Antibody Engineering Protocols

About this book

This comprehensive collection of recently developed methods for producing new antibody reagents by immunization and recombinant DNA techniques contains ready-to-use protocols that illuminate current areas of research on antibody structure, functions, and applications. The methods can be applied in basic immunological studies involving antibody specificity, catalysis, and evolution, and in the isolation of rare antibodies by phage display technology and the engineering of new antibodies by mutagenesis. They offer insight into new ways of developing clinically useful antibody reagents. Antibody Engineering Protocols constitutes a single-source volume for laboratory investigators who want to minimize extensive literature and methodology searches and to work productively in their fields with reproducible step-by-step protocols.

Contents

SEQHUNT: A Program to Screen Aligned Nucleotide and Amino Acid Sequences. Molecular Modeling of Antibody-Combining Sites. Structure and Properties of Human Immunoglobulin Light-Chain Dimers. Crystallographic and Chromatographic Methods for Study of Antibody Light Chains and Other Proteins. Detection of Human Variable Gene Family Expression at the Single-Cell Level. Purification of Reduced and Alkylated Antibody Subunits. Murine Monoclonal Antibody Development. Comparative Properties of Polyclonal and Monoclonal Antibodies. Evaluation of Antibody Clonality. Affinity Immunoblotting. Epitope and Idiotope Mapping Using Monoclonal Antibodies. Anti-Idiotypic Antibodies That Mimic Opioids. Catalytic Antibodies: Structure and Possible Applications. Preparation and Assay of Acetylcholinesterase Antibody. DNA Hydrolysis by Antibodies. Screening Strategies for Catalytic Antitransition-State Analog Antibodies. Expression of Chimeric Immunoglobulin Genes in Mammalian Cells. Single-Chain Anti-DNA Fv. Molecular Cloning of Antiground-State Proteolytic Antibody Fragments. Cloning and Bacterial Expression of an Esterolytic sFv. Site-Directed Mutagenesis of Antibody-Variable Regions. Synthetic Antibody Gene Libraries for In Vitro Affinity Maturation. Chaperonins in Phage Display of Antibody Fragments. Phage-Display Libraries of Murine and Human Antibody Fab Fragments. Selection of Human Immunoglobulin Light Chains from a Phage-Display Library. Purification of Antibody Light Chains by Metal Affinity and Protein L Chromatography. Rapid Purification of Recombinant Antibody Fragments for Catalysis Screening. Assay of Radiolabeled VIP Binding and Hydrolysis by Antibodies. Methods of Measuring Thyroglobulin and Peptide-Methylcoumarinamide Hydrolysis by Autoantibodies. Radiolabeling of Antibodies for Therapy and Diagnosis. Index.

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