Cytokine Protocols - Methods in Molecular Biology 2004 edition

Jacket Description/Back: Because cytokines regulate many different bodily functions in living organisms, mainly at the level of cell-cell communications, there is great interest in understanding their mode of action. In Cytokine Protocols, established researchers, physicians, and clinicians present their best biochemical, cellular, and molecular techniques for unraveling and quantifying the events occurring between the initial contact of a cytokine at the membrane receptor and the eventual activation of gene transcription. Described in step-by-step detail to ensure successful experimental results, these protocols cover the large-scale generation and purification of plasmid DNA, the identification of DNA-protein interactions via the gel mobility shift assay, RNA-level phenomena, and the isolation and characterization of cytokines, cytokine-related proteins, and their interactions. The techniques used include the generation of transfectants, the immunohistochemical detection of cytokines in tissue sections, and optimized staining for cytoplasmic detection. Highlights include RT-PCR of small amounts of mRNA, in situ hybridization, biosensor analysis, measurement of biological activities and standardization, immunohistochemical and single-cell detection, and receptor isolation, characterization, and crystallization. Each protocol includes a background introduction, equipment and reagent lists, tips on troubleshooting and avoiding pitfalls, and, where needed, a discussion of the interpretation of results. Comprehensive and highly practical, Cytokine Protocols offers novice researchers a concise introduction to the principles of cytokine research, and experienced investigators all the advanced techniques and time-saving tips needed to ensure productive research. Table of Contents: Large-Scale Generation of Plasmids that Express Type I Interferon Peter Harle and Daniel J. J. Carr Identification of trans-Acting Factors by Electrophoretic Mobility Shift Assay Rebecca J. Chan, Min You, and Gen-Sheng Feng Modulation of the Interferon-g Signal by Transfection of Cells with an Antisense-RNA Expressing Vector Leen Delrue and Marc De Ley Competitive RT-PCR to Quantify Small Amounts of mRNA Gabriela Aust In Situ Hybridization for Cytokines in Human Tissue Biopsies Emily Mathey, John Pollard, and Patricia Armati Purification, Identification, and Synthesis of Chemokines Patricia Menten, Anja Wuyts, and Jo Van Damme Receptor Isolation and Characterization: From Protein to Gene Daniela Novick and Menachem Rubinstein Crystallization of Cytokine-Receptor Complexes William J. Cook and Mark R. Walter Biosensor Analysis of Receptor-Ligand Interactions Sheng-Jiun Wu and Irwin Chaiken Analysis of SH2 Ligands and Identification of Sites of Interaction Manuel Baca Assays for Antiviral Activity Anthony Meager Assays for Cytotoxicity Anthony Meager Assays for Chemotaxis Anja Wuyts, Patricia Menten, Nancy Van Osselaer, and Jo Van Damme In Vitro and In Vivo Assays for High-Molecular-Weight Pegylated Muteins of Granulocyte Colony-Stimulating Factor James F. Eliason, Nadine Tare, Sharon Bowen, Tomoaki Inoue, and Ikuro Horii Development of a Mammalian Tet-On Expression Cell Line: Glucosylceramide Synthase Regulates TNF-a-Induced Apoptosis Yong-Yu Liu and Myles C. Cabot Immunohistochemical Detection of Cytokines in Human Tissue Sections Sandra Hulshof, Danny F. Dukers, Wim Vos, and Corline J. A. De Groot Detection of Cytokine- and Chemokine-Expressing Cells at the Single Cell Level Karin Lore and JanAndersson Detection of Cytokine Signal Transduction 'Cross-Talk' in Leukocyte Activation Howard R. Petty Assay System for the Effect of Prostaglandin E2 in the Determination of Polarized Cytokine Production Etsushi Kuroda and Uki Yamashita Index"Publisher Marketing: In these 19 chapters, researchers, physicians, and clinicians describe each step of their techniques for examining and quantifying the events occurring between the initial contact of a cytokine at the membrane receptor and the eventual activation of gene transcription. The techniques used include th