Hybridoma Preparation
Hybridoma Preparation - Related Videos
04:51
A Hybridoma Technology for Producing Monoclonal Antibodies against a Metallopeptidase
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2025
This video demonstrates an assay to produce monoclonal antibodies against porcine aminopeptidase N (APN), a membrane-bound metallopeptidase, using a traditional hybridoma technology. Upon immunizing a mouse with APN, the spleen cells are harvested and fused with immortal myeloma cells to produce hybridomas. The hybridomas are then selected and proliferated using the HAT (hypoxanthine-aminopterin-thymidine) medium to produce monoclonal antibodies specific to different APN epitopes.
13:21
Antibody Generation: Producing Monoclonal Antibodies Using Hybridomas
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2023
Source: Frances V. Sjaastad1,2, Whitney Swanson2,3, and Thomas S. Griffith1,2,3,4 1 Microbiology, Immunology, and Cancer Biology Graduate Program, University of Minnesota, Minneapolis, MN 55455 2 Center for Immunology, University of Minnesota, Minneapolis, MN 55455 3 Department of Urology, University of Minnesota, Minneapolis, MN 55455 4 Masonic Cancer Center, University of Minnesota, Minneapolis, MN 55455 Polyclonal antibodies are defined as a collection of antibodies directed against...
01:31
Hybridoma Technology
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2023
Hybridoma technology is used for the large-scale production of monoclonal antibodies. Monoclonal antibodies bind to only a single antigenic determinant or epitope. Such antibodies are used in research, diagnostics, and disease therapy. The hybridoma technology established in 1975 by Georges Köhler and Cesar Milstein was awarded the Nobel Prize in Medicine in 1984 for revolutionizing research and therapy. Hybridoma Selection Commonly used fusion techniques — electroporation, polyethylene glycol...
09:42
Generation of Murine Monoclonal Antibodies by Hybridoma Technology
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Cited by 68 •
2017
An optimized protocol is presented for the generation of monoclonal antibodies based on the hybridoma technology. Mice were immunized with an immunoconjugate. Spleen cells were fused by PEG and an electric impulse with immortal myeloma cells. Antibody-producing hybridoma cells were selected by HAT and antigen-specific ELISA screening.
03:00
Preparation of Epoxides
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2023
Overview Epoxides result from alkene oxidation, which can be achieved by a) air, b) peroxy acids, c) hypochlorous acids, and d) halohydrin cyclization. Epoxidation with Peroxy Acids Epoxidation of alkenes via oxidation with peroxy acids involves the conversion of a carbon–carbon double bond to an epoxide using the oxidizing agent meta-chloroperoxybenzoic acid, commonly known as MCPBA. Since the O–O bond of peroxy acids is very weak, the addition of electrophilic oxygen of peroxy acids to...
FAQs
The fusion combines two complementary properties: B cells provide the capacity to produce antibodies against the target antigen, while myeloma cells contribute immortality. The resulting hybrid cells can therefore continue growing while producing antibody. This pairing is central to obtaining a stable, renewable source rather than relying on short-lived antibody-producing cells.
Polyethylene glycol promotes fusion between antibody-producing spleen cells and myeloma cells. Its role is to bring the two cell types together so hybrid cells can form, creating the cellular basis for subsequent selection. Fusion efficiency influences how many candidate hybridomas become available for screening and eventual isolation.
HAT medium provides a selection step that enriches for hybrid cells after fusion. Because the fusion mixture contains more than the desired hybrids, selection narrows the population before antibody testing. Researchers then screen the surviving cultures for production of an antibody with the required specificity, separating selection from functional identification.
Screening determines whether a surviving culture produces an antibody that recognizes the desired target, while cloning helps establish a line derived from a single selected hybrid cell. Using both steps improves confidence that the resulting hybridoma has the intended specificity and can provide a consistent antibody source for later experiments.
The workflow begins with immunization using the target antigen, followed by isolation of antibody-producing B cells from the spleen. These cells are fused with myeloma cells using polyethylene glycol, and the fusion products are placed in HAT medium. Surviving cultures are screened for desired specificity and cloned to identify suitable lines.
Selected hybridomas provide consistent antibodies for pathogen detection, antigen characterization, and diagnostic assay development. They also support studies of immune recognition by allowing investigators to examine interactions with a defined target. Because the cell line is renewable, experiments can use antibody produced from the same selected source over time.