What is monoclonal antibody 03

Animal Biotechnology Group 05/03/2011 Outline content • Introduction • Production of Monoclonal Antibodies • Advantage and Disadvantages of MAbs • Application Beginning of Monoclonal Era Georg Kohler and Cesar Milstein fuse mouse lymphocytes with neoplastic mouse plasma cells to yield hybridomas that produce specific antibodies This offers a limitless supply of monoclonal antibodies Monoclonal antibodies permit diagnostic tests that are specific, and function as probes Discovery of Monoclonal Antibodies Monoclonal antibodies were produced in mice using a technique described by Köhler and Milstein et al They were awarded the Nobel Prize in 1984 (along with Jerne) for their work What is monoclonal antibody Antibodies are proteins produced by the B lymphocytes of the immune system in response to foreign proteins, called antigens Monoclonal antibodies are proteins that are made in a laboratory These proteins are designed to attach to areas on the surface of cancer cells and interfere with their growth and spread Monoclonal antibodies are monospecific antibodies that are identical because they are produced by one type of immune cell that are all clones of a single parent cell Given (almost) any substance, it is possible to create monoclonal antibodies that specifically bind to that substance Monoclonal antibodies are similar to the antibodies your body naturally produces when you are exposed to bacteria or viruses, such as a cold or the flu (influenza) Two features of the antibody-epitope relationship are key to the use of monoclonal antibodies as a molecular tool •specificity the antibody binds only to its particular epitope •sufficiency the epitope can bind to the antibody on its own Production of monoclonal antibodies • Monoclonal antibodies are antibodies produced by one cell line which are specific to one target (antigen) • Antibodies derived from mouse hybridomas are of limited use as human therapeutics, since they produce an adverse immune reaction with repeated use • The term hybridoma is myeloma cell culture applied to fused cells resulting due to fusion of following two types of cells: An antibody producing lymphocyte cell, and A single myeloma cell which is capable of multiplying indefinitely These fused hybrid cells or hybridoma have the antibody producing capability inherited from lymphocytes and have the ability to grow continuously (immortal) like malignant cancer cells Production of monoclonal antibodies • Steps in the production of monoclonal antibodies using hybridoma technology: Immunize a rabbit or mouse through repeated injection of a specific antigen for the production of specific antibody, facilitated due to proliferation of the desired B cells Produce tumors in a mouse or a rabbit From the above two types of animals, culture separately spleen cells that produce specific antibodies, and myeloma cells that produce tumors Production of monoclonal antibodies Induce fusion of spleen cells to myeloma cells, using polyethylene glycol (PEG), to produce hybridoma; the hybrid cells are grown in selective hypoxanthine aminopterine thymidine (HAT) medium Screening and propagating • The hybridomas now are ready to be diluted and grown, thus obtaining a number of different colonies, each producing only one type of antibody • The desired antibodies from the different colonies are then tested for their ability to bind to the antigen (ELISA), and the most effective one is picked out ELISA to test Mab production • The dark blue spot represents a positive clone • The light blue spots are positive controls, the next two wells to the right are negative controls Purification of Antibodies • Before final purification, the cultures may be subjected to cell fractionation for enrichment of the antibody protein • In E.coli, the antibodies may be secreted in the periplasm, used for enrichment of antibody, so that further purification is simplified • Alternatively the antibodies may be purified from cell homogenate or cell debris obtained from the medium • Antibodies can be purified by techniques: Ion-exchange chromatography Antigen affinity chromatography Monoclonal antibodies are produced by Hybridoma technique Advantages of Monoclonal Ab • Single Mabs are chemically defined, are specific for a particular type of antigen and can be used for standardization of specific assays • The Ab created are 100% active antibodies => Possible of high specific activity of labelling in radioimmunoassay, enzyme linked immunosorbent assay… • MAbs can be used as immunosorbents for antigen purification Advantages of Monoclonal Ab • MAb can be mass produced, easy to manipulate and mostly inexpensive • Distinct antigenic cross reactivities can be defined and very useful in diagnosis Disadvantages of Monoclonal Ab • MAbs are too specific Limited amounts of MAb migh miss important cross-reactive determinant • Any physical or chemical treatment that affect one molecule will affect MAbs, causing deactivation of MAbs activities • Time consuming to produce MAbs, 3-4 months for each fusion experiment Monoclonal antibodies’s applications Diagnostic tests • Detecting small amounts of drugs, toxins or hormones Ex : monoclonal antibodies to human chorionic gonadotropin (HCG) are used in pregnancy test kits (Biotech, 1989) Another diagnostic uses of antibodies is the diagnosis of AIDS by the ELISA test, diagnosis of hepatitis, influenza, herpes, streptococcal, and Chlamydia infections - Can be used to detect for the presence and quantity of substances by The Western blot test and immuno dot blot tests Therapeutic treatment • Limitations : The rejection of monoclonal antibodies by the human immune system • Solution : Mice have been genetically engineered to produce antibodies that have human constant regions By using these hybrid, the immune system only "sees" a human protein and does not react against them • Cancer treatment : Monoclonal antibodies bind to cancer cell-specific antigens and induce an immunological response against the target cancer cell Monoclonal antibodies can also be used as delivery vehicles, guiding radioactive molecules or toxins to the cancer cell Antibodies attached to a cell can trigger an immune response that destroys the cell Group member • Phạm Anh Dũng • Nguyễn Đức Thiện An • Lê Thị Kim Diệu • Ngơ Huỳnh Thế Hải • Nguyễn Việt Tuấn ... antibodies This offers a limitless supply of monoclonal antibodies Monoclonal antibodies permit diagnostic tests that are specific, and function as probes Discovery of Monoclonal Antibodies Monoclonal. .. Jerne) for their work What is monoclonal antibody Antibodies are proteins produced by the B lymphocytes of the immune system in response to foreign proteins, called antigens Monoclonal antibodies... of Monoclonal Ab • MAb can be mass produced, easy to manipulate and mostly inexpensive • Distinct antigenic cross reactivities can be defined and very useful in diagnosis Disadvantages of Monoclonal