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Lecture Notes forMethods in Cell Biology
(TRMD 623)Instructor:Mark F. Wiser
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TABLE OF CONTENTS
PREFACE—COURSE DESCRIPTION................................................................................7CHAPTER 1--MICROSCOPY..............................................................................................9LIGHT MICROSCOPY.............................................................................................9ELECTRON MICROSCOPY...................................................................................12MICROSCOPY APPENDIX. ALDEHYDE FIXATIVES......................................16CHAPTER 2--SPECTROPHOTOMETRY..........................................................................19SPECTROPHOTOMETRY THEORY....................................................................19INSTRUMENTATION............................................................................................21APPLICATIONS OF SPECTROPHOTOMETRY..................................................21FACTORS AFFECTING ABSORPTION...............................................................22VARIATIONS IN SPECTROPHOTOMETRY.......................................................22APPENDIX I. PHOTOMULTIPLIER TUBE..........................................................24APPENDIX II. CALCULATION OF ENZYME ACTIVITY.................................25CHAPTER 3--FLUORESCENCE........................................................................................29ENZYME ASSAYS AND FLUOROMETERS.......................................................29FLUORESCENCE MICROSCOPY.........................................................................30FLOW CYTOMETRY.............................................................................................33CHAPTER 4--RADIOCHEMISTRY...................................................................................35ATOMIC STRUCTURE AND RADIOACTIVE DECAY......................................35PROPERTIES OF RADIOISOTOPES....................................................................36MEASUREMENT OF ß-RADIATION...................................................................37GAMMA-RAY DETECTION.................................................................................42SAFETY...................................................................................................................42APPENDIX 1. INTERNAL STANDARDS.............................................................43APPENDIX 2. CHANNELS RATIO.......................................................................44APPENDIX 3. MULTIPLE ISOTOPES..................................................................45CHAPTER 5--pH AND BUFFERS......................................................................................47DEFINITIONS.........................................................................................................47HENDERSON-HASSELBACH...............................................................................47INSTRUMENTATION............................................................................................48COMPLICATIONS..................................................................................................48BUFFERING CAPACITY.......................................................................................49BUFFER SELECTION............................................................................................49BUFFER APPENDIX..............................................................................................50
3CHAPTER 6--CENTRIFUGATION....................................................................................53SEDIMENTATION THEORY.................................................................................53PREPARATIVE CENTRIFUGATION...................................................................55CENTRIFUGATION APPENDIX 1. RCF CALCULATION.................................58CHAPTER 7--INTRODUCTION TO PROTEINS..............................................................61PROTEIN STRUCTURE.........................................................................................61PROTEIN STABILITY............................................................................................63PROTEIN ASSAYS.................................................................................................64APPENDIX. AMINO ACIDS: PROPERTIES AND STRUCTURE......................66CHAPTER 8--DIFFERENTIAL PRECIPITATION OF PROTEINS..................................69PROCEDURE FOR (NH
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PRECIPITATION.................................................69OTHER PROTEIN PRECIPITATION METHODS................................................70APPENDIX. AMMONIUM SULFATE NOMOGRAM.........................................71CHAPTER 9--CHROMATOGRAPHY...............................................................................73BASIC PRINCIPALS...............................................................................................73EQUIPMENT...........................................................................................................74ADSORPTION CHROMATOGRAPHY.................................................................75ION EXCHANGE CHROMATOGRAPHY............................................................75HYDROPHOBIC CHROMATOGRAPHY.............................................................79Polar Solvent.........................................................................................................................80GEL FILTRATION CHROMATOGRAPHY..........................................................80AFFINITY CHROMATOGRAPHY........................................................................81CHAPTER 10--MEMBRANES AND DETERGENTS.......................................................83TYPES OF MEMBRANE PROTEINS....................................................................84DETERGENTS.........................................................................................................85DETERGENT REMOVAL......................................................................................87APPENDIX 1. CELL DISRUPTION TECHNIQUES.............................................88CHAPTER 11--ELECTROPHORESIS................................................................................91GEL ELECTROPHORESIS.....................................................................................91EQUIPMENT...........................................................................................................93SDS-PAGE...............................................................................................................94ISOELECTRIC FOCUSING....................................................................................96TWO-DIMENSIONAL GEL ELECTROPHORESIS..............................................97PROTEIN DETECTION FOLLOWING ELECTROPHORESIS............................98PREPARATIVE ELECTROPHORESIS...............................................................101ELECTROPHORESIS APPENDIX 1. SIZE CALCULATION............................104
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FACTORS AFFECTING HYBRIDIZATION for Methods in Cell Biology PDF[1]
this will help who are interested in experimental cell and molecular biology. all the contents are laboratory oriented based on basic experimental procedure.
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