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Full Description
This second edition volume expands on the previous edition with new protocols that offer biochemical, biophysical, and biological assays for studying macrophage migration inhibitory factor (MIF) and D-dopachrome tautomerase (D-DT). The chapters in this book cover topics such as generation of MIF peptides via solid phase peptide synthesis; fragment screening of MIF by surface plasmon resonance; MIF NMR chemical shift perturbation mapping; methods to determine the role of MIF in cilia biogenesis; and immunofluorescent quantification of MIF in larval zebrafish tailfin transfection. Written in the highly successful Methods in Molecular Biology series format, chapters include introductions to their respective topics, lists of the necessary materials and reagents, step-by-step, readily reproducible laboratory protocols, and tips on troubleshooting and avoiding known pitfalls.
Cutting-edge and comprehensive, Macrophage Migration Inhibitory Factor: Methods and Protocols, Second Edition is a valuable resource for any researcher hoping to contribute to the ever expanding world of MIF biology.
Contents
Expression and Purification of Recombinant Macrophage Migration Inhibitory Factor (MIF).- Generation of MIF Peptides via Solid Phase Peptide Synthesis.- Expression and Purification of Recombinant D-Dopachrome Tautomerase (D-DT).- Structural Determination and Ligand Interaction of Macrophage Inhibitory Factor: A Standardized X-Ray Crystallography Approach.- Generation of High-Quality D-Dopachrome Tautomerase Crystals for Structural Studies.- Fragment Screening of MIF by Surface Plasmon Resonance.- Detection of Noncovalent Protein-Ligand Complexes of D-DT Variants by MALDI-TOF Mass Spectrometry.- Analysis of D-Dopachrome Tautomerase Conformational Flexibility Through Molecular Dynamics Solutions.- MIF NMR Chemical Shift Perturbation Mapping.- Assessing the Tautomerase Activity of MIF Using L-Dopachrome.- Using 4-HPP and PP as Substrates to Assess Purified MIF and D-DT Tautomerase Activity and Inhibition.- Protocols for Assessing the Nuclease Activity of MIF.- Detection of Oxidized Macrophage Migration Inhibitory Factor (oxMIF) in Biological Samples Using Enzyme-Linked Immunosorbent Assay (ELISA).- Detection of Oxidized Macrophage Migration Inhibitory Factor (oxMIF) in Tissue Samples Using Immunohistochemistry (IHC).- Exploring the Chemotactic Effects of MIF on Neutrophils and T Cells and Methods for Selective Receptor Targeting.- Polarization of Bone Marrow-Derived Macrophages from Wild-Type and High Human MIF-Expressing CATT7 Polymorphic Mice.- Using Luciferase Reporter Cells to Assess the Effects of MIF Inhibitors on NF-kB Activation.- Monitoring Inflammasome Activity through ASC Speck Formation in THP-1 ASC-GFP Cells.- Small Interfering RNA (siRNA)-Mediated Knockdown of Macrophage Migration Inhibitory Factor Transcript in Adherent and Suspension Cultures of Human Cell Lines.- Stable Transfection of CD74 Gene Variants into A549 Lung Adenocarcinoma Cell Line.- Methods to Determine the Role of MIF in Cilia Biogenesis.- Modelling Fungal and Amoebic Infections: Co-Culture Approaches to Study MIF's Role in Infection Induced Inflammatory Responses.- Immunofluorescent Quantification of MIF in Larval Zebrafish Tailfin Transfection.
