Basic Methods in Protein Purification and Analysis: A Laboratory Manual

by Simpson, Richard J; Adams, Peter D; Golemis, Erica A

• ISBN13: 9780879698676
• ISBN10: 0879698675
• Edition: 1st
• Format: Paperback
• Copyright: 2008-09-30
• Publisher: Cold Spring Harbor Laboratory Press

Summary

"This new manual is designed for routine, day-to-day use at the bench. By integrating both established in vitro and in vivo molecular techniques with more modern in silico methods, this manual takes the user from the initial steps of obtaining cellular and subcellular extracts, through the purification and isolation steps appropriate for the protein of interest, and, finally, to the steps involved in characterizing and identifying proteins, protein complexes, and protein-protein interactions. These step-wise methods are combined with troubleshooting advice and guidance on selecting an appropriate experimental strategy, making this new manual a handy yet informative resource for bench scientists with all levels of experience."--BOOK JACKET.

Table of Contents

A Note from the Publisher
Introduction to Protein Purification Strategies	p. 1
Preparation of Cellular and Subcellular Extracts	p. 39
Purification Using Gel Electrophoresis
One-dimensional Polyacrylamide Gel Electrophoresis	p. 79
Preparative Two-dimensional Gel Electrophoresis with Immobilized pH Gradients	p. 113
Purification Using Column Chromatography
Ion-exchange Chromatography	p. 135
Size-exclusion Chromatography	p. 153
Reversed-phase High-performance Liquid Chromatography	p. 171
Affinity and Immunoaffinity Chromatography	p. 191
Metal Chelate Affinity Chromatography	p. 209
Multidimensional Chromatography of Intact Proteins	p. 231
Characterizing Protein Complexes
Identification of Protein-Protein Interactions by Conventional Column Chromatography	p. 245
Identification of Associated Proteins by Coimmunoprecipitation	p. 265
Chromatin Immunoprecipitation of Protein Complexes	p. 279
Identification of Protein-Protein Interactions with Glutathione-S-Transferase Fusion Proteins	p. 293
Direct Visualization of Protein Interactions in Living Cells Using Bimolecular Fluorescence Complementation Analysis	p. 315
In Silico Tools: Analysis for Creating Focused Interaction Networks	p. 327
Measuring the Concentration of Proteins	p. 347
Concentrating Solutions of Proteins	p. 369
Stabilization of Proteins for Storage	p. 385
Immunoblotting and Staining of Membrane-bound Proteins	p. 401
General Cautions	p. 417
Index	p. 425
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