Monoclonal antibodies are identical antibodies produced by a single clone of B cells that recognize a specific epitope. They are produced through the hybridoma technology which involves fusing antibody producing B cells with myeloma cells to form a hybridoma cell line. This allows for the mass production of antibodies that recognize a single epitope. Monoclonal antibodies have various applications including use in biochemical analysis through techniques like ELISA and RIA. They can also be used for diagnostic imaging by labeling them with radioisotopes.
Monoclonal antibodies are identical antibodies produced by one type of immune cell that are clones of a single parent cell. They are produced using hybridoma technology which involves fusing antibody producing B cells from an immunized animal with myeloma tumor cells to create a hybridoma cell line. This hybridoma cell line is capable of indefinite division in culture while producing the same monoclonal antibody. The monoclonal antibodies are then purified from the culture supernatant and have various diagnostic and therapeutic applications such as cancer treatment.
This document discusses edible vaccines, which involve genetically engineering plants to produce vaccine antigens that can be delivered orally through consumption of plant parts. Some key points:
- Edible vaccines could provide low-cost, easy to administer vaccines that don't require cold storage. Several plant species have been investigated for their ability to produce vaccine antigens.
- Early research demonstrated that transgenic potatoes, tobacco plants, and other species could produce antigens for diseases like hepatitis B, cholera, rabies, and norovirus when eaten, and stimulated protective immune responses in animal models.
- Advantages of edible vaccines include low-cost mass production and distribution without need for medical personnel. However, challenges remain around consistent dos
To synthesize a live attenuated vaccine, the disease-causing organism is grown under special laboratory conditions ,Vaccine production and purification
Introduction
History
Antibody
Catalytic antibodies
Structure of antibodies
Different strategies for catalytic antibodies
Application
Conclusion
References
Hybridoma technology and production of monoclonal antibodyRajpal Choudhary
Hybridoma technology allows for the mass production of monoclonal antibodies. It involves fusing antibody-producing B cells from the spleen of an immunized mouse with myeloma cancer cells, creating a hybridoma cell. These hybridoma cells are selected using HAT medium, which causes them to continuously secrete identical monoclonal antibodies. Monoclonal antibodies have many medical applications, including diagnosing diseases and infections through detection of specific antigens. For example, pregnancy tests detect the HCG hormone using monoclonal antibodies, and HIV tests detect HIV antibodies in blood serum using a multi-step process with monoclonal antibodies.
Students of medical and allied subjects must be exposed to the concept of monoclonal antibodies for the efficient practice of clinical and laboratory medicine.
Hybridoma Technology ( Production , Purification , and Application ) Sakshi Ghasle
Hybridoma technology revolutionized the field of immunology by enabling the production of monoclonal antibodies with high specificity and affinity. This presentation delves into the principles of DNA hybridoma technology, highlighting its significance in antibody production, therapeutic applications, and biomedical research. Learn about the key steps involved in generating hybridomas, from immunization to antibody screening, and discover the potential of recombinant DNA techniques in enhancing antibody engineering. Whether you're a student, researcher, or industry professional, this overview will provide valuable insights into the innovative world of hybridoma technology."
Uncover the wide-ranging applications of monoclonal antibodies in areas such as cancer therapy, autoimmune diseases, infectious diseases, and beyond. Learn about the latest advancements in antibody engineering and the development of novel therapeutic modalities, including bispecific antibodies, antibody-drug conjugates, and immune checkpoint inhibitors.
Whether you're a seasoned researcher or a newcomer to the field, this SlideShare presentation serves as a valuable resource for understanding the principles, techniques, and applications of hybridoma technology in modern biomedicine. Join a journey through the fascinating world of monoclonal antibodies and the groundbreaking science behind their creation.
Unlock the potential of hybridoma technology and propel your research to new heights. Dive into this SlideShare presentation now and explore the limitless possibilities of monoclonal antibody production with hybridoma technology.
Immunoblotting assays such as Western blotting allow detection of specific proteins in complex mixtures by separating proteins by gel electrophoresis, transferring them to a membrane, and using antibodies to identify target proteins on the membrane. The presentation provides details on the key steps of tissue preparation, gel electrophoresis, protein transfer, blocking, detection using labeled antibodies, analysis, and applications of immunoblotting assays like Western blotting.
Monoclonal antibodies are identical antibodies produced by one type of immune cell that are clones of a single parent cell. They are produced using hybridoma technology which involves fusing antibody producing B cells from an immunized animal with myeloma tumor cells to create a hybridoma cell line. This hybridoma cell line is capable of indefinite division in culture while producing the same monoclonal antibody. The monoclonal antibodies are then purified from the culture supernatant and have various diagnostic and therapeutic applications such as cancer treatment.
This document discusses edible vaccines, which involve genetically engineering plants to produce vaccine antigens that can be delivered orally through consumption of plant parts. Some key points:
- Edible vaccines could provide low-cost, easy to administer vaccines that don't require cold storage. Several plant species have been investigated for their ability to produce vaccine antigens.
- Early research demonstrated that transgenic potatoes, tobacco plants, and other species could produce antigens for diseases like hepatitis B, cholera, rabies, and norovirus when eaten, and stimulated protective immune responses in animal models.
- Advantages of edible vaccines include low-cost mass production and distribution without need for medical personnel. However, challenges remain around consistent dos
To synthesize a live attenuated vaccine, the disease-causing organism is grown under special laboratory conditions ,Vaccine production and purification
Introduction
History
Antibody
Catalytic antibodies
Structure of antibodies
Different strategies for catalytic antibodies
Application
Conclusion
References
Hybridoma technology and production of monoclonal antibodyRajpal Choudhary
Hybridoma technology allows for the mass production of monoclonal antibodies. It involves fusing antibody-producing B cells from the spleen of an immunized mouse with myeloma cancer cells, creating a hybridoma cell. These hybridoma cells are selected using HAT medium, which causes them to continuously secrete identical monoclonal antibodies. Monoclonal antibodies have many medical applications, including diagnosing diseases and infections through detection of specific antigens. For example, pregnancy tests detect the HCG hormone using monoclonal antibodies, and HIV tests detect HIV antibodies in blood serum using a multi-step process with monoclonal antibodies.
Students of medical and allied subjects must be exposed to the concept of monoclonal antibodies for the efficient practice of clinical and laboratory medicine.
Hybridoma Technology ( Production , Purification , and Application ) Sakshi Ghasle
Hybridoma technology revolutionized the field of immunology by enabling the production of monoclonal antibodies with high specificity and affinity. This presentation delves into the principles of DNA hybridoma technology, highlighting its significance in antibody production, therapeutic applications, and biomedical research. Learn about the key steps involved in generating hybridomas, from immunization to antibody screening, and discover the potential of recombinant DNA techniques in enhancing antibody engineering. Whether you're a student, researcher, or industry professional, this overview will provide valuable insights into the innovative world of hybridoma technology."
Uncover the wide-ranging applications of monoclonal antibodies in areas such as cancer therapy, autoimmune diseases, infectious diseases, and beyond. Learn about the latest advancements in antibody engineering and the development of novel therapeutic modalities, including bispecific antibodies, antibody-drug conjugates, and immune checkpoint inhibitors.
Whether you're a seasoned researcher or a newcomer to the field, this SlideShare presentation serves as a valuable resource for understanding the principles, techniques, and applications of hybridoma technology in modern biomedicine. Join a journey through the fascinating world of monoclonal antibodies and the groundbreaking science behind their creation.
Unlock the potential of hybridoma technology and propel your research to new heights. Dive into this SlideShare presentation now and explore the limitless possibilities of monoclonal antibody production with hybridoma technology.
Immunoblotting assays such as Western blotting allow detection of specific proteins in complex mixtures by separating proteins by gel electrophoresis, transferring them to a membrane, and using antibodies to identify target proteins on the membrane. The presentation provides details on the key steps of tissue preparation, gel electrophoresis, protein transfer, blocking, detection using labeled antibodies, analysis, and applications of immunoblotting assays like Western blotting.
This document contains lecture notes on major histocompatibility complex (MHC) and related topics from a biotechnology course. It discusses antigen-presenting cells, the structure and function of MHC class I and II molecules, similarities and differences between the two classes, MHC-associated genes, and important immune signaling molecules like cytokines, interleukins, interferons, and chemokines. Diagrams are included to illustrate MHC pathway and types of interferons. The notes provide an overview of key concepts in MHC and immunology for students in the biotechnology course.
Enzyme definition, Enzyme immobilization introduction , Enzyme immobilization definition, Explanation about support/ matrix, Examples about immobilized enzymes and their product, Advantages of immobilization, Applications of immobilization, Methods of immobilization in different categories like Adsorption method, Covalent bonding method, Entrapment method, Co polymerization /Cross linking method, Encapsulation method, Applications of immobilized enzymes, Diagrammatic explanation about methods of immobilization.
This document provides an introduction to animal cell culture by Dr. Anu P. Abhimannue. It discusses the history and development of animal cell culture from the early 20th century. It describes different types of animal cell culture such as primary versus secondary culture and finite versus continuous cell lines. It also discusses various cell culture methods like monolayer, suspension, types of culture vessels used and morphology of cultured cells. The document provides advantages and limitations of animal cell culture techniques.
Monoclonal antibodies are produced using hybridoma technology which involves fusing myeloma cells with antibody producing spleen cells from immunized mice. The fused cells or hybridomas are selected in HAT medium which allows only the fused cells to survive and grow. The resulting hybridoma cells secrete a single antibody clone that can be mass produced. Key steps include immunizing mice, isolating spleen and myeloma cells, fusing the cells to generate hybridomas, screening and selecting hybridomas that secrete the desired antibody clone, and culturing the selected hybridoma cells to produce monoclonal antibodies. Monoclonal antibodies find various therapeutic applications and can be purified using techniques like affinity chromatography.
This document discusses subunit and peptide vaccines. Subunit vaccines contain purified antigens from pathogens rather than whole pathogens. They often require adjuvants and multiple doses to provide long-lasting immunity. Peptide vaccines use short amino acid sequences from pathogens to stimulate immune responses. While they are stable and inexpensive to produce, peptides may not stimulate T-cells on their own and require carriers or adjuvants. The document outlines advantages and disadvantages of both subunit and peptide vaccines.
Hepatitis B is a viral infection that attacks liver cells and can cause chronic liver failure, liver cancer, and death. The hepatitis B vaccine contains the hepatitis B surface antigen (HBsAg) and is produced using genetically modified yeast cells. The HBsAg gene is isolated and inserted into a yeast expression system. The yeast then produces and secretes the recombinant HBsAg protein, which is purified and used in the hepatitis B vaccine to prevent hepatitis B infection. A course of three vaccine doses provides long-term immunity by establishing antibodies against HBsAg in the bloodstream.
Hybridoma
Hybridomas are cells that have been engineered to produce a desired antibody in large amounts, to produce monoclonal antibodies.
Monoclonal antibodies can be produced in specialized cells through a technique now popularly known as hybridoma technology.
Hybridoma technology was discovered in 1975 by two scientists, G. Kohler and C. Milstein, were awarded Noble prize for physiology and medicine in 1984.
An immobilized enzyme is an enzyme that has had its movement restricted by attaching it to a solid support. There are several reasons to immobilize enzymes, including protection from degradation, ability to reuse the enzyme for multiple reactions at a lower cost, and easy separation of products from enzymes. Common methods for immobilization include adsorption, entrapment, covalent binding, and cross-linking. Each method has advantages and disadvantages related to stability, activity loss, and reusability of the immobilized enzyme.
A vaccine is a biological preparation that improves immunity to a particular disease.
In the edible vaccine, Transgenic plants are used as vaccine production systems.
The genes encoding antigens of bacterial and viral pathogens can be expressed in plants in a form in which they retain native immunologic properties.
This document discusses cell culture contamination, including sources, types, signs, and methods for monitoring and preventing contamination. Common sources of contamination include failed sterilization of equipment, airborne particulates, and poorly maintained incubators. Major types of microbial contaminants are bacteria, molds, mycoplasma, yeasts, and viruses. Signs of contamination vary but may include cloudy cultures, pH changes, and visible microbes under a microscope. Proper aseptic technique and regular monitoring of cell cultures can help reduce contamination. Cross-contamination between cell lines must also be avoided.
Hybridoma technology is a method for producing large numbers of identical antibodies (also called monoclonal antibodies). This process starts by injecting a mouse (or other mammals) with an antigen that provokes an immune response.
A PERFECT BLEND OF INDUSTRIAL AND LABORATORY INFORMATION WITH FIRST HAND TECHNIQUES EXPLAINED IN DETAIL ABOUT VARIOUS FILTRATION TECHNIQUES, CHROMATOGRAPHY TECHNIQUES AND SEPRATION AND CELL LYSIS TECHNIQUE WITH ALL THE BASIC INFORMATION TO BEGINNERS
Immobilization of cells involves encapsulating cells in polymers to prevent division while maintaining viability. This allows increasing cell density in bioreactors for higher metabolite production. Methods include entrapment in gels, on fiber mats, or behind semi-permeable membranes. Alginate is commonly used for gel entrapment. Immobilized systems offer advantages like prolonged biomass use, higher product yields, and genetic stability. They also simplify downstream processing when products are extracellular. Various bioreactor designs are used depending on the immobilization method. Applications show enhanced production of metabolites like serpentine, anthraquinones, and capsaicin using immobilized plant cells.
Plasmids are circular DNA molecules that can exist independently of the bacterial chromosome and are used as cloning vectors. Cloning vectors are DNA molecules that can carry foreign genetic material into a host cell. Plasmids have features like cloning sites, selectable markers, and reporter genes that make them useful for cloning. Common types of cloning vectors include plasmids, bacteriophages, cosmids, and artificial chromosomes. Plasmids are advantageous as cloning vectors because they are small, can replicate independently of the host, and often confer antibiotic resistance, allowing for selection of transformed cells.
Use of microbes in industry. Production of enzymes-General consideration-Amyl...Steffi Thomas
Industrial uses of microbes, properties of useful industrial microbes, various industrial products, production of enzymes-general consideration-amylase, catalase, peroxidase, lipase, protease, penicillinase, procedure for culturing bacteria and inoculum preparation, submerged fermentation and solid state fermentation, uses of different enzymes
This document discusses genetic transformation, which is the alteration of a cell by incorporating exogenous DNA. It describes natural transformation in bacteria, which occurs when competent bacteria take up exogenous DNA from their environment. The document also discusses artificial transformation techniques used in labs, such as treating bacteria with calcium chloride to make them competent, then exposing them to exogenous DNA. It explains how the DNA enters the cell and can become incorporated into the genome. Selection methods for transformed cells, such as antibiotic resistance markers, are also summarized.
Monoclonal antibodies are produced from single clones of B cells and recognize a specific antigen. They have advantages over polyclonal antibodies like homogeneity, specificity, and unlimited production. Monoclonal antibodies are created through cell fusion between B cells and myeloma cells to form immortal hybridoma cells that produce the same antibody. They have various medical uses including cancer therapy, diagnostics, and treatment of autoimmune disorders.
This document contains lecture notes on major histocompatibility complex (MHC) and related topics from a biotechnology course. It discusses antigen-presenting cells, the structure and function of MHC class I and II molecules, similarities and differences between the two classes, MHC-associated genes, and important immune signaling molecules like cytokines, interleukins, interferons, and chemokines. Diagrams are included to illustrate MHC pathway and types of interferons. The notes provide an overview of key concepts in MHC and immunology for students in the biotechnology course.
Enzyme definition, Enzyme immobilization introduction , Enzyme immobilization definition, Explanation about support/ matrix, Examples about immobilized enzymes and their product, Advantages of immobilization, Applications of immobilization, Methods of immobilization in different categories like Adsorption method, Covalent bonding method, Entrapment method, Co polymerization /Cross linking method, Encapsulation method, Applications of immobilized enzymes, Diagrammatic explanation about methods of immobilization.
This document provides an introduction to animal cell culture by Dr. Anu P. Abhimannue. It discusses the history and development of animal cell culture from the early 20th century. It describes different types of animal cell culture such as primary versus secondary culture and finite versus continuous cell lines. It also discusses various cell culture methods like monolayer, suspension, types of culture vessels used and morphology of cultured cells. The document provides advantages and limitations of animal cell culture techniques.
Monoclonal antibodies are produced using hybridoma technology which involves fusing myeloma cells with antibody producing spleen cells from immunized mice. The fused cells or hybridomas are selected in HAT medium which allows only the fused cells to survive and grow. The resulting hybridoma cells secrete a single antibody clone that can be mass produced. Key steps include immunizing mice, isolating spleen and myeloma cells, fusing the cells to generate hybridomas, screening and selecting hybridomas that secrete the desired antibody clone, and culturing the selected hybridoma cells to produce monoclonal antibodies. Monoclonal antibodies find various therapeutic applications and can be purified using techniques like affinity chromatography.
This document discusses subunit and peptide vaccines. Subunit vaccines contain purified antigens from pathogens rather than whole pathogens. They often require adjuvants and multiple doses to provide long-lasting immunity. Peptide vaccines use short amino acid sequences from pathogens to stimulate immune responses. While they are stable and inexpensive to produce, peptides may not stimulate T-cells on their own and require carriers or adjuvants. The document outlines advantages and disadvantages of both subunit and peptide vaccines.
Hepatitis B is a viral infection that attacks liver cells and can cause chronic liver failure, liver cancer, and death. The hepatitis B vaccine contains the hepatitis B surface antigen (HBsAg) and is produced using genetically modified yeast cells. The HBsAg gene is isolated and inserted into a yeast expression system. The yeast then produces and secretes the recombinant HBsAg protein, which is purified and used in the hepatitis B vaccine to prevent hepatitis B infection. A course of three vaccine doses provides long-term immunity by establishing antibodies against HBsAg in the bloodstream.
Hybridoma
Hybridomas are cells that have been engineered to produce a desired antibody in large amounts, to produce monoclonal antibodies.
Monoclonal antibodies can be produced in specialized cells through a technique now popularly known as hybridoma technology.
Hybridoma technology was discovered in 1975 by two scientists, G. Kohler and C. Milstein, were awarded Noble prize for physiology and medicine in 1984.
An immobilized enzyme is an enzyme that has had its movement restricted by attaching it to a solid support. There are several reasons to immobilize enzymes, including protection from degradation, ability to reuse the enzyme for multiple reactions at a lower cost, and easy separation of products from enzymes. Common methods for immobilization include adsorption, entrapment, covalent binding, and cross-linking. Each method has advantages and disadvantages related to stability, activity loss, and reusability of the immobilized enzyme.
A vaccine is a biological preparation that improves immunity to a particular disease.
In the edible vaccine, Transgenic plants are used as vaccine production systems.
The genes encoding antigens of bacterial and viral pathogens can be expressed in plants in a form in which they retain native immunologic properties.
This document discusses cell culture contamination, including sources, types, signs, and methods for monitoring and preventing contamination. Common sources of contamination include failed sterilization of equipment, airborne particulates, and poorly maintained incubators. Major types of microbial contaminants are bacteria, molds, mycoplasma, yeasts, and viruses. Signs of contamination vary but may include cloudy cultures, pH changes, and visible microbes under a microscope. Proper aseptic technique and regular monitoring of cell cultures can help reduce contamination. Cross-contamination between cell lines must also be avoided.
Hybridoma technology is a method for producing large numbers of identical antibodies (also called monoclonal antibodies). This process starts by injecting a mouse (or other mammals) with an antigen that provokes an immune response.
A PERFECT BLEND OF INDUSTRIAL AND LABORATORY INFORMATION WITH FIRST HAND TECHNIQUES EXPLAINED IN DETAIL ABOUT VARIOUS FILTRATION TECHNIQUES, CHROMATOGRAPHY TECHNIQUES AND SEPRATION AND CELL LYSIS TECHNIQUE WITH ALL THE BASIC INFORMATION TO BEGINNERS
Immobilization of cells involves encapsulating cells in polymers to prevent division while maintaining viability. This allows increasing cell density in bioreactors for higher metabolite production. Methods include entrapment in gels, on fiber mats, or behind semi-permeable membranes. Alginate is commonly used for gel entrapment. Immobilized systems offer advantages like prolonged biomass use, higher product yields, and genetic stability. They also simplify downstream processing when products are extracellular. Various bioreactor designs are used depending on the immobilization method. Applications show enhanced production of metabolites like serpentine, anthraquinones, and capsaicin using immobilized plant cells.
Plasmids are circular DNA molecules that can exist independently of the bacterial chromosome and are used as cloning vectors. Cloning vectors are DNA molecules that can carry foreign genetic material into a host cell. Plasmids have features like cloning sites, selectable markers, and reporter genes that make them useful for cloning. Common types of cloning vectors include plasmids, bacteriophages, cosmids, and artificial chromosomes. Plasmids are advantageous as cloning vectors because they are small, can replicate independently of the host, and often confer antibiotic resistance, allowing for selection of transformed cells.
Use of microbes in industry. Production of enzymes-General consideration-Amyl...Steffi Thomas
Industrial uses of microbes, properties of useful industrial microbes, various industrial products, production of enzymes-general consideration-amylase, catalase, peroxidase, lipase, protease, penicillinase, procedure for culturing bacteria and inoculum preparation, submerged fermentation and solid state fermentation, uses of different enzymes
This document discusses genetic transformation, which is the alteration of a cell by incorporating exogenous DNA. It describes natural transformation in bacteria, which occurs when competent bacteria take up exogenous DNA from their environment. The document also discusses artificial transformation techniques used in labs, such as treating bacteria with calcium chloride to make them competent, then exposing them to exogenous DNA. It explains how the DNA enters the cell and can become incorporated into the genome. Selection methods for transformed cells, such as antibiotic resistance markers, are also summarized.
Monoclonal antibodies are produced from single clones of B cells and recognize a specific antigen. They have advantages over polyclonal antibodies like homogeneity, specificity, and unlimited production. Monoclonal antibodies are created through cell fusion between B cells and myeloma cells to form immortal hybridoma cells that produce the same antibody. They have various medical uses including cancer therapy, diagnostics, and treatment of autoimmune disorders.
Monoclonal antibodies (MAbs) are antibodies that are directed against a single antigen. They can be produced through hybridoma technology which involves fusing antibody-producing B cells with myeloma cells to form a hybrid cell line. This document outlines the process for producing MAbs including immunizing an animal, fusing B cells with myeloma cells, selecting antibody-producing hybridomas, cloning and mass producing the antibodies. MAbs have various diagnostic and therapeutic applications for diseases.
Monoclonal antibodies are identical antibodies produced by a single B cell clone that recognize a specific epitope. They are produced through the fusion of B cells from an immunized animal with myeloma cells to form a hybridoma cell line. Monoclonal antibodies have various applications, including use in diagnostic tests to detect substances like hormones and tumor markers, diagnostic imaging by delivering radioisotopes to target areas, and directly targeting diseases or purifying proteins through immunoaffinity chromatography. Their specificity and ability to target single epitopes makes them useful research and medical tools.
This document discusses monoclonal antibody production and applications. It begins by defining monoclonal and polyclonal antibodies, noting that monoclonal antibodies are identical because they are derived from a single parent cell. It then covers the history of monoclonal antibody development, the monoclonal antibody production method involving mouse immunization and cell fusion, and the types and uses of monoclonal antibodies including diagnostic and cancer treatment applications. Potential side effects of monoclonal antibody therapy are also mentioned.
Monoclonal antibodies are identical antibodies produced by a single clone of B cells that recognize a specific epitope on an antigen. They are produced by fusing B cells from immunized animals with myeloma cells. This produces a hybridoma that can produce identical antibodies indefinitely. Monoclonal antibodies have advantages over polyclonal antibodies in being specific to a single epitope. They are used diagnostically and therapeutically in areas like cancer, chronic diseases, and infections. However, producing monoclonal antibodies is an expensive and time-consuming process.
Hybridoma technology involves fusing antibody-producing B cells with myeloma cells to create immortal hybridoma cells that produce monoclonal antibodies. Georges Kohler and Cesar Milstein developed this technique in 1975 and were awarded the 1984 Nobel Prize. Hybridoma cells are cultured using HAT medium to produce monoclonal antibodies that have various diagnostic, therapeutic, and research applications.
Various diagnostic tools now a days relied on the Hybriodoma technology and monoclonal antibodies,so this presentation will give some basic information about mAb and Hybridoma technology.
Hybridoma Technology & its application in fisheriesUday Das
The document discusses the process of producing monoclonal antibodies through hybridoma technology. It involves fusing antibody-producing B-lymphocytes with myeloma tumor cells to create a hybrid cell that can divide indefinitely and produce antibodies of a single specificity. The process includes immunizing mice, fusing their spleen cells with tumor cells, selecting antibody-producing hybridomas through HAT medium, and harvesting monoclonal antibodies through in vitro or in vivo methods for various medical applications. While this technology has improved disease diagnosis and treatment, it remains inefficient and labor-intensive.
Monoclonal antibodies (mAb or moAb) are antibodies that are made by identical immune cells that are all clones of a unique parent cell. Monoclonal antibodies can have monovalent affinity, in that they bind to the same epitope (the part of an antigen that is recognized by the antibody). In contrast, polyclonal antibodies bind to multiple epitopes and are usually made by several different plasma cell (antibody secreting immune cell) lineages. Bispecific monoclonal antibodies can also be engineered, by increasing the therapeutic targets of one single monoclonal antibody to two epitopes. Given almost any substance, it is possible to produce monoclonal antibodies that specifically bind to that substance; they can then serve to detect or purify that substance. This has become an important tool in biochemistry, molecular biology, and medicine. When used as medications, non-proprietary drug names end in -mab and many immunotherapy specialists use the word mab anacronymically.
This document provides an overview of monoclonal antibodies including their history, development, preparation, and applications. It discusses how monoclonal antibodies are produced through the fusion of B cells and myeloma cells to form hybridomas. The document outlines the key applications of monoclonal antibodies in diagnosis (e.g. biochemical analysis, imaging), therapy (e.g. cancer, transplantation), and protein purification. It also provides examples of commercially available monoclonal antibodies for various diseases and conditions.
Monoclonal antibodies are laboratory-made proteins that mimic the immune system's ability to fight pathogens. They are generated from a single B-cell clone and recognize a unique epitope on a single antigen. The process involves immunizing an animal to produce B cells that secrete antibodies. The B cells are then fused with myeloma cells to create a hybridoma that is selected if it secretes the desired antibody. The hybridoma can then be cultured indefinitely to produce large quantities of monoclonal antibodies, which have applications in identification, diagnosis, passive immunity, and cancer therapy.
What are antibodies?
An antibody is a protein used by immune system to identify and neutralize foreign agents like bacteria and viruses.
Each antibody recognizes a specific antigen unique to its target.
Monoclonal antibodies are produced through the fusion of antibody-producing B cells with myeloma cells, creating a hybridoma that produces antibodies with a single specificity. They have applications in diagnosis, therapy, protein purification, and other areas. For diagnosis, they are used in biochemical assays and diagnostic imaging. Therapeutically, they can directly target conditions or be linked to toxins/drugs to target delivery. They are also useful for purifying specific proteins through immunoaffinity chromatography. Other applications include catalytic monoclonal antibodies and using them for autoantibody fingerprinting to identify individuals.
The document discusses the principles of antibody production, including how antibodies are produced by B cells in response to antigens, the process of antibody production including antigen presentation, activation of helper T cells and B cells, proliferation of plasma cells, and the role of memory cells in the secondary response. It also covers the structure and classes of antibodies, as well as the production and uses of monoclonal and polyclonal antibodies in research, diagnosis and therapy.
Hybridoma technology allows for the production of monoclonal antibodies. It involves fusing antibody-producing B cells with myeloma cells to create a hybridoma cell that is immortal and produces antibodies targeted against a specific antigen. Georges Kohler and Cesar Milstein discovered this technique in 1975 and were later awarded the Nobel Prize for their work. Hybridoma cells are cloned to generate identical daughter cells that secrete monoclonal antibodies, which have various diagnostic, therapeutic, and research applications.
Monoclonal antibodies are antibodies that are identical because they come from identical immune cells that are clones of a unique parent cell. They are produced through hybridoma technology which involves fusing myeloma cells with antibody producing immune cells to produce immortal hybridoma cell lines. Monoclonal antibodies have various applications including diagnostic tests, purification of proteins, cancer therapy, and treatment of autoimmune and inflammatory diseases. Common types of monoclonal antibodies include murine, chimeric, humanized, and fully human antibodies.
- Georges Köhler and Cesar Milstein developed hybridoma technology in 1975 for which they received the Nobel Prize in 1984. Hybridoma technology involves fusing myeloma cells with antibody-producing immune cells to create immortal cell lines that produce monoclonal antibodies.
- Monoclonal antibodies are identical antibodies produced by a single clone and recognize a specific antigen, whereas polyclonal antibodies are derived from different cell lines.
- Hybridoma technology involves immunizing an animal, fusing its immune cells with myeloma cells, selecting and cloning hybridoma cells that produce the desired antibody, and characterizing and storing the monoclonal antibody produced. Monoclonal antibodies have important applications in diagnostics and treatment of diseases like cancer.
BIOTECHNOLOGY IS
CHALLENGING SUBJECT TO TEACH AND UNDERSTAND ......
ITS A VERY INTERESTING TO LEARN ABOUT HYBRIDOMA TECHNOLOGY .. THEIR PRODUCTION AND
APPLICATION ALSO ....
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Monoclonal antibody production by hybridoma technology
1. Presented by : Hasnat Tariq
Gulab Devi institute of Pharmacy (GDIP) Lahore , Pakistan
2. What are Antibodies?
• An antibody is a protein used by immune system to identify and neutralize foreign
objects like bacteria and viruses. Each antibody recognizes a specific antigen unique to its
target.
• The high specificity of antibodies makes them an excellent tool for detecting and
quantifying a broad array of targets, from drugs to serum proteins to microorganisms.
• With in vitro assays, antibodies can be used to precipitate soluble antigens, agglutinate
(clump) cells, opsonize and kill bacteria with the assistance of complement, and
neutralize drugs, toxins, and viruses.
3. Antibody
An antibody binds to a specific region on an antigen called an Epitope. A
single antigen can have multiple epitopes for different, specific antibodies.
4. Monoclonal Antibody
• Monoclonal antibodies are identical immunoglobulins, generated from a single B-cell clone.
• These antibodies recognize unique epitopes, or binding sites, on a single antigen.
• Derivation from a single B-cell clones and subsequent targeting of a single epitope is what
differentiates monoclonal antibodies from polyclonal antibodies.
• Polyclonal antibodies are antibodies that are derived from different cell lines. They differ in
amino acid sequences.
5. Characters of Monoclonal Antibodies
• Monoclonal antibodies (mAB) are single type of antibody that are identical and are
directed against a specific epitope (antigen, antigenic determinant)
• Produced by B-cell clones of a single parent or a single hybridoma cell line.
• A hybridoma cell line is formed by the fusion of one B-cell lymphocyte with a
myeloma cell.
• Some myeloma cell synthesize single mAB antibodies naturally.
6. Monoclonal Antibody VS Polyclonal Antibody
Parameters Monoclonal antibodies Polyclonal antibodies
Produced by: A single B cell clone Many B cell clone
Binds to: A single epitope of a cell Multiple epitopes of all
Antibody class All of a single Antibodies
class.
A mixture of different Antibodies
classes
Ag-binding sites Antibodies have the same
antigen binding sites
A mixture of Antibodies with
different Ag-binding sites classes
Potential for cross reactivity Low High
7. Advantages of using Monoclonal Antibodies
• Side effects can be treated and reduced by using mice-human hybrid cells or by using
fractions of antibodies.
• They bind to specific diseased or damaged cells needing treatment.
• They treat a wide range of conditions.
8. Disadvantages of Monoclonal Antibody
• Very expensive and needs considerable effort to produce them.
• Small peptide and fragment antigens may not be good antigens- monoclonal antibody
may not recognize the original antigen.
• Hybridoma culture may be subject to contamination.
• System is only well developed for limited animal and not for other animals.
• More than 99% of the cells do not survive during the fusion process - reducing the range
of useful antibodies that can be produced against an antigen
• It is possibility of generating immunogenicity.
• Time consuming project - anywhere between 6-9 months.
9. History of Monoclonal Antibody
• Paul Enrlich at the beginning of 20th century coined the term
"magic bullets" and postulated that, if a compound could be made
that selectively targets a disease-causing organism, then a toxin
for that organism could be delivered along with the agent of
selectivity.
• In the 1970s, the B-cell cancer multiple myeloma was known. It
was understood that these cancerous B-cells all produce a single
type of antibody (a paraprotein).
10. History of Monoclonal Antibody
• In 1975, Kohler and Milstein provided the most outstanding proof of the clonal selection
theory by fusion of normal and malignant cells (Hybridoma technology) for which they
received Nobel prize in 1984.
• In 1986, first monoclonal antibody was licensed by FDA. Orthoclone OKT3
(muromonab-CD3) which was approved for use in preventing kidney transplant
rejection.
11. Preparation of Monoclonal Antibody
• Monoclonal Antibody (mAB) is produced by cell lines or clones obtained from the
immunized animals with the substances to be studied. Cell lines are produced by fusing B
cells from the immunized animal with myeloma cells.
• To produce the desired mAB, the cells must be grown in either of two ways:
1. by injection into the peritoneal cavity of a suitably prepared mouse (In vivo method)
2. by In vitro tissue culture.
• The vitro tissue culture is the method used when the cells are placed in culture outside the
mouse the mouse's body in flask.
13. 1.Immunization of specific animal which generate B cells
Need to create antigen of interest against which want to
create a monoclonal antibody and use an organism such
as mouse or rabbit as vehicle to make monoclonal
antibodies
So immunizing these mouse
generate B cells against these
antigens in spleen
Antigen of interest is made by
DNA recombinant technology
and injected in mouse
intraperitoneally
14. 2. Extraction of B cells
After some days, B cell which are
produce in spleen against
Antigens are extracted
Then the extracted B cells are cultured
15. Need of Hybridoma technology
But there is a fundamental problem in terms of culturing
the B cells. B cells don’t survive in a culture more than a
week then we can initially get some amount of
monoclonal antibodies from these B cells but the problem
is we can’t get it for long.
SO WHAT’S THE SOLUTION?
In order to solve this problem
we use Hybridoma technology
16. 3. Fusion of B cells with Myeloma cells
In hybridoma technology , two types of cells are used Antigen specific B cells extracted from
the spleen from immunized organism and it would produce antibodies (mortal)
Now , fuse these Antibodies with Myeloma cells (Immortal) actually they are cancer cell line.
By fusion of above these cells they would generate Hybridoma cells (immortal and generate
antibodies as well)
17. 3. Fusion of B cells with Myeloma cells
Antigen specific B cells
Myeloma cells
18. 4. Different cell types in culture
When we trying to add B cells along with myeloma cells
to generate hybridoma cells there are a lot of cell types
which would be present in these culture and they are
following:
20. 5. Selection of HAT medium
Let us review that how we can get only hybridoma cells.
Selection procedure is required by suing HAT media which
contains (Hypoxanthine + Aminopterin + Thymidine).
21. Basic concept of cell division
When any cells divide it needs to replicate its DNA and
DNA replication requires a lot of nucleotides so
nucleotides biosynthesis is crucial for cells to divide and
inside the cells nucleotide are generated using two
pathways De novo pathway and Salvage pathway
22. Pathways of Nucleotide synthesis
In the absence of De novo pathway , Salvage pathway is only way of
generating these nucleotide. In salvage pathway, Activated ribose +
Base are used to produce nucleotide & It is done by hypoxanthine +
thymidine + HGPRT (Hypoxanthine-guanine phosphoribosyltransferase
) . HGPRT is present in B cells but absent in myeloma cells .
So , HGPRT + ve cells would survive because salvage pathway would
occur but HGPRT –ve cells would die due to absence of salvage
pathway and no way of generating nucleotide and cell unable to get
divide and would die.
Aminopterin present in the HAT media
inhibit Dihydrofolate reductase enzyme
which further terminate synthesis of
nucleotide by De novo pathway
23. 5. Isolation of Hybridoma Cells
The remaining two cells like Antigen
specific B cells and fused B cells would die
because they are immortal and after 2
weeks only Hybridoma cells are left.
Both myeloma cells and fused myeloma cells
are HGPRT –ve and they would die
eventually
24. 6.Screening of Hybridoma cells
The colonies of hybridoma cells are picked
up and each clones would be cultured in a
multiple plate.
25. 6.Screening of Hybridoma cells
From these multiple plate , the culture would be
screened for the specificity of antibodies with the
help of ELISA and some other techniques like
western blot or immunofluorescence
26. 7. Bulk production of Monoclonal Antibodies
When 2 or 3 tubes of specific monoclonal antibodies are
generated than these cells which are residing in these
tubes are cultured in bulk in a big flask, That’s how we
get a lot of hybridoma cells which yield monoclonal
Antibodies
31. 1.Biochemical Analysis
• Routinely used in radioimmunoassay (RIA) and enzyme-linked immunosorbent assays
(ELISA) in the laboratory.
• These assays measure the circulating concentrations of hormones (insulin, human
chorionic gonadotropin, growth hormone, progesterone, thyroxine, triiodothyronine,
thyroid stimulating hormone) and several other tissue and cell Products (blood group
antigens, blood clotting factors, interferon's, interleukins, tumor markers).
• E.g. Pregnancy by detecting the urinary levels of human chorionic gonadotropin.
• Hormonal disorders analysis of thyroxine, triiodothyronine.
• Cancers estimation of plasma carcinoembryonic antigen in colorectal cancer, and prostate
specific antigen for prostate cancer
History of History of
32. 2.Diagnostic imaging
• Radiolabeled-MAbs are used in the diagnostic imaging of diseases, and this technique is
referred to as immunoscintigraphy. The radioisotopes commonly used for labeling MAb
are iodine-131 and technetium-99. The MAb tagged with radioisotope are injected
intravenously into the patients.
• These MAbs localize at specific sites (say a tumor) which can be detected by imaging the
radioactivity. In recent years, single photon emission computed tomography (SPECT)
cameras are used to give a more sensitive three dimensional appearance of the spots
localized by radiolabeled-MAbs.
• Myocardial infarction, DVT, atherosclerosis etc.
33. 3.Direct use of Monoclonal Antibody as therapeutic agents
• In destroying disease-causing organisms: MAbs promote efficient
opsonization of pathogenic organisms (by coating with antibody) and
enhance phagocytosis.
• In the immunosuppression of organ transplantation: In the normal
medical practice, immunosuppressive drugs such as cyclosporin and
prednisone are administered to overcome the rejection of organ
transplantation. In recent years, MAbs specific to T-lymphocyte surface
antigens are being used for this purpose
•
34. 3.Direct use of Monoclonal Antibody as therapeutic agents
• In the treatment of cancer:
MAbs, against the antigens on the surface of cancer cells, are useful for the treatment of
cancer. The antibodies bind to the cancer cells and destroy them via different pathways.
• In the treatment of AIDS:
Genetic engineers have been successful to attach Fc portion of mouse monoclonal antibody
to human CD4 molecule. This complex has high affinity to bind to membrane glycoprotein
gp120 of virus infected cells. The Fc fragment induces cell-mediated destruction of HIV
infected cells.
35. 4.Monoclonal Antibody as Targeting agents
• The drugs can be coupled with MAb (directed against a cell surface antigen of the cells,
say a tumor) and specifically targeted to reach the site of action.
E.g. Alkaline phosphatase for the conversion of phosphate pro-drugs.
Carboxy peptidase for converting inactive carboxyl pro-drugs to active drugs.
Lactamase for hydrolyzing ẞ-lactam ring containing antibiotics.
• MAbs in the dissolution of blood clots:
Fibrin is the major constituent of blood clot which gets dissolved by plasmin. Plasmin in
turn is formed by the activation of plasminogen E.g. activator. By Tissue plasminogen
activator (tPA) can be used as a therapeutic agent to remove the blood clots.
36. 5.Drug delivery through liposomes coupled to tissue-specific MAbs
• Liposomes are sacs or vesicles formed spontaneously when certain lipid molecules are
exposed to aqueous environment.
• Drug entrapped in liposomes that are coated with MAbs directed against tissue-specific
antigens are being tried for drug delivery.
• Unfortunately, the progress in this approach has been limited, since such liposomes do
not reach the target cells.
• They are retained mostly in the liver and spleen (reticuloendothelial cells), and degraded.
37. 6.Protein purification
• Monoclonal antibodies can be produced for any protein. And the so produced MAb can
be conveniently used for the purification of the protein against which it was raised.
• MAbs columns can be prepared by coupling them to cyanogen bromide activated
Sepharose (chromatographic matrix). The immobilized MAbs in this manner are very
useful for the purification of proteins by immunoaffinity method.
• There are certain advantages of using MAbs for protein purification. These include the
specificity of the MAb to bind to the desired protein, very efficient elution from the
chromatographic column and high degree of purification.
39. References
• Shivanand P. (2010). "Hybridoma technology for production of monoclonal antibodies" International Journal of
Pharmaceutical Sciences Review and Research vol.1, issue 2 (017)
• U. Marx et al. (1997) "Monoclonal Antibody Production" The Report and Recommendations of ECVAM Workshop
ATLA 25, 121.137
• Edward A. Greenfield, (2014) "Antibodies: A Laboratory Manual" Cold Spring Harbor Laboratory Press, 2 ^ (nd) ed.
Chapter 7
• Justin K.H. Liu, (2014) "The history of monoclonal antibody development Progress, remaining challenges and future
innovations" Annals of Medicine and Surgery (2014) 113-116
• Andrew S., Otavia C ., (2014) "Monoclonal antibodies for the therapy of cancer" Simpson andCaballero BMC
Proceedings 2014, 8 (Suppl 4)
• WHO Guidelines on evaluation of monoclonal antibodies as similar biotherapeutic products,