Signal Transduction Protocols

In 1995, Signal Transduction Protocols, edited by David A. Kendall and Stephen J. Hill, was published in the Methods in Molecular Biology series. This second edition represents an update to that previous work with an emp- sis on new methodologies that have developed in the last few years. The goal, then and now, is to provide procedures written by experts with first-hand ex- rience in a detail that goes far beyond what is generally encountered in the methods section of most journals and thus actually permits a particular p- cedure to be replicated. In addition, we have had as a secondary goal the id- tification of protocols for the assay of general classes of signal transduction components that, ideally, can be adapted to the assay of any member of that class. The ability to do this has resulted in large part from the use of affini- based assays, the ease with which specific proteins can be specifically tagged, and an explosion in the availability of highly specific antibodies from comm- cial sources, especially antibodies raised against signaling proteins of human origin. The number of available approaches is, fortunately for those working in signaling research, far too great to fit within the confines of this volume, so hard choices as to what to include had to be made.

Includes supplementary material: sn.pub/extras

Klappentext
Carrying on the high standards of the much-acclaimed first edition, highly experienced investigators have extensively updated Signal Transduction Protocols with many of the new approaches that have been transforming the field. Included in this new edition are readily reproducible immunoassays, fluorescence-based assays, high-throughput methods, protein modification assays, lipid second messenger assays, and chromatin immunoprecipitation techniques. Wherever possible, protocols for the assay of general classes of signal transduction components have been identified so they can be adapted to the assay of any member of that class. The protocols follow the successful Methods in Molecular Biology™ series format, each one 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.
State-of-the-art and eminently practical, Signal Transduction Protocols, 2nd ed. offers novice and experienced researchers alike an indispensable set of standardized, but readily adaptable, approaches for the assay of key signaling molecules-an invaluable tool for all laboratories newly entering the field, expanding the breadth of previous investigations, or seeking to improve an existing approach.

Inhalt
Making Protein Immunoprecipitates.- Signal Transduction Inhibitors in Cellular Function.- Two-Dimensional Gel Electrophoresis for the Identification of Signaling Targets.- A High-Throughput Mammalian Cell-Based Transient Transfection Assay.- Determining Protein Half-Lives.- Assaying Protein Kinase Activity.- Comparative Phosphorylation Site Mapping From Gel-Derived Proteins Using a Multidimensional ES/MS-Based Approach.- Studies of Calmodulin-Dependent Regulation.- Measurement of Protein-DNA Interactions In Vivo by Chromatin Immunoprecipitation.- Characterization of Protein-DNA Association In Vivo by Chromatin Immunoprecipitation.- Nonradioactive Methods for Detecting Activation of Ras-Related Small G Proteins.- Nucleocytoplasmic Glycosylation, O-GlcNAc.- Techniques in Protein Methylation.- Assaying Lipid Phosphate Phosphatase Activities.- Assaying Phosphoinositide Phosphatases.- Assaying Phospholipase A2 Activity.- Measurement and Immunofluorescence of Cellular Phosphoinositides.- Measuring Dynamic Changes in cAMP Using Fluorescence Resonance Energy Transfer.- In Vivo Detection of Protein-Protein Interaction in Plant Cells Using BRET.- Revealing Protein Dynamics by Photobleaching Techniques.- Assaying Cytochrome c Translocation During Apoptosis.