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Full Description
This volume discusses the latest advancements in the field of antigen processing and presentation, and the complex assays and techniques that have been devised to help pick apart molecular events along this pathway. The chapters in this book cover topics such as how to make and analyze MHC molecules and their cargo; analyzing the function chaperones that regulate antigen presentation; enzymes that generate antigenic peptides; how to make and analyze T cell receptors; and presenting a series of functional assays to follow adaptive immune responses to antigen presentation. 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, Antigen Processing and Presentation: Methods and Protocols is a valuable resource for all researchers who are looking to carry the field forward, validate key findings, and help newcomers make meaningful contributions.
Contents
High-Throughput Production of Class I MHC Tetramers for Immunological Studies using Chaperone-Mediated Peptide Exchange.- Empty MHC Class I Protein Production.- Determining the Half-Life of MHC Class I Molecules by Blocking the Protein Synthesis Using Cycloheximide.- Probing Thermal Transitions of Peptide-Major Histocompatibility Complexes by Differential Scanning Fluorimetry.- Method for Measuring the Kinetic Stability of Peptides Bound onto MHC Class I using MALDI-TOF Mass Spectrometry.- Methods for Isolating and Analyzing the HLA Class I Immunopeptidome.- Expression and Functional Characterization of Recombinant HLA-DM.- Utilizing TAPBPR for Peptide Loading, Dissociation, and Exchange on Plasma Membrane-Expressed MHC-I.- Expression, Purification, and Functional Analysis Methods of Antigen-Processing Aminopeptidases ERAP1, ERAP2, and IRAP.- Methods and Guidelines for Analyzing the Conformational Dynamics of Proteins in the Antigen Presentation Pathway.- Guidelines for the Design of Custom Affinity-Based Probes for Metalloproteases.- Method for Producing Recombinant Soluble T-Cell Receptors.- Method for Performing T Cell Receptor-Peptide-Human Leukocyte Antigen Co-Complexation and Structural Analysis.- Method for Determining the Affinity of the T Cell Receptor for the Peptide-Human Leukocyte Antigen Complex.- Methods for Probing NK Cell Surveillance of Antigen Presentation.- Cellular Assays to Monitor Antigen Cross-Presentation In Vitro.- T Cell Assay to Track Presentation of an Influenza Hemagglutinin Epitope Presented by the MHC-II Protein HLA-DR1.- Transient Transfection and T Cell Activation in Assessment of Endoplasmic Reticulum Aminopeptidase 1 and 2 Peptide Trimming Function.- Methods for Analyzing Human T Cell Activation through TCR or CAR Engagement.
