting multidimensional life science data using methods that support these conversations.

Siebert JC, Wang L, Haley DP, Romer A, Zheng B, Munsil W, Gregory KW, and Walker EB (2010), “Exhaustive expansion: A novel technique for analyzing complex data generated by higher-order polychromatic flow cytometry experiments”, Journal of Translational Medicine 8: 106. Siebert JC, Inokuma M, Waid DM, Pennock ND, Vaitaitis GM, Disis ML, Dunne JF, Wagner DH, and Maecker HT (2008), “An analytical workflow for investigating cytokine profiles”, CytometryA 73:4.

Adam Treister, President, Tree Star. Adam is the primary author of FlowJo analysis software. He has a BA in Fine Arts from Amherst College, and a MS in Electrical and Computer Engineering from UC Santa Barbara. His work as a scientific programmer at Stanford University led him to the Herzenberg laboratory where he picked up the thread of cytometry analysis and created FlowJo. He has been a member of the ISAC Data Standards Committee and Data Standards Task Force for fifteen years, and is very interested in developing the protocols necessary to describe and rationalize cytometric analysis.

Nikesh Kotecha, PhD. President, Cytobank, Inc. Dr. Kotecha develops informatics approaches to incorporating molecular technologies in translational research. His current focus is developing infrastructure and analysis methods for signaling-based flow cytometry assays, including a diagnostic for juvenile myelomonocytic leukemia (Cancer Cell 2008, cover) and development of Cytobank, a web-based resource for managing, sharing, and analyzing flow cytometry data. Cytobank provides a platform for delivering novel analyses and visualizations – components he believes to be crucial for the next generation of flow cytometry. Dr. Kotecha’s experience includes a PhD in Biomedical Informatics at Stanford University (laboratory of Dr. Garry Nolan) and work at TIBCO Spotfire Inc. and Wyeth Ayerst Inc. developing software applications for visualization and analysis of various types of data in the drug development pipeline.

Kotecha N, Krutzik PO, Irish JM (2010), “Web-based analysis and publication of flow cytometry experiments”, Current Protocols in Cytometry”, 53:10.17.1–10.17.24. PMID: 20578106

Friday noon panel discussion, “Cytometry informatics in 2020”

Juliane Hill, Scientist II, Analytical Development, Dendreon. Julie has extensive experience in clinical, pre-clinical, and research flow cytometry. She sees first hand and on a daily basis the need for efficient and flexible data management tools that allow diverse types of data to be analyzed in a comprehensive way. Julie spearheads local continuing cytometry education in the Seattle area, and is currently working to bring cytometry education to K-12 and community college classrooms.

Edwin Walker, PhD, Director, Laboratory of Immunological Monitoring, Earl A. Chiles Research Institute, Robert W. Franz Cancer Research Center, Providence Portland Medical Center. Dr. Walker’s research has long involved use of cutting-edge polychromatic flow cytometry tools to analyze the phenotype and functional properties of long-term self-tumor antigen-specific effector and memory T cells in cancer patients receiving different immunotherapy regimens. His laboratory studies the tolerogenic mechanisms which regulate differentiation and maintenance of tumor-specific memory T cells. The laboratory is known to push the envelope on data analysis and visualization tools, for example in their innovative use of heat maps, to facilitate better understanding of high parameter data.

FICCS7 + NWRCM bioinformatics - genomic and flow sessions

Obi Griffith, PhD, CIHR Postdoctoral fellow, Cancer & DNA Damage Responses, Lawrence Berkeley National Laboratory. Dr. Griffith completed his PhD at the University of British Columbia and with Steven Jones at BCGSC in the field of bioinformatics and clinical statistics, with research focusing on gene regulation and expression. He developed the ORegAnno database (www.oreganno.org) for community-driven curation and collection of gene regulatory sequences, developed novel methods for cross-platform integration, meta-analysis, and subspace clustering of large-scale gene expression data, and worked in collaboration with Sam Wiseman to identify new diagnostic and prognostic signatures for thyroid cancer from tissue microarray data. He recently joined Joe Gray's lab where his research focuses on analysis of next-generation sequencing data and clinical statistics in breast cancer, with the goal of identifying -omic predictors of drug response in breast cancer.

Laura Heiser, PhD, Scientist, Lawrence Berkeley National Lab. Laura earned her Ph.D. in Neuroscience from the University of Pittsburgh in 2004. She completed post-doctoral studies under the direction of Paul Spellman at Lawrence Berkeley National Laboratory, where she developed algorithms to analyze large genomic and proteomic datasets and integrate them into a computational model of deregulated cell signaling in breast cancer. Her research focuses on the genomic and proteomic changes that drive oncogenicbehavior in breast cancer, with the goal of identifying aberrations associated with therapeutic response.

Robert Searles, PhD, Director, OHSU Massively Parallel Sequencing Shared Resource, and Associate Director, OHSU Integrated Genomics Laboratory. Bob has degrees in biochemistry and biophysics and has worked for 20 years in gene expression analysis and genomic studies. He also has a long term interest in the practical aspects of managing genomic data. He is the prior director of the OHSU Spotted Microarray and Illumina Microarray Cores, for which he wrote software packages to transfer, consolidate, and normalize data, as well as to develop a server-based platform for data distribution. He has primary responsibility for data management and security in the Integrated Genomics Laboratory and has recently focused on the management and distribution of terabyte-size datasets commonly produced in "next generation" sequencing.

Greg Finak, PhD, is a postdoctoral fellow in Dr. Raphael Gottardo’s lab, Fred Hutchinson Cancer Research Center. He transitioned from biochemistry to computational biology and bioinformatics at McGill University, where he worked on prognostic biomarker development from breast cancer functional genomics data. In the Gottardo lab he develops new computational algorithms and tools for high throughput analysis of clinical flow cytometry data.

Soren Impey, PhD, Assistant Professor, Oregon Stem Cell Center, and Department of Cell and Developmental Biology, OHSU. The Impey lab utilizes functional genomic approaches to characterize the transcriptional and epigenetic networks that regulate stem cell self-renewal and neural differentiation. The lab developed a novel technology, termed Serial Analysis of Chromatin Occupancy (SACO), that enables the identification of transcription factor target sites or epigenetic marks across entire mammalian genomes. High-throughput sequencing methodologies (ChIP-Seq and RNA-Seq) are utilized to profile transcription factor occupancy, epigenetic modifications, and transcription in an unbiased manner in stem cells and developing neuronal cells. The lab also develops bioinformatics tools to extract and visualize novel biology from functional genomic and transcriptomescreens.

FICCS7 + NWRCM bioinformatics – imaging sessionSteven Jacques, PhD, Professor, Departments of Dermatology and Biomedical Engineering, OHSU. Steve has undergraduate degrees in Biology (MIT) and Electrical Engineering and Computer Science, and a PhD degree in Biophysics and Medical Physics from the University of California-Berkeley (1984). His postdoctoral work was at the Wellman Center for Photomedicine at Massachusetts General Hospital and Dermatology/Bioengineering at Harvard Medical School, where his team developed the use of Monte Carlo computer simulations to study optical transport in biological tissues. At OHSU he continues on develop novel uses of optical technologies for therapy and diagnosis, such as a hand-held polarized light camera to visualize skin cancer margins and guide surgical excision, now in clinical trials. He has also developed in vivo sub-nm measurements of vibration of the cochlear membrane of the inner ear in animal models, and is developing novel microscopes sensitive to the ultrastructure of cells and tissues.

Daniel Gareau, PhD has undergraduate degrees in electrical engineering, and a PhD in biomedical engineering from OHSU. Dr. Gareau’s postdoctoral training was completed at Memorial Sloan Kettering and he is published in the Journal of Biomedical Optics, British Journal of Dermatology, Journal of Investigative Dermatology, and Journal of Microscopy. Dr. Gareau is currently a postdoctoral fellow at the Oregon Health & Science University in the departments of dermatology and biomedical engineering.

Niloy Choudhury, PhD, is a senior research associate in the Department of Biomedical Engineering. Niloy’s PhD is in Physics with an emphasis on optoelectronics and photonics. After 4 years in industry developing opto-electronic instrumentation, he joined OHSU to work on biomedical optics. With NIH funding he has been developing low-coherence measurements of vibration in the middle ear of animal models.

Kevin Phillips, PhD, is a Physical Sciences in Oncology Postdoctoral Fellow with a joint appointment in the Department of Dermatology and Biomedical Engineering. He recently won a PS-OC Trans-network award to investigate the nanoscale architecture of circulating tumor cells detected in the peripheral blood of melanoma/breast/colorectal cancer patients using partial wave spectroscopic microscopy and quantitative phase microscopy.

Scattering pattern from a 5μm sphere, illuminated at λ = 460nm, and numerically recovered from a DIC image (scatter pattern and scatter intensity, azimuthal average v. polar angle).

8am – 3pm, FloCyte course “Data analysis for flow”, OL 219

Schedule: 2011 Northwest Regional Cytometry MeetingThursday, March 24

8:30am – 2:30pm 2011 NWRCM + FICCS78:30am – 8:45am, J. Paul Robinson and Ryan Brinkman, chair’s introductory remarks, Auditorium8:50am – 9:20am, Obi Griffith, “-Omic predictors of drug response in breast cancer”, Auditorium9:25am – 9:55am, Laura Heiser, “Pathway analysis of breast cancer subtypes: Implications for targeted therapies”, Auditorium

10:35am – 11:05am, Bob Searles, “Considerations for real-time handling of terabyte-sized datasets”, Auditorium

11:20am – 11:50am, Steve Jacques, “Label-free scatter-based imaging”, Auditorium11:55am – 12:25pm, Dan Gareau, “Non-invasive morphometric digital pathology”, Auditorium12:25pm – 1:00pm, Lunch break, and Niloy Choudhury, “Oblique angle low coherence imaging and vibrometry for 3D sensing of vibration in the Organ of Corti”, OL 2171:00pm – 1:30pm, Nikesh Kotecha and Chad Rosenberg, “Opportunities in cloud computing for flow cytometry”, Auditorium1:35pm – 2:05pm, Adam Triester, “The goodness of gates: Metrics and visualizations to compare event classification“, Auditorium2:10pm – 2:40pm, Greg Finak, “Bayesian gating of flow cytometry data”, Auditorium2:50 pm, NWRCM introduction, Auditorium3:00pm – 3:40pm, Garry Nolan, “Systems immunology in the post-fluorescence era”, Auditorium3:45pm – 4:25pm, Norman Dovichi, “Chemical and metabolic cytometry”, Auditorium4:30pm – 5:10pm, Manfred Auer, “Imaging biological function across scales: From macromolecules to cells, tissues, and microbial communities”, Auditorium5:15pm – 5:35pm, “BD PermKey: A compendium of results for guiding the design of multiplexed intracellular protein and surface marker staining for flow cytometry”, OL 2175:15pm – 5:35pm, Shervin Javadi, “Performing multi-color analysis using Stratedigm’s S1000Ex analyzer with ‘Smart Detect’ technology”, OL 215 5:15pm – 5:35pm, Mark Munson, “An update on Gemstone: A guided journey through multi-dimensional space”, OL 2195:40pm – 6:20pm, Masanobu Yamamoto, “50 years of laser technology: Innovating flow cytometry with Blu-ray technology”, Auditorium6:25pm – 7:00pm, Gregory Kaduchak, “Acoustic focusing flow cytometry”, Auditorium7pm reception dinner, Old Library7:30pm, dinner speaker, Joe Gray, “Spatial systems biology – the central role of cytometry in the interpretation of the genome”

Friday, March 258:00am Introduction, Auditorium8:05am – 8:45am, J. Paul Robinson, “Coordinated parallel systems approach to high content/high throughput flow cytometry drug screening”, Auditorium8:50am -9:30am, Holden Maecker, “Standardization and validation of fluorescence and mass cytometry panels”, Auditorium9:35am -10:15am, Chris Novak, “Astrios: A new frontier in cell sorting”, OL 2179:35am – 9:55am, Lani Duenas, “A new dye from BioLegend”, OL 2159:35am – 9:55am, Dave Novo, “What’s new in FCS Express 4?”, OL 2199:55am – 10:15am, Kevin Phillips, “Quantitative phase imaging of circulating tumor cells”, OL 2159:55am – 10:15am, Kevin Krouse, “Reagent database”, OL 21910:15am – 10:45am, “Benchtop instruments from Life Technologies to complement your work-FLOW”, OL 217

11:05am – 11:20am, Coffee break

10:00am – 10:30am, Soren Impey, “High throughput sequencing of the transcriptome and epigenome from limiting amounts of cells”, Auditorium

Schedule: 2011 Northwest Regional Cytometry MeetingFriday, March 25 continued10:15am – 10:45am, Matt Alexander, “4 lasers, plate loader, and electronic volume: The new Eclipse is an affordable cytometer for DNA, flourescent proteins, and immunophenotyping”, OL 21510:15am – 10:45am, Bill Ortyn, “Advances in applications for imaging cytometry”, OL 21910:45am – 11:05am, Nikesh Kotecha, “Cytobank: A web-based approach for analysis and publication of flow cytometry experiments”, OL 21710:45am – 11:05am, Kim Luu, “Validation of a 96-well high-throughput flow cytometer screening assay for simultaneously measuring 5 indicators of apoptosis”, OL 21510:45am – 11:05am, Weon Bae, “BioSorter: Flow cytometry for large objects”, OL 21911:10am – 11:50am, Maria Jaimes, “Quality assurance and variability in intracellular staining”, Auditorium11:55am – 12:35pm, Ryan Brinkman, “Automated gating and analysis: State of the art and prospects”, Auditorium12:40pm – 1:20pm, “Cytometry informatics in 2020” lunch-time panel discussion, OL 21712:40pm – 1:00pm, Jeff Harvey, “JSAN cell sorter update and advances in platelet analysis with the Apogee A50 Analyzer”, OL 2151:00pm – 1:20pm, Colin Fitzpatrick, “FlowCytomix – Multiple analyte detection on the flow cytometer”, OL 21512:40pm – 1:00pm, Katye Fichter, “Quantum dot probes to measure the spatio-temporal dynamics of cell signaling”, OL 219 1:00pm – 1:20pm, Jared Jimenez [Cytek], “Taking your cytometry from the past into the future”, OL 219 1:25pm – 1:55pm, Ray Lefebvre, “Innovative approaches to analyzing and visualizing multivariate data using InCyte software”, OL 2171:25pm – 1:55pm, Clare Rogers, “Simplifying fluorescence quantification by standardizing fluorescence detection”, OL 2151:25pm – 1:55pm, Jay Almarode, “A look at new FlowJo metrics for analyzing the quality of human or algorithmic gating operations”, OL 2192:00pm – 2:40pm, Ger van den Engh, “Parsing cytometric noise”, Auditorium2:45pm – 3:25pm, Bob Zigon, “Architectures for real time processing and mining of flow cytometry data”, Auditorium3:30pm – 4:10pm, Bill Ortyn, “High parameter morphological classification replicates the sensitivity and specificity of immunofluorescence labeling in whole blood samples”, Auditorium4:15pm – 4:50pm, Joe Trotter, “New Influx software: Fundamentally changing how software supports cell sorting”, OL 2174:15pm – 4:50pm, Woodside Logic, TBA, OL 215 4:15pm – 4:50pm, TBA, OL 2194:55pm – 5:35pm, C. Bruce Bagwell, “Probability State Modeling for automated high-dimensional cytometry”, Auditorium5:40pm – 6:20pm, Dave Novo, “Consolidated software for flow and image cytometry”, Auditorium6:20pm FRIDAY NIGHT FUN

Saturday, March 268am – TBA*, C. Bruce Bagwell, “Analysis of high dimensional data in Gemstone”8am – TBA*, Dave Novo, “Interactive session on image cytometry analysis”8am – TBA*, Nikesh Kotesha, “Management and analysis of flow cytometry based signaling assays using Cytobank”8am – TBA*, Ray Lefebvre, “Innovative approaches to visualizing and analyzing multivariate data using InCyte software“8am – TBA*, Clare Rogers, “Standardized fluorescence detection”8am – TBA*, Bob Zigon, “Kaluza: Exploring new dimensions in flow cytometry analysis”10am – 5pm, FloCyte course, “Standardization, validation, characterization, and controls”, OL 21510am – 3pm, Adam Triester, “FlowJo for integrators”, 10am – 3pm, Julie Hill, “Cytometry workshop for middle and high school teachers”, OL 217* These practicums will be scheduled based on the information received from registrations. The practicums have a summary front-end for the benefit of those taking the 10am courses. Non-conflicting practicums may be scheduled for later in the morning.

RegistrationRegistration is optional, but required if you wish to attend the Thursday evening reception dinner and Friday lunch. The $25 registration helps defray some of the cost. The number for dinner and lunch will be determined from the number of registrants. Registrants will get a name-tag, which also serves as a ticket for dinner and lunch. Registration closes March 19. You can register and pay online by going to www.ohsu.edu/ohrc , and clicking on the 2011 NWRCM caption to the left. (Online registration is preferred to the fax alternative, as it requires less hand processing.)

Attendance at sessions does not require registration. Walk-ins are encouraged. In addition, providing information on the registration form without payment (no name-tag) is also an option if you wish to be put on the mailing list.

If you have questions about registration or payment, please contact Allan Kachelmeier, 2011 NWRCM coordinator, at kachelme@ohsu.edu .

Posters. We have limited space for posters. Abstracts should be sent to the coordinator by February 15th to be considered, and those selected will be notified by February 20th. Posters not associated with vendors will have priority.

Hotels and transportation.

Transportation from the airport is convenient using the Max line (38 min and $2.35 to downtown). The # 8 bus goes from downtown to OHSU, with tickets valid for 2 hours. Downtown to OHSU (walking distance) is close and convenient using public transport.

When making hotel reservations, mention you’ll be attending a conference at OHSU. A partial listing of hotels downtown giving special OHSU conference rates include:

Benson Hotel, 503-228-2000, $128Paramount Hotel, 503-223-9900, $114Governor Hotel, 503-224-3400, $109Marriott Downtown Waterfront, 503-226-7600, $99University Place, 503-221-0140, $67

Parking at OHSU on the hill, particularly for March 24, is extremely limited. Take the bus (#8) if you can. If you must drive, park in the Schnitzer lot on the South Waterfront ($7) and take the complimentary aerial tram ride up. The Schnitzer parking booth closes at 1:30pm. Parking on Saturday is free. More parking information and map are available at http://www.ohsu.edu/parking .

The 2011 NWRCM steering committee includes Jack Dunne, Vice-President, BD Biosciences, Janet Siebert, President, CytoAnalytics, Julie Hill, Dendreon Corporation, and Shannon McWeeney, Associate Professor, Bioinformatics and Computational Biology, OHSU. Support for this program was obtained from Oregon Hearing Research Center, ISAC, Knight Cancer Institute, OHSU Department of Physiology and Pharmacology, OHSU Department of Biomedical Engineering, Portland State University Physics Department, BD Biosciences (including support for Maria James and Ger van den Engh), Beckman Coulter (including support for Bob Zigon and the Friday evening happy hour), Life Technologies (including support for Gregory Kaduchak and the Friday happy hour), Sony Life Electronics (including support for Masanobu Yamamoto), Accuri Cytometers, Millipore, Amnis Corporation (including support of Dave Basiji), Automation Lab Technologies, Bay Bioscience, BioLegend, Cytek Development, CytoBank, DeNovo Software (including support of Dave Novo), eBioscience, Labkey, Immudex, Intellicyt, Miltenyi Biotec, Partec, Propel Labs, StemCell Technologies, Stratedigm, Tree Star, Union Biometrica, Verity Software (including support of Bruce Bagwell), and Woodside Logic.

Automation Lab Technologies

Department of Physiology and Pharmacology, OHSU

Department of Physics

Biomedical EngineeringOregon Health & Science University

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