Derek Wong

Monoclonal Lecture

Monoclonal Antibodies

            Generally antibodies generated in a natural immune response are the result of a mixture of different specificities and affinities.  To determine which exact antibody did what, a series of test was used, however there were problems.  First, each antiserum is different from all other antisera. The antisera used can only be produced in limited quantities, making it impossible to use in a series of tests.  Thirdly, even if antisera was somehow purified, there would still be minor populations of antibodies that may give unexpected results (ex. cross-reactivity).  Therefore, the only way seen was to make an unlimited supply of homogenous antibody molecules with a known specificity.  This is done through monoclonal engineering.

            A monoclonal antibody is made in mice (or in any other organism in principle) by immunizing it with Ag A so that the spleen cells produce the specific antibody.  To produce the unlimited supply these spleen cells are fused with immortal myeloma cells using polyethleneglycol (PEG) to produce a hybrid cell line called a hybridoma.  The myeloma cells are selected to ensure that it is not secreting any antibodies and that they are sensitive to the HAT medium (hypoxanthine-aminopterim-thymidine) because they lack the HGPRT enzyme (hypoxanthine:guanine phophoribosyl transferase).  Once transferred to the HAT medium, the immortal hybridomas proliferate while the mortal spleen cells and unfused HGRPT- myeloma die.  Individual hybridomas are then screened for antibody production by ELISA (enzyme-linked-immunosorbent-assay), and cells which make the desired specificity are cloned from the single antibody-producing cell.

            The main advantage to monoclonal antibodies is that it provides a limitless supply of antibodies of a single and known specificity.  This can then be used in serological assays such as diagnostic probes and as therapeutic agents.  The disadvantage is that it is very difficult to make, so far only mouse monoclonals are routinely produced.