Methods of Hybridoma Formation


Book Description

Laymen often consider modern laboratory research to be based on an endless array of sophisticated technologies whose complex capabilities are as important to the outcome of any project as the inventiveness and creativity of the scientists who employ them. Scientists at times may share this point of view until they are con fronted by unexpected findings that demand new approaches, and they discover that yesterday's "sophisticated tools" are today's "blunt instruments." This experience provides a more sobering view of the current state of our scientific methods. It also serves as an impetus for the further development of technology that prepares us for the next stage of advance. Immunologists were confronted by such a technological crises in the late 1970s when they finally were forced to admit that poly clonal antibodies, although quite sensitive reagents, were not spe cific enough to answer many of the questions then confronting virologists and tumor biologists. The answer to the need for specific ity came with the development of monoclonal antibody technology. In the last ten years there have been considerable advances in monoclonal antibody techniques. Today these reagents are much more versatile than they were initially and can be applied to a broad range of problems. Still, most workers who are using these anti bodies are convinced that their potential is far from exhausted, and that at least in some fields we are currently in the early stages of learning how to use them properly.




Monoclonal Antibody Production


Book Description

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.







Bispecific Antibodies


Book Description

The concept of using bispecific antibodies for cancer therapy by retargeting immune effector cells was developed several years ago. Initial clinical studies were rather disappointing mainly due to low efficacy, severe side effects and the immunogenicity of the bispecific antibodies. The progress in antibody engineering finally led to the generation of new classes of bispecific antibodies lacking these obstacles. In addition, new applications were established, such as pre-targeting strategies in radioimmunotherapy and dual targeting approaches in order to improve binding, selectivity and efficacy. In this book, the different ways of generating bispecific antibodies are described, with emphasis on recombinant formats. The various applications of bispecific antibodies, e.g. in cellular cancer immunotherapy, radioimmunotherapy and pretargeting strategies are covered, and emerging applications such as dual targeting strategies, which involve the simultaneous inhibition of two targets, are addressed.




Biosimilars of Monoclonal Antibodies


Book Description

Addressing a significant need by describing the science and process involved to develop biosimilars of monoclonal antibody (mAb) drugs, this book covers all aspects of biosimilar development: preclinical, clinical, regulatory, manufacturing. • Guides readers through the complex landscape involved with developing biosimilar versions of monoclonal antibody (mAb) drugs • Features flow charts, tables, and figures that clearly illustrate processes and makes the book comprehensible and accessible • Includes a review of FDA-approved mAb drugs as a quick reference to facts and useful information • Examines new technologies and strategies for improving biosimilar mAbs




Monoclonal Antibodies


Book Description

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.




ELISA


Book Description

This volume is a practical biochemical guide to the Enzyme-Linked Immunosorbent Assay (ELISA), used to detect a target substance in a liquid sample. The ELISA is an important and widely used diagnostic tool in medicine, animal health, botany and quality assurance processes in food and beverage production. An introductory chapter orients the reader on the basic structure and function of immunoglobulins and their fragments while subsequent chapters outline the methodology to generate monoclonal antibodies using hybridoma technology and the general methods used to purify antibodies. Multiple chapters demonstrate how to creatively use the properties of the antibody to identify, localize and quantify target analytes to answer questions and resolve problems. The reader will learn how to use a variety of immunoassay strategies, reporters and detection systems that will undoubtedly facilitate their efforts to gain answers to their own questions. 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, ELISA: Methods and Protocols seeks to provide both professionals and novices with the technical information necessary for the reader to successfully use the immunoassay as part of the discovery process.




Introduction to Cell and Tissue Culture


Book Description

It is a pleasure to contribute the foreword to Introduction to Cell and Tissue Culture: The ory and Techniques by Mather and Roberts. Despite the occasional appearance of thought ful works devoted to elementary or advanced cell culture methodology, a place remains for a comprehensive and definitive volume that can be used to advantage by both the novice and the expert in the field. In this book, Mather and Roberts present the relevant method ology within a conceptual framework of cell biology, genetics, nutrition, endocrinology, and physiology that renders technical cell culture information in a comprehensive, logical for mat. This allows topics to be presented with an emphasis on troubleshooting problems from a basis of understanding the underlying theory. The material is presented in a way that is adaptable to student use in formal courses; it also should be functional when used on a daily basis by professional cell culturists in a- demia and industry. The volume includes references to relevant Internet sites and other use ful sources of information. In addition to the fundamentals, attention is also given to mod ern applications and approaches to cell culture derivation, medium formulation, culture scale-up, and biotechnology, presented by scientists who are pioneers in these areas. With this volume, it should be possible to establish and maintain a cell culture laboratory devot ed to any of the many disciplines to which cell culture methodology is applicable.




Adverse Reactions to Biologics


Book Description

In many areas of medicine physicians still face the great challenge of finding therapies that will meet the patients’ needs. In dermatology the challenge has arisen on multiple fronts through advances in the understanding of the immunopathogenesis of many inflammatory and malignant cutaneous disorders. Breakthroughs, combined with significant developments in targeted immunotherapy, have resulted in improved outcomes as these newer therapies are being used for both approved indications and as off-label therapies for various chronic inflammatory skin disorders and many forms of skin cancer. In the expectation that by truly understanding the safety profile of these targeted therapies patients’ outcomes will be significantly improved, this book offers insights into topics such as adverse reactions, infectious complications and the perioperative use of biologics in psoriasis, immunogenicity of biologic therapies, paradoxical reactions, safety of biologics used to treat autoimmune bullous diseases and primary cutaneous lymphomas, adverse reactions and skin manifestations of therapies targeting melanoma and non-melanoma skin cancer and other neoplastic diseases. Eminent researchers with extensive clinical experience have contributed to this publication, providing an in-depth overview of the latest knowledge in this field.




Cell Line Development


Book Description

Mammalian cell lines command an effective monopoly for the production of therapeutic proteins that require post-translational modifications. This unique advantage outweighs the costs associated with mammalian cell culture, which are far grater in terms of development time and manufacturing when compared to microbial culture. The development of cell lines has undergone several advances over the years, essentially to meet the requirement to cut the time and costs associated with using such a complex hosts as production platforms. This book provides a comprehensive guide to the methodology involved in the development of cell lines and the cell engineering approach that can be employed to enhance productivity, improve cell function, glycosylation and secretion and control apoptosis. It presents an overall picture of the current topics central to expression engineering including such topics as epigenetics and the use of technologies to overcome positional dependent inactivation, the use of promoter and enhancer sequences for expression of various transgenes, site directed engineering of defined chromosomal sites, and examination of the role of eukaryotic nucleus as the controller of expression of genes that are introduced for production of a desired product. It includes a review of selection methods for high producers and an application developed by a major biopharmaceutical industry to expedite the cell line development process. The potential of cell engineering approch to enhance cell lines through the manipulation of single genes that play important roles in key metabolic and regulatory pathways is also explored throughout.