Many investigations into the structure and function of cells and tissues require the isolation of a particular membrane or subcellular component (organelle). This book covers all the necessary aspects, from breaking up the cells (homogenization), via a variety of separation techniques (the isolation and fractionation chapters), to characterization of the separated organelles.
Eukaryotic cells are remarkably complex structures, containing a vast repertoire of macromolecules, organelles, and other compartments that orchestrate the tasks required for life. For in-depth studies of their function and composition, reliable methods for the isolation of specific subcellular structures are often required. This laboratory manual provides step-by-step protocols for the extraction of subcellular components from animal tissues, yeasts, plants, and cultured cells. Each chapter focuses on a particular eukaryotic organelle, vesicle, membrane, or macromolecular complex. Strategies for breaking cells while maintaining the structural and functional integrity of the component of interest, enriching for that component based on its physical and biochemical characteristics, and monitoring and ensuring the success of the purification procedure are provided. The contributors describe both traditional approaches (e.g., density gradient centrifugation) and innovative techniques (e.g., the use of SPIONs) for isolating subcellular constituents. This manual is therefore an essential laboratory resource for all cell biologists seeking a comprehensive collection of dependable subcellular fractionation methods.
Subcellular Components: Preparation and Fractionation talks about cells and particles' components, including their preparation and fractionation. The book includes theories and answers to questions that are relevant to the study. The first chapter of the book details various facts about homogenization of mammalian cells. This chapter presents the results of studies on solid tissues and single-cell suspensions; the author then offers his conclusion of the study. The next two chapters highlight the methods on isolating nuclei, including the guides for standard assessment and the procedure of isolation, along with analysis of nuclei biochemical properties. The main topics in Chapter 4 are mitochondria from animal tissues and yeasts; this chapter also discusses the preparation for a rat-liver, blowfly flight-muscle, yeast, and brain mitochondria. The chapter that follows widely talks about lysosomes, including its historical background, centrifugal method, and related topics. In the next several chapters, the topics covered include purification, isolation, preparation, and separation of cells including plasma-membrane, polysomes, ribosomes, microsomes, and microvilli. The book serves as a great reference for undergraduates and postgraduates in the field, as it contains a thorough discussion of various relevant studies.
This volume summarizes the new developments that made subcellular proteomics a rapidly expanding area. It examines the different levels of subcellular organization and their specific methodologies. In addition, the book includes coverage of systems biology that deals with the integration of the data derived from these different levels to produce a synthetic description of the cell as a system.
Adipose tissue is recognized to be exquisitely sensitive to hormone action, and is also now recognized as a secretory and endocrine organ required for reproduction and good health. Adipocytes are “smart” cells able within the tissue to communicate with surrounding cells, but also with various organs, particularly via leptin acting on the central nervous system. Brown adipose tissue (BAT) and white adipose tissue (WAT) are known to be distinct tissues, whereas the heterogeneity of WAT depots is well est- lished. Unfortunately, excess WAT leads to obesity, which is the most common health problem in industrialized countries. Therefore, from both a scientific and a technical point of view, the time has come to create a survey of adipose tissues and their neglected adipocytes. In Adipose Tissue Protocols, I have attempted to gather together chapters from all areas of adipose tissue research—from in vivo to in vitro studies—and to provide methods covering a wide variety of techniques, including the choice of adipose tissue depot and of morphological techniques for the study of BAT and WAT; the isolation, subcellular fractionation, and transfection of adipocytes where the low density of these cells must be taken into account; assays of nutrient and ion fluxes and the metabolic aspects of nutrient uptake; assays of lipid-related enzymes; biopsies and quantification of lipid-related mRNAs; cultures of adipose precursor cells from WAT and BAT of various species, including human tissue; measurements of adipose secretory products; and assessment of WAT metabolism in vivo.
Many investigations into the structure and function of cells and tissues require the isolation of a particular membrane or subcellular component (organelle). This book covers all the necessary aspects, from breaking up the cells (homogenization), via a variety of separation techniques (the isolation and fractionation chapters), to characterization of the separated organelles.
This volume details protocols that cover the broad arsenal of techniques used to study a secretion system from A to Z. Chapters focus on identifying and localizing the different subunits, defining interactions within subunits, monitoring conformational changes, purifying and imaging of large complexes, defining the assembly pathway by fluorescence microscopy and the role of energy during assembly and/or secretion, identifying secreted effectors as well as reporters to follow effector transport. Written in the highly successful Methods in Molecular Biology series format, chapters include introductions to their respective topics, lists of the necessary materials and reagents, step-by-step, readily reproducible laboratory protocols, and tips on troubleshooting and avoiding known pitfalls. Authoritative and cutting-edge, Bacterial Protein Secretion Systems: Methods and Protocol aims to provide techniques that are not restricted to the study of secretion systems but are also of specific interest for any researcher interested on multi-protein complexes of the bacterial cell envelope.
Subcellular Fractions: Advances in Research and Application: 2011 Edition is a ScholarlyEditions™ eBook that delivers timely, authoritative, and comprehensive information about Subcellular Fractions. The editors have built Subcellular Fractions: Advances in Research and Application: 2011 Edition on the vast information databases of ScholarlyNews.™ You can expect the information about Subcellular Fractions in this eBook to be deeper than what you can access anywhere else, as well as consistently reliable, authoritative, informed, and relevant. The content of Subcellular Fractions: Advances in Research and Application: 2011 Edition has been produced by the world’s leading scientists, engineers, analysts, research institutions, and companies. All of the content is from peer-reviewed sources, and all of it is written, assembled, and edited by the editors at ScholarlyEditions™ and available exclusively from us. You now have a source you can cite with authority, confidence, and credibility. More information is available at http://www.ScholarlyEditions.com/.
It is a truism of science that the more fundamental the subject, the more universally applicable it is. Neverthelens, it is important to strike a level of "fundamentalness" appropriate to the task in hand. For example, an in-depth study of the mechanics of motor cars would tell one nothing about the dynamics of traffic. Traffic exists on a different "level" - it is dependent upon the existence of motor vehicles but the physics and mathematics of traffic can be adequately addressed by considering motor vehicles as mobile "blobs", with no consideration of how they become mobile. To start a discourse on traffic with a consideration of the mechanics of motor vehicles would thus be inappropropriate. In writing this volume, I have wrestled with the question of the appropriate level at which to address the physics underlying many of the techniques used in protein isolation. I have tried to strike a level as would be used by a mechanic (with perhaps a slight leaning towards an engineer) - i.e. a practical level, offering appropriate insight but with minimal mathematics. Some people involved in biochemical research have a minimal grounding in chemistry and physics and so I have tried to keep it as simple as possible.
