Methods of Immunization to Enhance the Immune Response to Specific Antigens In Vivo in Preparation for Fusions Yielding Monoclonal Antibodies -- Methods of Immunization to Enhance the Immune Response to Specific Antigens In Vitro -- Culture Conditions that Optimize Outgrowth of Hybridomas -- Production of Stable Heterohybridomas Producing Human Monoclonal Antibodies -- Production of Stable Bovine-Murine Interspecies Hybrids -- Production of Monoclonal Antibodies in Swine -- Production of Monoclonal Antibodies in Horses -- Use of Electric-Field-Mediated Cell Fusion to Produce Hybridomas Secreting Monoclonal Antibodies -- Rapid Isotyping of Mouse Monoclonal Antibodies -- Methods of Measuring Antibodies in Sera/Plasma, Ascites, and Tissue-Culture Media -- Measurement of Immunoglobulin Synthesis Using the ELISPOT Assay -- Use of Monoclonal Antibodies in Immunohistochemistry -- Use of Monoclonal Antibodies in Immunoelectron Microscopy -- Use of Monoclonal Antibodies for Western Blotting with Enhanced Chemiluminescent Detection -- The Use of Directly and Indirectly Labeled Monoclonal Antibodies in Flow Cytometry -- Use of Monoclonal Antibodies and Flow Cytometry to Cluster and Analyze Leukocyte Differentiation Molecules -- In Vivo and In Vitro Production of Monoclonal Antibodies -- Separation of Monoclonal Antibodies from Cell-Culture Supernatants and Ascites Fluid Using Thiophilic Agarose -- Detection, Purification, and Utilization of Murine Monoclonal IgM Antibodies -- Preparation and Use of Immunoaffinity Columns with Monoclonal Antibodies Without Purification from Ascites and Tissue-Culture Medium -- Antigen Purification by Monoclonal Antibody Immunoaffinity Chromatography -- Coupling of Monoclonal Antibodies with Fluorophores -- Coupling of Monoclonal Antibodies with Biotin -- Coupling of Monoclonal Antibodies with Enzymes -- Use of Monoclonal Antibodies with Magnetic Particles to Separate Cell Subpopulations by Negative Selection -- Use of Monoclonal Antibodies with Magnetic Particles to Separate Cell Subpopulations by Positive Selection.

Since the initial description of techniques to immortalize anti­ body-producing B-lymphocytes by fusion with tissue culture-adapted myeloma cells, methods have been developed to produce monoclonal antibodies of defined specificity in multiple animal species. Stable hybrids can be readily produced in mice using a number of myeloma and hybridoma cell lines. To obviate the problem of identifying fusion partners in other animal species, xenohybrids have been produced using B-lymphocytes from the relevant species and mouse myeloma cells. The use of xenohybrids has minimized the problem of obtain­ ing stable antibody-producing hybrids in all species examined thus far. Although alternative techniques are being developed to produce monoclonal antibodies by molecular methods, hybridoma technol­ ogy will remain the technology of choice for producing monoclonal antibodies for a variety of applications in research and industry. The objective of Monoclonal Antibody Protocols is to provide investigators with a set of methods for producing and using mono­ clonal antibodies in biomedical, agricultural, and biological sciences. The book is not intended to provide methodology for all possible applications, but rather a series of methods presented in an easy-- follow format that can be used by new and established investiga­ tors, graduate and postgraduate fellows, and technical staff.