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 Filtration Chromatography -- 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. New technologies and the manner in which scientific research is conducted have been fully incorporated into this thoroughly revised second edition of PROTEIN ANALYSIS AND PURIFICATION. Among the many topics introduced to this new edition are the use of robotics in the lab, the impact of the codification of the human genome, functional proteomics, mass spectrometry, and the expanding use of bioinformatics as a tool to improve efficiency and productivity. The author has also included an annotated appendix of the educational materials and technical databases available online to enhance the researcher's benchside access to the latest resources. With solid explanations of the science asking the "why" and literally dozens of step-by-step protocols providing the "how," this new edition is a complete resource for the benchside.