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Description
The successful isolation of specific cellular structures permits in-depth studies of their function and composition. This laboratory manual provides step-by-step protocols for the extraction of subcellular components from animal tissues, yeasts, plants, and cultured cells. Each chapter describes methods for isolating a particular organelle, vesicle, membrane, or macromolecular complex. Strategies for breaking cells without damaging 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.
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Content
Contents
(preliminary) 1. Introduction: Subcellular Fractionation—An introduction to cellular compartmentalisation and fractionation techniques. Discussion of fractionation of different samples (plant, human/animal including tissue culture cells), yeast and bacteria.—Methods of fractionation (ways to break open/homogenise tissues) and then organelle/sample isolation such as differential centrifugation, density gradient centrifugation and direct isolation (magnetic, affinity chromatography).2. Plasma membrane isolation using immobilised concanavalin ASue-Hwa Lin3. Isolation of clathrin coated vesicles from tissue culture cells Stephen Royle 4. Purification of endosomes from cellsLukas Huber,Mariana Eca, Guimaraes de Araujo, Giorgia Lamberti5. The synaptosome as a model system for studying synaptic physiologyGareth Evans6. Isolation of phagosomes from tissue culture cellsPaul Pryor7. Isolation of exosomes from cell culture media, blood plasma, serum, urine and cerebrospinal fluidEmily Zeringer, Mu Li, Tim Barta, Alexander V. Vlassov8. Isolation of lysosomes from rat tissue and tissue culture cellsPaul Pryor9. Isolation of Specialised Secretory Compartments: GLUT4 Storage VesiclesJessica Sadler, Christopher Lamb, Gwyn Gould, Nia Bryant10. Isolation of peroxisomes from yeastJana Cramer, Daniel Effelsberg, Wolfgang Girzalsky, Ralf Erdmann11. Isolation of microtubules, MAPs, and mictrotubule-dependent motor proteins12. Golgi isolationDanming Tang, Yanzhuang Wang13. Isolation of mitochondria from cells and tissuesD.A. Clayton, HHMI Janelia Farm with Gerald Shadel14. Topic: Chloroplast isolation15. Subcellular fractionation of rough microsomesD.D. Sabatini16. Purification of ribosomes, ribosomal subunits, and polysomesM. Rivera, A.M. Aguinaldo, T. Atha, J. Lake, A. Scheinman PROTOCOLSIsolation of Ribosomes by ChromatographyPurification of Ribosomes, Ribosomal Subunits, and PolysomesIsolation of Ribosomes and Polysomes by Sucrose Density Gradient CentrifugationPurification of 70S RibosomesPurification of PolysomesDissociation of Ribosomes into Large and Small Subunits 17. Rapid isolation of nuclei from cells in vitroKarl Riabowol18. The nuclear matrix: Preparation for microscopy and biochemical analysisDawn Coverley, Rosemary Wilson GlossaryAppendixReferencesIndex
Audience/Reader
Zellbiologie, Zytologie, Biochemie
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CURRENT PROTOCOLS Wiley USA