Benchtop Techniques
Buch, Englisch, 520 Seiten, Format (B × H): 186 mm x 249 mm, Gewicht: 2080 g
ISBN: 978-0-8176-4341-6
Verlag: Birkhauser Boston
Fully revised and newly enhanced, PROTEIN ANALYSIS AND PURIFICATION provides the research with the most complete picture of the state of the art technology and this expanding and dynamic field. Now enhanced with data on mass spectrometry techniques, the use of robotics and in silicon science, and in appendix of online databases, this new edition provides the most current, complete, and user-friendly bench side resource to both the experienced research and the novice investigator.
Zielgruppe
Research
Autoren/Hrsg.
Fachgebiete
Weitere Infos & Material
1 An Overview of This Manual.- Kits, Cores, and Computers.- How to Use This Book.- Basic Laboratory Equipment.- Laboratory Automation.- Beyond Protein Analysis and Purification.- 2 Protein Structure.- A. The Amino Acids.- B. The Four Levels of Protein Structure.- C. Chemical Characteristics of Proteins.- 3 Tracking the Target Protein.- A. Labeling Cells and Proteins.- B. Lysis—Preparation of the Cell Free Extract.- C. Principles of Immunoprecipitation.- D. Additional Methods to Identify Associated Proteins.- 4 Electrophoretic Techniques.- to Polyacrylamide Gel Electrophoresis (PAGE).- A. Preparation of SDS-Polyacrylamide Gels.- B. 2-Dimensional (2-D) Gel Systems.- D. Recovery of Proteins from the Gel.- E. Identification of Enzyme Activity in Polyacrylamide Gels.- 5 Getting Started with Protein Purification.- A. Making a Cell Free Extract.- B. Protein Quantitation.- C. Manipulating Proteins in Solution.- D. Precipitation Techniques.- 6 Membrane Proteins.- A. Peripheral Membrane Proteins.- B. Integral Membrane Proteins.- C. Detergents.- 7 Transfer and Detection of Proteins on.- Membrane Supports.- A. Transfer of Proteins to Membrane Supports.- B. Staining the Blot.- C. Recovery of Proteins from the Blot.- D. Detection of the Target Protein.- 8 Identification of the Target Protein.- A. Peptide Mapping.- B. Enzymatic Cleavage of Proteins.- C. Chemical Cleavage.- D. Microsequencing from PVDF Membranes.- MS Basics.- Protein Identification by MS.- Protein Database Searches.- The Rise of Biological Databases.- 9 Identifying and Analyzing Posttranslational.- Modifications.- A. Glycosylation.- B. Phosphorylation.- C. Lipid Modification of Proteins.- D. Selected Modifications.- 10 Chromatography.- A. Important Terminology Used in Chromatography.- B. Gel FiltrationChromatography.- C. Introduction to HPLC.- D. Ion Exchange Chromatography: Separation on the Basis of Charge.- E. Hydrophobic Interaction Chromatography (HIC).- Flow Rate.- 11 Recombinant Protein Techniques.- Recombinant Protein for Antibody Production.- Protein for Biochemical or Cell Biological Studies.- A. In vitro Transcription and Translation.- B. Recombinant Gene Products in E. coli: Expression, Identification and Characterization.- D. Expression of Foreign Proteins in Eukaryotic Cells.- Appendices.- A. Safety Considerations.- First Aid: Emergency Procedures.- B. Antibody Preparation.- Production of Polyclonal Antisera in Rabbits.- Purification of Antibody Using Protein A Affinity Columns.- Numbering Mice.- Molarity.- Choosing and Preparing Buffers.- Common Laboratory Solutions.- Extinction Coefficients.- D. Nucleic Acids.- Spectrophotometric Conversions.- DNA/Protein Conversions.- Oligonucleotide Concentrations.- RNA Precipitation.- E. Modifications and Motifs.- Nomenclature.- Protein Modification Sequences.- Protein Kinase Recognition Sequence Motifs.- Subcellular Localization Motifs.- Protein Databases.- F. Centrifugation.- Nomogram.- General Purpose Centrifuge Rotors.- Ultracentrifuge Rotors.- G. Proteases and Proteolytic Enzyme Inhibitors.- Commonly Used Proteases.- Protease Inhibitors.- H. Radioactivity.- Manual and Machine Film Processing.- L Tissue Culture.- Transwell Permeable Supports.- J. Miscellaneous.- Unit Prefixes.- The Greek Alphabet.- Abbreviations.- HPLC Pump Pressure Conversion.- Dipeptide Masses.- Mass Differences Considered in Molecular Weight Analysis of Proteins.- K. List of Suppliers, Vendors, Manufacturers.
