Hands-on researchers describe in step-by-step detail 73 proven laboratory methods and bioinformatics tools essential for analysis of the proteome. These cutting-edge techniques address such important tasks as sample preparation, 2D-PAGE, gel staining, mass spectrometry, and post-translational modification. There are also readily reproducible methods for protein expression profiling, identifying protein-protein interactions, and protein chip technology, as well as a range of newly developed methodologies for determining the structure and function of a protein. The bioinformatics tools include those for analyzing 2D-GEL patterns, protein modeling, and protein identification. All laboratory-based protocols follow the successful Methods in Molecular BiologyT series format, each offering step-by-step laboratory instructions, an introduction outlining the principle behind the technique, lists of the necessary equipment and reagents, and tips on troubleshooting and avoiding known pitfalls.
The Proteomics Handbook Ed. John M. Walker 1. Extraction and Solubilization of Proteins for Proteomic Studies - Richard M. Leimgruber 2. Preparation of Bacterial Samples for 2-D PAGE Brian Berg Vandahl, Gunna Christiansen and Svend Birkelund 3. Preparation of Yeast Samples for 2-D PAGE - Joakim Norbeck 4. Preparation of Mammalian Tissue Samples for Two-Dimensional Electrophoresis - Frank Witzmann 5. Differential Detergent Fractionation of Eukaryotic Cells - Melinda L. Ramsby and Gregory S. Makowski 6. Serum or Plasma Sample Preparation for Two-Dimensional Gel Electrophoresis - Anthony G. Sullivan, Stephen Russell, Henry Brzeski, Richard I. Somiari and Craig D. Shriver 7. Preparation of Plant Protein Samples for 2-D PAGE - David W.M. Leung 8. Laser Assisted Microdissection for Sample Preparation Darrell L. Ellsworth, Stephen Russell, Brenda Deyarmin, Anthony Sullivan, Henry Brzeski, Richard I. Somiari and Craig D. Shriver 9. Purification of Cellular and Organelle Populations by Fluorescence-Activated Cell Sorting for Proteome Analysis - William L. Godfrey, Colette J. Rudd, Sujata Iyer and Diether Recktenwald 10. Purification of Nucleoli From Lymphoma Cells and Solubilization of Nucleolar Proteins for 2-DE Separations - Regis Dieckmann, Yohanne Coute, Denis Hochstrasser, Jean-Jacques Diaz and Jean-Charles Sanchez 11. Pre-Fractionation of Complex Protein Mixtures for 2-D PAGE Using Reversed Phase Liquid Chromatography Volker Badock and Albrecht Otto 12. Fractionation of Complex Proteomes by Microscale Solution Isoelectrofocusing Using ZOOM IEF Fractionators to Improve Protein Profiling - Xun Zuo, Ki-Beom Lee and David W. Speicher 13. Large-Format Two-Dimensional Polyacrylamide Gel Electrophoresis - Henry Brzeski, Stephen Russell, Tony G. Sullivan, Richard Somiari & Craig Shriver 14. Analysis of Membrane Proteins by Two-Dimensional Gels - Michael Fountoulakis, 15. 2-D PAGE of High Molecular Mass Proteins - Masamichi Oh-Ishi and Tadakazu Maeda 16. Using Ultra-Zoom Gels for High Resolution Two-Dimensional Polyacrylamide Gel Electrophoresis Sjouke Hoving, Hans Voshol and Jan van Oostrum 17. NEPHGE and pI Strip Proteomic Two-Dimensional Gel Electrophoretic Mapping of Lipid-Rich Membranes - Steven E. Pfeiffer, Yoshihide Yamaguchi, Cecilia B. Marta, Rashmi Bansal and Christopher M. Taylor 18. Silver Staining of 2-D gels - Julia Poland, Thierry Rabilloud and Pranav Sinha 19. Zn2+ Reverse Staining Technique Carlos Fernandez-Patron 20. Multiplexed Proteomics Technology for the Fluorescence Detection of Glycoprotein Levels and Protein Expression Levels Using Pro-Q Emerald and SYPRO Ruby dyes - Birte Schulenberg and Wayne F. Patton 21. Multiplexed Proteomics Technology for the Fluorescence Detection of Phosphorylation and Protein Expression Levels Using Pro-Q Diamond and SYPRO Ruby Dyes - Birte Schulenberg, Terrie Goodman, Thomas Steinberg and Wayne F. Patton 22. Sensitive Quantitative Fluorescence Detection of Proteins in Gels Using SYPRO Ruby Protein Gel Stain - Birte Schulenberg, Nancy Ahnert and Wayne F. Patton 23. Rapid, Sensitive Detection of Proteins in Minigels with Fluorescent Dyes: Coomassie Fluor Orange, SYPRO Orange, SYPRO Red, and SYPRO Tangerine protein gel stains - Thomas H. Steinberg, Courtenay R. Hart and Wayne F. Patton 24. Difference Gel Electrophoresis in a High Throughput Environment Richard I. Somiari, Stephen Russell, Stella B. Somiari, Anthony G. Sullivan, Darrell L. Ellsworth, Henry Brzeski and Craig D. Shriver 25. Statistical Analysis of 2-D Gel Patterns - Francoise Seillier-Moiseiwitsch 26. 2-DE Databases on the World Wide Web - Christine Hoogland, Khaled Mostaguir and Ron D. Appel 27. Computer Analysis of 2-D images Patricia M. Palagi, Daniel Walther, Gerard Bouchet, Sonja Voordijk and Ron D. Appel. 28. Comparing 2D Electrophoretic Gels Across Internet Databases - An Open Source Application - Peter F. Lemkin, Gregory C. Thornwall and Jai Evans 29. Sample Cleanup by Solid Phase Extraction/Pipette Tip Chromatography - Alastair Aitken 30. Protein Identification by In-Gel Digestion and Mass Spectrometric Analysis - Michele Learmonth and Alastair Aitken 31. Peptide sequences of 2-D Gel Separated Protein Spots by Nanoelectrospray Tandem Mass Spectrometry - Alastair Aitken 32. Identification of Proteins by MALDI-tof MS - Alastair Aitken 33. Sequencing of Tryptic Peptides Using Chemically-Assisted Fragmentation MALDI-PSD - John Flensburg and Maria Liminga 34. The In Situ Characterization of Membrane Immobilized 2-D PAGE Separated Proteins Using Ink-Jet Technology - Patrick W. Cooley, Janice L. Joss, Femia G. Hopwood, Nicole L. Wilson and Andrew A. Gooley 35. Protein Identification by Peptide Mass Fingerprinting - Alastair Aitken 36. Analysis of the Proteomes in Human Tissues by In-Gel Isoelectric Focusing and Mass Spectrometry - Francesco Giorgianni and Sarka Beranova-Giorgianni 37. Liquid Chromatograph Coupled to MS for Proteome Analysis Alastair Aitken 38. Quantitative Analysis of Proteomes and Subproteomes by Isotope Coded Affinity Tags (ICAT) and Solid-Phase Glycoprotein Capture - Eugene Yi, Hui Zhang, Kelly Cooke, Ruedi Aebersold and David R. Goodlett 39. Amino Acid-Coded Mass Tagging for Quantitative Profiling of Differentially Expressed Proteins Xian Chen 40. Mass-Coded Abundance Tagging (MCAT) for Protein Identification and Relative Abundance Determination - Gerard Cagney and Andrew Emili 41. Virtual 2-D Gel Electrophoresis by Mass Spectrometry - Angela Walker, Gary Rymar and Philip Andrews 42. Identification of Post-Translational Modification by Mass Spectrometry Alastair Aitken 43. Approaches to the O-Glycoproteome - Franz-Georg Hanisch and Stefan Muller 44. Identification of Protein Phosphorylation Sites by Mass Spectrometry Alastair Aitken 45. Quantitative Analysis of Protein Phosphorylation Status and Protein Kinase Activity on Microassays Using Pro-Q Diamond Dye Technology - Karen Martin and Wayne F. Patton 46. New Challenges and Strategies for Multiple Sequence Alignment in the Proteomics Era - Julie D. Thompson and Olivier Poch 47. The Clustal Series of Programs for Multiple Sequence Alignment Julie D. Thompson 48. FASTA Servers for Sequence Similarity Search - Biju Issac and Gajendra P.S. Raghava 49. Protein Sequence Analysis and Domain Identification - Chris P. Ponting and Ewan Birney 50. Mammalian Genes and Evolutionary Genomics - Leo Goodstadt and Chris Ponting 51. Computational Identification of Related Proteins: BLAST and PSI-BLAST and Other Tools Qunfeng Dong and Volker Brendel 52. Protein Identification and Analysis Tools on the ExPASy Server - Elizabeth Gasteiger, Christine Hoogland, Alexandre Gattiker, Severine Duvaud, Mark Wilkins, Ron D. Appel and Amos Bairoch 53. Protein Sequence Databases - Michele Magrane, Maria Jesus Martin, Claire O'Donovan and Rolf Apweiler 54. In Silico Characterization of Proteins InterPro and Proteome Analysis - Nicola Jane Mulder, Manuela Pruess and Rolf Apweiler 55. Computational Prediction of P0rotein-Protein Interactions - Anton J. Enright, Lucy Skrabanek and Gary D. Bader 56. The Yeast Two-Hybrid System for Detecting Interacting Proteins Ilya G. Serebriiskii, Erica A. Golemis and Peter Uetz 57. Antibody-Affinity Purification to Detect Interacting Proteins - Sonia Navarro and Lucio Comai 58. Biomolecular Interaction Analysis Coupled with Mass Spectrometry to Detect Interacting Proteins - Setsuko Hashimoto, Toshiaki Isobe and Tohru Natsume 59. Assessment of Antibody-Antigen Interaction Using SELDI Technology - Li-Shen Hsieh, Ramy Moharram , Emilio Caputo and Brian M. Martin 60. Protein and Peptide Microarray-Based Assay Technology - Scott T. Clarke 61. Production of Protein Microarrays Using Robotic Pin Printing Technologies Ye Fang, Ann M. Ferrie and Fang Lai 62. PCR-Directed Protein In Situ Arrays - Joe Boutell and Mingyue He 63. Site-Specific Immobilization of Proteins in a Microarray - Yee Peng R. Lue, Su-Yin D. Yeo, Lay Pheng Tan, Grace Y.J. Chen and Shao Q. Yao 64. A Guide to Protein Interaction Databases - Tiffany B. Fischer, Melissa Paczkowski, Michael F. Zettel and Jerry Tsai 65. Deriving Function From Structure: Approaches and Limitations Annabel E. Todd 66. Comparative Protein Structure Modelling - M.S. Madhusudhan, Marc A. Marti Renom, Narayanan Eswar, Bino John, Ursula Pieper, Rachel Karchin, Min-Yi Shen and Andrej Sali 67. Classification of Protein Sequences and Structures - Shalom Rackovsky 68. How to Use Protein 1D Structure Predicted by PROFphd - Burkhard Rost 69. Classification of Proteins Folds - Rob Russell 70. Protein Threading - Andrew E. Torda 71. High Throughput Crystallography for Structural Proteomics Jeff Yon, Mladen Vinkovic and Harren Jhoti 72. Automated High-Throughput Protein Crystallization - Arezou Azarani 73. NMR-Based Structure Determination of Proteins in Solution - Andrzej Ejchart and Igor Zhukov