Monoclonal Antibody Protocols

Monoclonal Antibody Protocols provides researchers in biomedical, agricultural, and biological science with a set of detailed, easy-to-follow methods for developing and using monoclonal antibodies. The protocols emphasize techniques that optimize the outgrowth of hybridomas from primary cultures of fused cells and the use of an alternative, electric-field-mediated cell fusion technique to increase the yield of hybridomas. The book stresses antibodies produced in mice, but includes methods of producing xenogeneic hybrids that yield human, bovine, equine, and porcine monoclonal antibodies. With its detailed instructions, its comments on how to alter the various steps of a protocol in order to accommodate different materials, and its hints and tips that often make the difference between success and failure, Monoclonal Antibody Protocols provides you with a ready and indispensable source of information for preparing and using monoclonal antibodies successfully in your laboratory.

Monoclonal Antibodies: Methods and Protocols, Second Edition expands upon the previous edition with current, detailed modern approaches to isolate and characterize monoclonal antibodies against carefully selected epitopes. This edition includes new chapters covering the key steps to generate high quality monoclonals via different methods, from antigen generation to epitope mapping and quality control of the purified IgG. Chapters are divided into four parts corresponding to four distinct objectives. Part I covers monoclonal antibody generation, Part II deals with monoclonal antibody expression and purification, Part III presents methods for monoclonal antibody characterization and modification, and Part IV describes selected applications of monoclonal antibodies. 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. Authoritative and practical, Monoclonal Antibodies: Methods and Protocols, Second Edition provides crucial initial steps of monoclonal antibody generation and characterization with state-of-the art protocols.

This book examines a collection of state-of-the-art methods that employ monoclonal antibodies in a clinical setting. The chapters offer in-depth description for generating mouse and recombinant humanized antibodies, and a comprehensive review of how antibodies are being used in bead-based methods for measuring proteins. This field will continue to expand and provide new and innovative techniques in the laboratory and as a basis that complements targeted therapy.

The introduction of monoclonal antibodies revolutionized immunology. The development of human monoclonal antibodies was inspired primarily by the enormous clinical benefits promised by these reagents which can be used as anti-inflammatory reagents, anti-tumor reagents and reagents for passive immunization in a variety of pathologies. Human Monoclonal Antibodies: Methods and Protocols presents technical protocols of cellular and molecular methods for the production, purification and application of human monoclonal antibodies, as well as review articles on related topics of human monoclonal and polyclonal antibodies. Written in the 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 protocols, and notes on troubleshooting and avoiding known pitfalls. Authoritative and easily accessible, Human Monoclonal Antibodies: Methods and Protocols seeks to serve both professionals and novices with its well-honed methodologies which will prove invaluable in a clinical setting.

The American Anti-Vivisection Society (AAVS) petitioned the National Institutes of Health (NIH) on April 23, 1997, to prohibit the use of animals in the production of mAb. On September 18, 1997, NIH declined to prohibit the use of mice in mAb production, stating that "the ascites method of mAb production is scientifically appropriate for some research projects and cannot be replaced." On March 26, 1998, AAVS submitted a second petition, stating that "NIH failed to provide valid scientific reasons for not supporting a proposed ban." The office of the NIH director asked the National Research Council to conduct a study of methods of producing mAb. In response to that request, the Research Council appointed the Committee on Methods of Producing Monoclonal Antibodies, to act on behalf of the Institute for Laboratory Animal Research of the Commission on Life Sciences, to conduct the study. The 11 expert members of the committee had extensive experience in biomedical research, laboratory animal medicine, animal welfare, pain research, and patient advocacy (Appendix B). The committee was asked to determine whether there was a scientific necessity for the mouse ascites method; if so, whether the method caused pain or distress; and, if so, what could be done to minimize the pain or distress. The committee was also asked to comment on available in vitro methods; to suggest what acceptable scientific rationale, if any, there was for using the mouse ascites method; and to identify regulatory requirements for the continued use of the mouse ascites method. The committee held an open data-gathering meeting during which its members summarized data bearing on those questions. A 1-day workshop (Appendix A) was attended by 34 participants, 14 of whom made formal presentations. A second meeting was held to finalize the report. The present report was written on the basis of information in the literature and information presented at the meeting and the workshop.

The introduction of monoclonal antibodies revolutionized immunology. The development of human monoclonal antibodies was inspired primarily by the enormous clinical benefits promised by these reagents which can be used as anti-inflammatory reagents, anti-tumor reagents and reagents for passive immunization in a variety of pathologies. Human Monoclonal Antibodies: Methods and Protocols presents technical protocols of cellular and molecular methods for the production, purification and application of human monoclonal antibodies, as well as review articles on related topics of human monoclonal and polyclonal antibodies. Written in the 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 protocols, and notes on troubleshooting and avoiding known pitfalls. Authoritative and easily accessible, Human Monoclonal Antibodies: Methods and Protocols seeks to serve both professionals and novices with its well-honed methodologies which will prove invaluable in a clinical setting.

Includes all of the information required to produce monoclonal antibodies in the laboratory and to prepare them for use in a multitude of given applications. Production procedures are treated in chronological order, beginning with basic tissue culture techniques, immunization strategies and screening test design, followed by production of hybridoma cell lines and basic antibody characterization, purification and labeling. Each chapter contains explanatory text on each step with comparative analysis of methods where appropriate. All necessary experimental protocols are presented in a self-contained format that is easy to follow in the laboratory. Alternative protocols are provided where relevant; for others not included in full, source references are presented. Surveys the current status of human hybridoma production and antibody engineering using molecular biology techniques.

The development of the hybridoma technology created the possibility to obtain unlimited amounts of monoclonal antibodies (mAb) with high specificity and affinity for any target and to introduce mAbs in a wide range of applications; however, the bulky size of mAbs, costly production, and cumbersome engineering hampered regularly their streamlined development in some applications. In Single Domain Antibodies: Methods and Protocols, expert researchers examine single variable domain antibody fragments, referred to as VH, VL, VHH or VNAR. These fragments are the smallest intact antigen-binding fragments that can be produced recombinantly at low cost. Written in the highly successful Methods in Molecular BiologyTM 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.