Production of Monoclonal Antibodies


Monoclonal antibodies are the secreted products of a hybridoma cell line that is created from the fusion of an antibody‐producing lymphocyte and a histocompatible myeloma cell. The hybrid cell inherits the ability to survive in tissue culture from the parent myeloma cell and the capacity to produce antibody from the parent lymphocyte. Each hybrid cell can develop into a clone of identical cells producing identical antibodies that bind to one epitope. Production of such monoclonal cell lines from the fusion mixture requires several rounds of dilution and selection on the basis of specific antibody secretion. Once established, the pure cell line can be stored frozen and recovered to produce unlimited quantities of identical antibodies as required. The vast majority of monoclonal antibodies are derived from mouse cells, although it is also possible to produce them from rat cells. There are a few human monoclonal antibodies derived from hybridomas, but these tend to be very difficult to establish and maintain. All the steps required to make mouse monoclonal antibodies are described here.

Keywords: monoclonal antibodies; hybridoma; cell fusion; cell cloning; myeloma

Figure 1.

Summary of procedures involved in monoclonal antibody production.

Figure 2.

Mouse spleen prior to preparation of single cell suspension.

Figure 3.

Mouse myeloma cells.

Figure 4.

Neubauer cell‐counting chamber grid. The cells in each of the large numbered squares are counted. The average value is the number of cells in 10−4 mL.

Figure 5.

Fusion mixture after 3 days growth in HAT medium. Note that very few cells are visible and there is much debris.

Figure 6.

Cell mixture 7 days after fusion. Clones develop rapidly under the debris.

Figure 7.

A monoclone. The small cells in the background are thymocytes.



Campbell AM (1991) Monoclonal Antibody and Immunosensor Technology Amsterdam: Elsevier Science.

Goding JW (1996) Monoclonal Antibodies: Principles and Practice3rd edn. San Diego: Academic Press.

Liddell JE (2001) Antibodies. In: Wild D (ed.) The Immunoassay Handbook 2nd edn. London: Nature Publishing Group.

Liddell JE and Cryer A (1991) A Practical Guide to Monoclonal Antibodies Chichester: Wiley.

Further Reading

Köhler G and Milstein C (1975) Continuous cultures of fused cells secreting antibody of predefined specificity. Nature 256: 495–497.

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Liddell, JE(May 2005) Production of Monoclonal Antibodies. In: eLS. John Wiley & Sons Ltd, Chichester. [doi: 10.1038/npg.els.0003773]