Developing Monoclonal Antibodies

How are Monoclonal Antibodies Generated

To facilitate the generation of monoclonal antibodies, various techniques have been developed, including hybridoma technology and phage display.

Monoclonal antibodies are produced by introducing an antigen to a mouse and then fusing polyclonal B cells from the mouse's spleen to myeloma cells. In the subsequent identification step, the culture supernatants of all hybridoma cells are screened weekly for the production of the antibody of interest. Hybridoma cells producing the antibody of interest are cloned by limited dilution till a monoclonal hybridoma is found. This is called hybridoma technology which is a very time-consuming and laborious process.

Antibody phage display is the most widely used method for in vitro selection of monoclonal antibodies. The power of phage display technology results from its ability to establish a physical connection between phenotype (displayed peptide) and genotype (DNA sequence encoding the displayed peptide). This phenotype-genotype link forms the cornerstone of phage display methodology and makes it possible for researchers to isolate target-avid ligands displayed on the phage.

The Fig 1. shows the timeline of monoclonal antibody development since 1975. In 1986, the first therapeutic monoclonal antibody Orthoclone OKT3 (also called muromonab) was approved by FDA. More and more monoclonal antibodies (mAbs) are undergoing clinical trials.

Timeline of major improvements in monoclonal antibody development
Fig 1. Timeline of major improvements in monoclonal antibody development

Monoclonal Antibody Development Methods

For hybridoma development, there are four steps to produce monoclonal antibodies, including 1) animal immunization, 2) fusion & selection, 3) antibody purification and 4) Quality control. View more about "Hybridoma antibody production".

For phage display antibody development, it involves isolating B-lymphocytes from the blood of humans and then isolating the mRNA and converting it into cDNA using PCR to amplify all the VH and VL segments. These segments can then be cloned into a vector (usually as scFv) next to the PIII protein of a bacteriophage before being used to infect E. coli to generate a library containing approximately 1010 cells by inoculating the library with an additional helper phage.

E. coli can then secrete the bacteriophage containing the VH and VL segments as part of the bacteriophage coat. Specific VH and VL segments against the antigen can then be selected and used to reinoculate E. coli with the bacteriophage. Cells containing the plasmid can then be isolated and sequenced. Its advantages include: once the library is made, the same library can be used to generate new antibodies and does not have to be remade, no immunisations are required as the entire process is done in vitro, antibodies can be obtained much more quickly than the traditional hybridoma technique and the library can be used to generate antibodies to toxic antigens that could not be used to immunise an animal.

Sino Biological is capable of developing and manufacturing monoclonal antibodies by hybridoma and phage display, offering researchers worldwide good quality monoclonal antibody services at a very affordable price to support your project.

Monoclonal Antibody Generation

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Applications of Monoclonal Antibodies

Today monoclonal antibodies are used in a variety of applications as cancer treatment, medical diagnosis and basic research.

How are monoclonal antibodies used in cancer treatment

Monoclonal antibodies used in cancer treatment are designed to bind to antigens that are generally more numerous on the surface of cancer cells than healthy cells. By targeting those antigens, the antibodies can latch onto the cancer cells and act as a "call to arms" for other disease-fighting warriors in the immune system.

More than a dozen monoclonal antibodies have been approved by the Food and Drug Administration (FDA) to fight different types of cancer, including breast, head and neck, lung, liver, bladder, and melanoma skin cancers, as well as Hodgkin lymphoma. First tried in advanced melanoma, monoclonal antibodies have extended some patients' survival as much as 10 years.

Related sections: Immunotherapy | Cancer Drug Target

How are monoclonal antibodies used in medical diagnosis

Monoclonal antibodies have revolutionized the laboratory diagnosis of various diseases. They are employed as diagnostic reagents for biochemical analysis or as tools for diagnostic imaging of diseases. Diagnostic tests based on the use of mAbs as reagents are routinely used in radioimmunoassay (RIA) and enzyme-linked immunosorbent assays (ELISA) in the laboratory.

Owing to mAbs, it is now possible to do the early diagnosis of the following conditions/diseases, including pregnancy, hormonal disorders, infectious diseases, and cancers.

How are monoclonal antibodies used in research

In basic research, researchers can use monoclonal antibodies to identify molecules of interest by assays such as Western Blot, flow cytometry, immunohistochemistry (IHC), enzyme-linked immunosorbent assay (ELISA), and many others.

References

1. Hanack, K., Messerschmidt, K., & Listek, M. (2016). Antibodies and selection of monoclonal antibodies. In Protein Targeting Compounds (pp. 11-22). Springer, Cham.
2. Scott, A. M., Allison, J. P., & Wolchok, J. D. (2012). Monoclonal antibodies in cancer therapy. Cancer Immunity Archive, 12(1), 14.
3. Aghebati-Maleki, L., Bakhshinejad, B., Baradaran, B., Motallebnezhad, M., Aghebati-Maleki, A., Nickho, H., ... & Majidi, J. (2016). Phage display as a promising approach for vaccine development. Journal of biomedical science, 23(1), 66.
4. Abell, C. W., & Denney, R. M. (1985). Application of monoclonal antibodies in basic research, diagnosis, and therapy. Journal of natural products, 48(2), 193-202.

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