Proteomics in Practice presents the various techniques and routes required in approaching proteomic analysis.

Key methods explained are electrophoretic techniques including isoelectric focusing, SDS page, 2-D page and DIGE, and liquid chromatography, and reversed-phase HPLC.

The second part of Proteomics in Practice is a comprehensive, laboratory manual spanning the full range of methods from sample preparation to protein identification.

The third part is a unique troubleshooting guide, designed to answer many of the frequently asked questions regarding proteome analysis.

The final section of Proteomics in Practice contains a thorough reference list to guide interested readers towards further detail.

Contents

• History
• Critical Points
• Proteomics Strategies
• Concepts of Experimental Planning

Proteomics Technology

1. Electrophoretic Techniques

• The Principle of Electrophoresis and Some Methodological Background
• Polyacrylamide Gel Electrophoresis
• Blotting
• Isoelectric Focusing
• Two-Dimensional Electrophoresis
• Image Analysis
• Spot Handling

2. Liquid Chromatography Techniques

• Basic Principles of Important Liquie Chromatography Techniques
• Strategic Approach and General Applicability
• Liquid Chromatography Techniques and Applications in Proteome Analysis
• Practical Considerations and Application of LC-Based Protein Pre-Fractionation
• Critical Review and Outlook

3. Mass Spectrometry

• Ionization
• Ion Separation
• Generating MS Data for Protein Identification
• Protein Characterization
• Protein Quantification Using Mass Spectrometry
• MS Strategies

4. Functional Proteomics: Studies of Protein-Protein Interactions

• Non-immunological Methods
• Antibody-based Techniques

Practical Manual of Proteome Analysis

Equipment, Consumables, Reagents

1. Sample Preparation
2. Fluorescence Difference Gel Electrophoresis
3. Isoelectric Focusing
4. SDS Polyacrylamide Gel Electrophoresis
5. Scanning of Gels Containing Pre-Labled Proteins
6. Staining of Gels
7. Image Analysis and Evaluation of DIGE Gels
8. Spot Excision
9. Sample Destaining
10. Protein Digestion
11. Microscale Desalting and Concentrating of Sample
12. Chemical Derivatization of the Peptide Digest
13. MS Analysis
14. Calibration of MALDI-ToF MS
15. Preparing for a Database Search

Trouble Shooting

• Two-dimensional Electrophoresis
• Mass Spectrometry

Index

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