Mouse monoclonal antibodies are a valuable tool in the biomedical field. They can be used for a variety of applications, from research to clinical diagnostics. In this blog post, we will discuss the basics of mouse monoclonal antibody production, including its protocol and application. The major difference between monoclonal antibodies and polyclonal antibodies is that monoclonal antibodies can have a single-specificity, meaning they only bind to the same epitope while polyclonal antibodies can bind to multiple epitopes and are generally synthesized by several different plasma cell lineages. Bispecific monoclonal antibodies can also be developed by wedging two epitopes from one monoclonal antibody into the binding site of another.
Steps to produce Mouse monoclonal antibody - Hybridoma Technology
- The main protocol of monoclonal antibody production is that a single antibody-producing cell produces only one type of antibody with a specificity. Because immunization causes a polyclonal response, with the production of many antibodies with various sensitivities and isotypes, it must be elicited in order to obtain a monoclonal antibody. However, because these cells have a limited lifespan, they must be bred with a non-producing myeloma cell line to create a hybrid cell that is both immortal and antibody producing. Cancereous cells being immortal in nature fusing them with normal cell is taken into consideration and this technology is called as hybridoma technology.
- Mouse is first immunized with the antigen of interest with the antigen of interest, then harvesting its spleen after determination of successful polyclonal antibody production. these spleen cells are the fused with myeloma cell line using polyethylene glycol. The hybrid cells are then selected and allowed to grow. Cells producing desired antibodies are the selected but the success rate is low, so a selective medium in which only fused cells can grow is used.
- Because myeloma cells no longer have the capacity to make hypoxanthine-guanine-phosphoribosyl transferase (HGPRT), an enzyme required for nucleic acid salvage synthesis, this is conceivable. If a defect in either the pyrimidine synthesis pathway or HGPRT is present, however, these cells are unable to grow. Cells that are exposed to aminopterin, a folic acid analog, which inhibits dihydrofolate reductase, or DHFR become unable to use the de novo pathway and become completely auxotrophic for nucleic acids, necessitating supplementation.
- The vitamin-depleted culture medium is known as HAT medium because it contains hypoxanthine, aminopterin, and thymidine. This media is ideal for the growth of fusion hybridoma cells. Unfused myeloma cells can't grow because they lack the HGPRT gene, so they are unable to replicate their DNA. Because of their short life span, unfused spleen cells cannot grow indefinitely. Because the spleen cell provides HGPRT and the myeloma companion has characteristics that make it immortal, hybrid cells known as fusion hybrids are capable of growing indefinitely in culture.
Monoclonal Antibody Application
- Monoclonal antibodies are being investigated as potential treatments for asthma, autoimmune disorders, cancer, poisoning, septicemia, substance abuse, viral infections, and a variety of other diseases.
- The protocol for producing mouse monoclonal antibodies is relatively simple and straightforward. Hybridoma technology is used to produce monoclonal mouse antibodies. This involves fusing a myeloma cell line with a lymphocyte that generates the desired antibody. Hybrid cells that have been selected and cultivated in culture are then utilized to manufacture large amounts of the wanted antibody.
- Monoclonal antibodies are being used for a variety of purposes, including research. They may be utilized to study disease processes or discover new therapeutic targets. Mouse monoclonal antibodies can also be utilized in clinical testing, such as cancer detection or disease monitoring.
- Clinical uses for mouse monoclonal antibodies include cancer detection and disease monitoring. Monoclonal antibodies are more costly to produce than small molecules due to the complicated procedures and overall size of the molecules, in addition to the significant research and development expenditures.
- Monoclonal antibodies that bind only to cancer cell-specific antigens and stimulate the immune system against the target cancer cell are one type of treatment for cancer. The ability to design and make monoclonal antibodies that can bind with their Fab regions to target antigen and a conjugate or effector cell" may be utilized as part of the drug development process. Such monoclonal antibodies can be modified for delivery of a toxin, radioisotope, cytokine, or other active conjugate, as well as designing bispecific antibodies that bind with their Fab regions not just to the target antigen but also to a conjugate or effector cell. With its Fc region, every intact antibody can bind to cell receptors or other proteins.
- Example- muromonab-CD3 (Orthoclone OKT3), which was developed from the mouse antibody muromonab-CD3. There's always a danger with this approach that some people exposed to mouse antibodies will develop an immune response to the mouse antibody sequence.