CELL BIOLOGY ASSAYS PROTEINS

Protein assay methods are used for protein identification with blood groups, cell surface markers, drugs and toxins. This text features comprehensive protocols essential for researchers studying various areas of biological and medical sciences. The techniques in this text are presented in a friendly step-by-step fashion, providing useful tips and potential pitfalls while enabling researchers at all stages to embark on basic problems using a vareity of technologies and model systems.

• Focus on protein identification using mass spectrometry
• Step-by-step procedures detailing materials, procedures, comment and pitfalls
• Information on the plethora of technologies needed to tackle complex problems

1. Protein Determination
2. Phosphopeptide Mapping: A Basic Protocol
3. Coupling of Fluorescent Tags to Proteins
4. Radioiodination of Proteins and Peptides
5. Free-Flow Electrophoresis
6. Gel-Based Proteomics: High-Resolution Two-Dimensional Gel Electrophoresis of Proteins. Isoelectric Focusing and Nonequalibrium pH Gradient Electrophoresis.
7. High-Resolution Two-Dimensional Electrophoresis with Immobilized pH Gradients for Proteome Analysis
8. Two-Dimensional Difference Gel Electrophoresis: Application for the Analysis of Differential Protein Expression in Multiple Biological Samples
9. Affinity Electrophoresis for Studies of Biospecific Interations: High-Resolution Two-Dimensional Affinity Electrophoresis for Separation of Hapten-Specific Polyclonal Antibodies into Monoclonal Antibodies in Murine Blood Plasma
10. Image Analysis and Quantitation
11. Fluorescence Detection of Proteins in Gels Using SYPRO Dyes
12. Autoradiography and Fluorography: Film-Based Techniques for Imaging Radioactivity in Flat Samples
13. Two-Dimensional Gel Profiling of Posttanslationally Modified Proteins by in vivo Isotope Labeling
14. Immunoprecipitation of Proteins under Nondenaturing Conditions
15. Nondenaturing Polyacrylamide Gel Electrophoresis as a Method for Studying Protein Interations: Applications in the Analysis of Mitochrondrial Oxidative Phosphorylation Complexes
16. Affinity purification with Natural Immobilized Ligands
17. Analysis of Protein-Protein Interactions by Chemical Cross-Linking
18. Peroxisomal Targeting as a Tool to Assess Protein-Protein Interactions
19. Biomolecular Interaction Analysis Mass Spectrometry
20. Blot Overlays with P-Labedled GST-Ras Fusion Proteins: Application to Mapping Protein-Protein Interaction Sites
21. Ligand Blot Overlay Assay: Detetion of Ca+2- and Small GTP-Binding Proteins
22. Modular Scale Yeast Two-Hybrid Screening
23. Chromophore-Assisted Laser Inactivation of Proteins by Antibodies Labeled with Malachite Green
24. Chromatin Immunoprecipitation (ChIP)
25. Gel Mobility Shift Assay
26. DNA Affinity Chromatography of Transcription Factors: The Oligaonucleotide Trapping Approach
27. Protein Degradation Methods: Chaperone-Mediated Autophagy
28. Methods in Protein Ubiquitination
29. Protein Identification and Sequencing by Mass Spectrometry
30. Proteome Specific Sample Preparation Methods for Matrix-Assisted Laser Desorption/Ionization Mass Spectrometry
31. In-Gel Digestion of Protein Spots for Mass Spectrometry
32. Peptide Sequencing by Tandem Mass Spectrometry
33. Direct Database Searing Using Tandem Mass Spectra of Peptides
34. Identification of Proteins from Organisms with Unsequenced Genomes by Tandem Mass Spectrometry and Sequence-Similarity Database Searching Tools
35. Identification of Protein Phosphorylation Sites by Mass Spectrometry
36. Analysis of Carbohydrates/Glycoproteins by Mass Spectrometry
37. Stable Isotope Labeling by Amindo Acids in Cell Culture for Quantitative Proteomics
38. Site-Specific, Stable Isotopic Labeling of Cysteinyl Pepties in Complex Peptide Mixtures
39. Protein Hydrogen Exchange measured by Electrospry Ionization Mass Spectrometry
40. Nongel Based Proteomics: Selective Reversed-Phase Chromatographic Isolation of Methionine-Containing Peptides from Complex Peptide Mixtures
41. Mass Spectrometry in Noncovalent Protein Interactions and Protein Assemblies

Married, with three children and one cat (Max William), Dr. Celis currently resides in Denmark where he holds among his many titles the chair to the Symposium and Prize Committee of the Danish Cancer Society, as well as the acting Chairman of the Nordic Molecular Biology Association (NOMBA). Dr. Celis is currently the Vice President of the European Molecular Biology Laboratory (EMBL) Council and leader of the Danish Delegation to the OECD Megascience Forum subgroup on Bioinformatics. He is also the European Union Observer to the International Nucleotide sequence database collaboration and a member of the EMBO Course Committee.