2-D Proteome Analysis Protocols

ISBN-13: 9780896035249 / Angielski / Twarda / 1998 / 601 str.

Andrew J. Link

Opis

With the completion of sequencing projects and the advancement of a- lytical tools for protein identification, proteomics the study of the expressed part of the genome has become a major region of the burgeoning field of functional genomics. High-resolution 2-D gels can reveal virtually all p- teins present in a cell or tissue at any given time, including posttranslationally modified proteins. Changes in the expression and structure of most cellular proteins caused by differentiation or external stimuli can be displayed and eventually identified using 2-D protein gels. 2-D Proteome Analysis Protocols covers all aspects of the use of 2-D protein electrophoresis for the analysis of biological problems. The contri- tors include many of the leaders in the fields of biochemistry and analytical chemistry who were instrumental in the development of high-resolution 2-D gels, immobilized pH gradients, computer analysis, and mass spectromet- based protein identification methodologies. This book is intended as a benchtop manual and guide both for novices to 2-D gels and for those aficionados who wish to try the newer techniques. Any group using protein biochemistry especially in the fields of molecular biology, biochemistry, microbiology, and cell biology should find this book eminently useful. 2-D Proteome Analysis Protocols takes the researcher through the c- plete process of working with 2-D protein gels from making the protein - tract to finally identifying the proteins of interest. It includes protocols for generating 2-D protein extracts from most of the standard model organisms, including bacteria, yeast, nematode, Drosophila, plants, mouse, and human."

Recenzje

"Our readers will find in this book with its 55 chapters, written by 78 highly well-trained specialists, all they need to come to a good result, going out from the theory of the subject to the precise description of the protocols, the instrumentation, to additive notes, the problems and the eventual difficulties. In one word, an excellent and practical book."-Cellular and Molecular Biology

"...There are 55 chapters covering everything you really need to know to get you to speed in the analysis of the proteome. I really liked the layout of each chapter with a pithy introduction followed by materials and methods sections...an absolute must for the proteomic laboratory."-Microbiology Today

". . .the book covers practical aspects , is well documented, and clearly written . . . recommended for researchers in the field. This collection of protocols is an invaluable starting point for people embarking on attempts to develop new proteomes."-FEBS Letters

"The wealth of techniques and helpful hints presented in this book should be useful to both neophytes and advanced researchers engaged in the area of high resolution 2-D protein electrophoresis."-Analytical Biochemistry

Spis treści

2-D Protein Gel Electrophoresis: An Overview, Jenny Fichmann and Reiner Westermeier. Solubilization of Proteins in 2-D Electrophoresis: An Outline, Thierry Rabilloud. Preparation of Escherichia coli Samples for 2-D Gel Analysis, Ruth A. VanBogelen and Frederick C. Neidhardt. Preparing 2-D Protein Extracts from Yeast, David M. Schieltz. 2-D Protein Extracts from Drosophila melanogaster, Christer Ericsson. Preparing 2-D Protein Extracts from Caenorhabditis elegans, Robert Zwilling. Eukaryotic Cell Labeling and Preparation for 2-D, Nick Bizios. Differential Detergent Fractionation of Eukaryotic Cells: Analysis by Two-Dimensional Gel Electrophoresis, Melinda L. Ramsby and Gregory S. Makowski. Fractionated Extraction of Total Tissue Proteins from Mouse and Human for 2-D Electrophoresis, Joachim Klose. Preparation and Solubilization of Body Fluids for 2-D, Jean Charles Sanchez and Denis F. Hochstrasser. 2-D Electrophoresis of Plant Proteins, Akira Tsugita and Masaharu Kamo. Quantifying Protein in 2-D PAGE Solubilization Buffers, Louis S. Ramagli. Measuring the Radioactivity of 2-D Protein Extracts, Andrew J. Link and Nick Bizios. Advantages of Carrier Ampholyte IEF, Mary F. Lopez. 2-D Electrophoresis Using Carrier Ampholytes in the First Dimension (IEF), Mary F. Lopez. Nonequilibrium pH Gel Electrophoresis (NEPHGE), Mary F. Lopez. High Resolution, 2-D Protein Electrophoresis Using Nondedicated Equipment, Marion Sarmiento. Large-Gel 2-D Electrophoresis, Joachim Klose. Advantages of Immobilized pH Gradients, Jenny Fichmann. Casting Immobilized pH Gradients (IPGs), Elisabetta Gianazza. Analytical IPG-Dalt, Angelika Görg and Walter Weiss. IPG-Dalt of Very Alkaline Proteins, Angelika Görg. Running Preparative Carrier Ampholyte and Immobilized pH Gradient IEF Gels for 2-D, Neil M. Matsui, Diana M. Smith-Beckerman, Jenny Fichmann, and Lois B. Epstein. In-Gel Sample Rehydration of Immobilized pH Gradient, Jean-Charles Sanchez, Denis Hochstrasser, and Thierry Rabilloud.High-Resolution, IPG-Based, Mini Two-Dimensional Gel Electrophoresis, Jean-Charles Sanchez and Denis F. Hochstrasser. Horizontal SDS-PAGE for IPG-Dalt, Angelika Görg and Walter Weiss. Casting and Running Vertical Slab-Gel Electrophoresis for 2D-PAGE, Bradley J. Walsh and Benjamin R. Herbert. Nonreducing 2-D Polyacrylamide Gel Electrophoresis, Hong Ji and Richard J. Simpson. 2-D Diagonal Gel Electrophoresis, Joan Goverman. 2-D Phosphopeptide Mapping, Hikaru Nagahara, Robert R. Latek, Sergei A. Ezhevsky, and Steven F. Dowdy. Internal Standards for 2-D, Andrew J. Link. Autoradiography of 2-D Gels, Andrew J. Link. Double-Label Analysis, Kelvin H. Lee and Michael G. Harrington. Silver Staining of 2-D Electrophoresis Gels, Thierry Rabilloud. Staining of Preparative 2-D Gels: Coomassie Blue and Imidazole-Zinc Negative Staining, Neil M. Matsui, Diana M. Smith-Beckerman, and Lois B. Epstein. Electroblotting of Proteins from 2-D Polyacrylamide Gels, Michael J. Dunn. Detection of Total Proteins on Western Blots of 2-D Polyacrylamide Gels, Michael J. Dunn. Protein Detection Using Reversible Metal Chelate Stains, Wayne F. Patton, Mark J. Lim, and David Shepro. Gylcoprotein Detection of 2-D Separated Proteins, Nicolle H. Packer, Malcolm S. Ball, and Peter L. Devine. Image Acquisition in 2-D Electrophoresis, Wayne F. Patton, Mark J. Lim, and David Shepro. Computer Analysis of 2-D Images, Ron D. Appel and Denis F. Hochstrasser. 2-D Databases on the World Wide Web, Ron D. Appel, Amos Bairoch, and Denis F. Hochstrasser. Comparing 2-D Electrophoresis Gels Across Internet Databases, Peter F. Lemkin. Constructing a 2-D Database for the World Wide Web, Ron D. Appel, Christine Hoogland, Amos Bairoch, and Denis F. Hochstrasser. Absolute Quantitation of 2-D Protein Spots, Steven P. Gygi and Ruedi Aebersold. Generating a Bacterial Genome Inventory: Identifying 2-D Spots by Comigrating Products of the Genome on 2-D Gels, Ruth A. VanBogelen. Immunoaffinity Identification of 2-DE Separated

O autorze

Andrew J. Link, Associate Professor of Microbiology & Immunology, Assistant Professor of Biochemistry. Vanderbilt University School of Medicine, Nashville, Tennessee.

Blurb

In 2-D Gel Proteome Analysis Protocols, Andrew Link and his expert collaborators take today's researchers step-by-step through the complete process of doing proteomics. With easy-to-follow instructions, complete with many helpful hints and explanations, leading investigators and pioneers in the field show how to make protein extracts, reproducibly run them on a 2-D gels, detect them, analyze the data, and precisely identify each protein. The book covers the latest methods of using carrier ampholytes in the 1st dimension, casting and running immobilized pH gradient 2-D gels, MALDI-TOF-based peptide mapping, automated tandem mass spectrometry, and nanoelectrospray ionization technology. For the 2nd dimension, there are methods for running flatbed or vertical gels and for protein detection using autoradiography, and Coomassie, silver, and reversible metal-chelate stains. The book is a perfect complement to the genome sequencing project for answering biological questions.

2-D Gel Proteome Analysis Protocols is the most complete guide for using proteomics to answer biological questions. Whether it is a question of global protein analysis or evaluating a cell's response to internal or external stimuli, the advanced methods described here will enable today's researchers better to understand how cells work and open new possibilities for drug discovery.