コース：

ハイコンテント分析上級コース　HCAの最先端 *

Course Instructors:

• Anthony M. Davies, Ph.D., Director, Irish National Center for High-Content Screening and Analysis (INCHA)
• Paul A. Johnston, Ph.D., Research Associate Professor, Department of Pharmacology and Chemical Biology, Drug Discovery Institute, University of Pittsburgh School of Medicine
• Karol Kozak, Ph.D., Head, Data Handling Unit and High-Content Screening, ETH Zurich

8:30-9:00 am Registration for the Advanced Workshop and Morning Coffee

9:00-10:30 パート1：計装および自動化

Anthony M. Davies, Ph.D., Director, Irish National Center for High-Content Screening and Analysis (INCHA)

• What’s New in HCS: An examination of the functionality and unique selling points of the established and very latest HCA hardware systems, latest cell-based assay technologies, and image and data analysis tools to reach the marketplace. This section of the course will provide a comprehensive overview of the HCA technologies currently available and is a must for anyone looking to acquire an HCA platform.
• Choosing the Right HCA Platform for Your Research Requirements: An assessment of the most widely used methods employed when choosing an HCA instrument. In this part of the workshop we will examine the approaches that are commonly taken when choosing a new HCA platform and the key factors that need to be considered before procurement.
• Imaging, Robotics and People: To ensure maximum productivity as soon as possible it is essential that key issues surrounding the managment of your lab or research program are identifed. Success is not only dependent on choosing the right HCA platform but also in the managment of relationships with users and vendors. Additionally, it is crucal to plan your procurement trajectory to ensure that your current and future research needs are met seemlessly. Topics will include: (i) training your personnel for maximum productivity, (ii) effective strategies for managing equipment and users, (iii) procurement planning and justification for purchase of equipment, (iv) how to stay friends with your equipment vendor, and (v) robotics, automation and software—what is available and what you will need.

10:30-11:00 Networking Coffee Break

11:00 am-12:30 pm パート2：データ分析および管理

Karol Kozak, Ph.D., Head, Data Handling Unit and High-Content Screening, ETH Zurich

The amount of data produced within the HCA field is enormous. Together with the complexity of the data this makes the handling of HCA data not a simple task. This course will cover several data management and analysis-related topics that are of importance in the field of HCA. Next to different multivariate data analysis approaches, related topics like experimental design and data preprocessing will be discussed. In the course multivariate statistical methods will be introduced and illustrated by practical examples from the HCA department. This course reviews the different ways to analyze and manage large sets of HCS data, including the questions that can be asked and the challenges in interpreting the measurements. The main image-processing approaches used in HCA, such as image segmentation and image correction, will be outlined.

Main Course Objectives:

• Specify the steps of HCS data analysis
• Very simple data management and data structures
• Background in image processing
• Library management and library quality
• List the issues associated with HCS data analysis
• List various data standards
• Open source versus commercial software applications
• Role of workflow systems in HCS
• Laboratory Information Management Systems (LIMS): web versus desktop
• Quality control and normalization
• Explain what filtering is and how it should be properly used
• List different pattern discovery techniques such as clustering and classification
• Explain how biological interpretation is linked to pattern discovery
• List the different modules of an HCS informatics system

12:30-1:30 Enjoy Lunch on Your Own

1:30-3:00 パート3：HCA/HCS分析試料の開発

Paul A. Johnston, Ph.D., Research Associate Professor, Department of Pharmacology and Chemical Biology, Drug Discovery Institute, University of Pittsburgh School of Medicine

The development of high-content analysis/screening assays (HCA/HCS) involves the optimization of sample preparation methods, image acquisition procedures, and image analysis algorithms:

• HCS sample preparation is a complex, multi-component process that includes selection and optimization of cell line, microtiter plate, fixative, permeabilization buffer, blocking buffer, wash buffer, primary and secondary antibodies, and fluorescent probes. The choices made for automating cell plating, compound treatment, and sample preparation will also have a significant impact on the biology and the consistency of HCS assays.
• Image acquisition for HCS assays requires input on the objective, the number of channels to be acquired, the excitation and emission filters, focal offsets required relative to the autofocus point, exposure times, and number of image fields that need to be captured. The choice of magnification (objective) profoundly affects HCS assay performance and throughput by impacting the resolution, field of view, detection sensitivity, and the output from the image analysis algorithm. The selection of fluorophores, filter performance and mode of image acquisition all impact the sensitivity, signal-to-background, signal-to-noise, and throughput of the HCS system.
• Image analysis can be achieved at several levels: pixels, objects, semantic concepts, and at the pattern and knowledge level. Digital images are composed of pixels, or squares of uniform grey values captured by a CCD camera or PMT that are assigned to objects established through segmentation. The user defines the objects and features to be extracted automatically from every image prior to the analysis procedure. The selection and optimization of the final image analysis parameters typically involves the iterative use of a training set of images, most commonly the assay controls for the top and bottom of the HCS assay signal window.
• To illustrate the process and selections that are required during the HCS assay development process, several case histories will be presented.

3:00-3:30 Networking Refreshment Break

3:30-5:00 Part 4: Questions and Panel Discussion

5:00 Close of Day

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