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This volume of the acclaimed Methods in Cell Biology series provides specific examples of applications of confocal microscopy to cell biological problems. It is an essential guide for students and scientists in cell biology, neuroscience, and many other areas of biological and biomedical research, as well as research directors and technical staff of microscopy and imaging facilities. An integrated and up-to-date coverage on the many various techniques and uses of the confocal microscope (CM). Includes detailed protocols accessible to new users Details how to set up and run a "Confocal Microscope Core Facility" Contains over 170 figures
This volume of the acclaimed Methods in Cell Biology series provides specific examples of applications of confocal microscopy to cell biological problems. It is an essential guide for students and scientists in cell biology, neuroscience, and many other areas of biological and biomedical research, as well as research directors and technical staff of microscopy and imaging facilities. An integrated and up-to-date coverage on the many various techniques and uses of the confocal microscope (CM). - Includes detailed protocols accessible to new users - Details how to set up and run a "Confocal Microscope Core Facility" - Contains over 170 figures
In Confocal Microscopy Methods and Protocols, Stephen Paddock and a highly skilled panel of experts lead the researcher using confocal techniques from the bench top, through the imaging process, to the journal page. They concisely describe all the key stages of confocal imaging-from tissue sampling methods, through the staining process, to the manipulation, presentation, and publication of the realized image. Written in a user-friendly, nontechnical style, the methods specifically cover most of the commonly used model organisms: worms, sea urchins, flies, plants, yeast, frogs, and zebrafish. Centered in the many biological applications of the confocal microscope, the book makes possible the successful imaging of both fixed and living specimens using primarily the laser scanning confocal microscope. The powerful hands-on methods collected in Confocal Microscopy Methods and Protocols will help even the novice to produce first-class cover-quality confocal images.
There has been a great upsurge in interest in light microscopy in recent years due to the advent of a number of significant advances in microscopy, one of the most important of which is confocal microscopy. Confocal microscopy has now become an important research tool, with a large number of new fluorescent dyes becoming available in the past few years, for probing your pet structure or molecule within fixed or living cell or tissue sampies. Many of the people interested in using confocal microscopy to further their research do not have a background in microscopy or even cell biology and so not only do they find considerable difficulty in obtaining satisfactory results with a confocal microscope, but they may be mislead by how data is being presented. This book is intended to teach you the basic concepts ofmicroscopy, fluorescence, digital imaging and the principles of confocal microscopy so that you may take full advantage ofthe excellent confocal microscopes now available. This book is also an excellent reference source for information related to confocal microscopy for both beginners and the more advanced users. For example, do you need to know the optimal pinhole size for a 63x 1. 4 NA lens? Do you need to know the fluorescence emission spectrum of Alexa 568? Access to the wealth of practical information in this book is made easier by using both the detailed index and the extensive glossary.
Cell Biological Applications of Confocal Microscopy instructs researchers on methods of preparing biological material for confocal microscopy. Individual chapters describe the technical problems of imaging structures in thick specimens and useful techniques such as multi-color dye visualization. The book provides specific examples of applications of confocal microscopy to cell biological problems. This volume is an essential guide for students and scientists incell biology, neuroscience, and many other areas of biological and biomedical research, as well as research directors and technical staff of microscopy and imaging facilities.Describes proper specimen preparation for obtaining high-quality data and imagesCovers pitfalls in imaging different specimens and in employing different dyesContains detailed protocols accessible to new usersChapters are self-contained units, but describe strategies that can be combined to develop new protocolsTechniques are illustrated with full-color plates
The previous edition of this book marked the shift in technology from video to digital camera use with microscope use in biological science. This new edition presents some of the optical fundamentals needed to provide a quality image to the digital camera. Specifically, it covers the fundamental geometric optics of finite- and infinity-corrected microscopes, develops the concepts of physical optics and Abbe's theory of image formation, presents the principles of Kohler illumination, and finally reviews the fundamentals of fluorescence and fluorescence microscopy. The second group of chapters deals with digital and video fundamentals: how digital and video cameras work, how to coordinate cameras with microscopes, how to deal with digital data, the fundamentals of image processing, and low light level cameras. The third group of chapters address some specialized areas of microscopy that allow sophisticated measurements of events in living cells that are below the optical limits of resolution. - Expands coverage to include discussion of confocal microscopy not found in the previous edition - Includes "traps and pitfalls" as well as laboratory exercises to help illustrate methods
Basic Confocal Microscopy, Second Edition builds on the successful first edition by keeping the same format and reflecting relevant changes and recent developments in this still-burgeoning field. This format is based on the Confocal Microscopy Workshop that has been taught by several of the authors for nearly 20 years and remains a popular workshop for gaining basic skills in confocal microscopy. While much of the information concerning fluorescence and confocal microscopy that made the first edition a success has not changed in the six years since the book was first published, confocal imaging is an evolving field and recent advances in detector technology, operating software, tissue preparation and clearing, image analysis, and more have been updated to reflect this. Several of these advances are now considered routine in many laboratories, and others such as super resolution techniques built on confocal technology are becoming widely available.
Advances in technology have revolutionized the development of light microscopy techniques in biomedical research, thus improving visualization of the microstructure of cells and tissues under physiological conditions. Fluorescence microscopy methods are non-contact and non-invasive and provide high spatial and temporal resolution that other laboratory techniques cannot. This well-illustrated book targets graduate students and scientists who are new to the state-of-the-art fluorescence microscopy techniques used in biological and clinical imaging. It explains basic concepts and imaging procedures for wide-field, confocal, multiphoton excitation, fluorescence resonance energy transfer (FRET), lifetime imaging (FLIM), spectral imaging, fluorescence recovery after photobleaching (FRAP), optical tweezers, total internal reflection, high spatial resolution atomic force microscopy (AFM), and bioluminescence imaging for gene expression. The usage of these techniques in various biological applications, including calcium, pH, membrane potential, mitochondrial signaling, protein-protein interactions under various physiological conditions, and deep tissue imaging, is clearly presented. The authors describe the approaches to selecting epifluorescence microscopy, the detectors, and the image acquisition and processing software for different biological applications. Step-by-step directions on preparing different digital formats for light microscopy images on websites are also provided.
Immunohistochemistry and immunocytochemistry are invaluable tools for the visualization of tissue and cellular antigens in diagnostic and biological research environments. The need to obtain accurate, reliable and reproducible results is paramount. It is with this fundamental aim in mind that we have compiled Immunohistochemistry: Essential Methods. We have achieved this by examining each aspect of immunochemistry in turn, with each chapter including detailed information regarding the subject matter in question. Each chapter is written by an expert in their field and includes protocols that are typically used in their own research. Subjects covered are, amongst others, antibodies and their production; selection of reporter labels; immunochemical staining methods and experimental design (both using single and multiple reporter labels); quality assurance; automated immunochemistry; confocal microscopy and electron microscopy. In addition, benefits and limitations of each approach are discussed within the chapters.
Recent advances in imaging technology reveal, in real time and great detail, critical changes in living cells and organisms. This manual is a compendium of emerging techniques, organized into two parts: specific methods such as fluorescent labeling, and delivery and detection of labeled molecules in cells; and experimental approaches ranging from the detection of single molecules to the study of dynamic processes in organelles, organs, and whole animals. Although presented primarily as a laboratory manual, the book includes introductory and background material and could be used as a textbook in advanced courses. It also includes a DVD containing movies of living cells in action, created by investigators using the imaging techniques discussed in the book. The editors, David Spector and Robert Goldman, whose previous book was Cells: A Laboratory Manual,are highly respected investigators who have taught microscopy courses at Cold Spring Harbor Laboratory, the Marine Biology Laboratory at Woods Hole, and Northwestern University.