Proteomics Today Protein Assessment and

The last few years have seen an unprecedented drive toward the application of proteomics to resolving challenging biomedical and biochemical tasks. Separation techniques combined with modern mass spectrometry are playing a central role in this drive. This book discusses the increasingly important role of mass spectrometry in proteomic research, and emphasizes recent advances in the existing technology and describes the advantages and pitfalls as well. Provides a scientifically valid method for analyzing the approximatey 500,000 proteins that are encoded in the human genome Explains the hows and whys of using mass spectrometry in proteomic analysis Brings together the latest approaches combining separation techniques and mass spectrometry and their application in proteome analysis Comments on future challenges and how they may be addressed Includes sections on troubleshooting

MAHMOUD HAMDAN, PhD, is the head of Mass Spectrometry & Separation Technologies, GlaxoSmithKline, Verona, Italy. <p>PIER GIORGIO RIGHETTI, PhD, is Professor of Biochemistry and director of the Laboratory of Proteome Science at the University of Verona, Italy.

PREFACE.

ACKNOWLEDGMENT.

I INTRODUCTION TO PART I.

1 INSTRUMENTATION AND DEVELOPMENTS.

1.1 Introduction.

1.2 Ionization Techniques for Macromolecules.

1.3 Examples on Analytical Solutions Based on FAB–MS.

1.4 Electrospray Ionization.

1.5 Matrix-Assisted Laser Desorption Ionization.

1.6 Ion Detection.

1.7 Types of Analyzers.

1.8 Hybrid Analyzers.

1.9 Tandem Mass Spectrometry.

1.10 Current MS Instrumentation in Proteome Analyses.

1.11 Current MS-Based Proteomics.

1.12 Recent Achievements and Future Challenges.

1.13 Concluding Remarks.

References.

2 PROTEOMICS IN CANCER RESEARCH.

2.1 Introduction.

2.2 Pancreatic Ductal Adenocarcinoma.

2.3 Proteomic Analysis of Human Breast Carcinoma.

2.4 Proteomic Profiling of Chemoresistant Cancer Cells.

2.5 Signal Pathway Profiling of Prostate Cancer.

2.6 Emerging Role of Functional and Activity-Based Proteomics in Disease Understanding.

2.7 Activity-Based Protein Profiling.

2.8 Probing Protein Functions Using Chromophore-Assisted Laser Inactivation.

2.9 Role of Protein–Tyrosine Kinases.

2.10 Concluding Remarks and Future Prospects.

References.

3 CURRENT STRATEGIES FOR PROTEIN QUANTIFICATION.

3.1 Introduction.

3.2 Global Internal Standard Technology.

3.3 Differential In-Gel Electrophoresis.

3.4 Quantification of Modified Proteins.

3.5 Comments and Considerations.

3.6 Other Approaches.

3.7 Emerging Role of Microfluidic Devices.

3.8 Concluding Remarks.

References.

II PROTEOMICS TODAY: SEPARATION SCIENCE AT WORK.

4 CONVENTIONAL ISOELECTRIC FOCUSING IN GEL MATRICES AND CAPILLARIES AND IMMOBILIZED pH GRADIENTS.

4.1 Introduction.

4.2 Conventional Isoelectric Focusing in Amphoteric Buffers.

4.3 Immobilized pH Gradients.

4.4 Capillary Isoelectric Focusing.

4.5 Separation of Peptides and Proteins by CZE in Isoelectric Buffers.

4.6 Conclusions.

References.

5 SODIUM DODECYL SULFATE–POLYACRYLAMIDE GEL ELECTROPHORESIS.

5.1 Introduction.

5.2 SDS–Protein Complexes: a Refinement of the Model.

5.3 Theoretical Background of M<sub>r</sub> Measurement by SDS–PAGE.

5.4 Methodology.

5.5 Gel Casting and Buffer Systems.

5.6 Blotting Procedures.

5.7 Conclusions.

References.

6 TWO-DIMENSIONAL MAPS.

6.1 Introduction.

6.2 Some Basic Methodology Pertaining to 2D PAGE.

6.3 Prefractionation Tools in Proteome Analysis.

6.4 Multidimensional Chromatography Coupled to MS.

6.5 Protein Chips and Microarrays.

6.6 Nondenaturing Protein Maps.

6.7 Spot Matching in 2D Gels via Commercial Software.

6.8 Conclusions.

References.

INDEX.

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Proteomics Today Protein Assessment and Biomarkers Using Mass Spectrometry, 2D Electrophoresis,and Microarray Technology

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