Assignment lines. In this time used the hypoxanthine-aminoptenin-thymidine

“Production Of monoclonal Antibodies by Hybridoma technology”

Submitted to:
Najneen Ahmed
Senior lecturer
Department of Pharmacy
East West University
Submitted by:
Majedul HaqueID: 2014-3-70-20
Department of Pharmacy
East West University
In 1964 Littlefield developed a way. In this way to isolate a hybrid cells from to parents cell lines. In this time used the hypoxanthine-aminoptenin-thymidine (HAT) selection medium. On August 7, 1975 Cesar Milstein and Georges Kohler published their paper on the production of monoclonal antibodies. In 1990 Milstein 1st produced monoclonal antibodies. 1
Monoclonal Antibodies
Homogeneous immunoglobulins are also called monoclonal antibodies (mAb & moAb). It’s originated from a individual B cell clone. Unique epitopes, or binding sites, on a single antigen that is recognize from these antibodies. 3
Four types of monoclonal antibodies are
Murine (-omab)
Chimeric (-ximab)
Humanized (-zumab)
Human (-umab)
Hybridoma Technology
Cesar Milstein and Georges Kohler on August 7, 1975 published their paper on the production of monoclonal antibodies that is continuous cell culture. Production of monoclonal antibodies (MAbs) by hybridoma cell lines this technology called Hybridoma technology. This technology is used the purification of protein and the diagnosis treatment of disease. 1
Advantage of hybridoma technology 2
Reduced the variability of culture medium
Decreased risk of infectious agents.

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Potential for raise antibody secretion.

Low or independence on animals.

Production Of monoclonal Antibodies by Hybridoma technology

Adapted from: Anon, (2018). online Available at: Accessed 6 Nov. 2018.

A hybridoma, also known as a harry cell can be produced by the injection of a specific antigen into a mouse, procuring the antigen specific plasma cells from the mouse spleen and also the subsequent fusion of the cell with a cancerous immune cell. That is also known as myeloma cell. The produced hybrid cell can be cloned so that it can produce many identical daughter clones and those daughter clones then secrete the immune cell product. As these antibodies come from only one type of cell ,they are known as monoclonal antibodies. With the help of this process, it is possible to combine the qualities of the two different types of cells, increase the ability to grow continually and produce larger amount of pure antibody.
Preparation of monoclonal antibodies is done with the help of HAT medium (hypoxanthine aminopetrin thymidine). At first, laboratory animals are exposed to an antigen to which we are interested in isolating an antibody against. The B cells are fused with immortalized myeloma cells which lack HGPRT gene using polyethylene glycol as the splenocytes are isolated from the mammal. Then the fused cells are incubated in the HAT medium. Aminopterin that is present in the myeloma cell die because they cannot produce nucleotide by the slavage medium that blocks the pathway that allows for nucleotide synthesis. As a result, infused D and infused B cell die as they have short life span. Because of the HGPRT gene coming from the B cell is functional, only the B-cell myeloma hybrids survive. These cells help to produce antibodies and they are immortal. Since the incubated medium is diluted into multiwall plates, each well contains only one cell. Then the supernatant in each well can be checked for the desired antibody. As the antibodies in a well are produced from the same B cell, they will be directed to the same epitope known as monoclonal antibodies. Once a hybrid colony is produced, it will keep growing in the culture medium like RPMI-1640 (with foetal bovine serum and antibiotics) and produce antibody. Then the next stage known as rapid primary screening process, which helps to identify and select only the hybridomas. The hybridoma culture supernatant, secondary enzyme labeled conjugate, and chromogenic substrate will be incubated, and the formation of a colored product will indicate a positive hybridoma. 1
Monoclonal antibodies an essential tool for biomedical research, medical value and are of great commercial value. Production of monoclonal antibody by hybridoma technology is most valuable in vitro conditions. The route of administration of monoclonal antibodies is intravenous.1
Reference (2018). Monoclonal antibody production. online Available at: Accessed 7 Nov. 2018.

GUPTA, P. (2018). Elements of biotechnology. 3rd ed. pp.203-211. (2018). What are Monoclonal antibodies(MABs) – GenScript. online Available at: Accessed 6 Nov. 2018.


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