A monoclonal antibody is an antibody made by cloning a unique white blood cell. All subsequent antibodies derived this way trace back to a unique parent cell.
Monoclonal antibodies can have monovalent affinity, binding only to the same epitope (the part of an antigen that is recognized by the antibody). In contrast, polyclonal antibodies bind to multiple epitopes and are usually made by several different antibody secreting plasma cell lineages. Bispecific monoclonal antibodies can also be engineered, by increasing the therapeutic targets of one monoclonal antibody to two epitopes.
Much of the work behind production of monoclonal antibodies is rooted in the production of hybridomas, which involves identifying antigen-specific plasma/plasmablast cells (ASPC) that produce antibodies specific to an antigen of interest and fusing these cells with myeloma cells.[6] Rabbit B-cells can be used to form a rabbit hybridoma. Polyethylene glycol is used to fuse adjacent plasma membranes,[9] but the success rate is low, so a selective medium in which only fused cells can grow is used. This is possible because myeloma cells have lost the ability to synthesize hypoxanthine-guanine-phosphoribosyl transferase (HGPRT), an enzyme necessary for the salvage synthesis of nucleic acids. The absence of HGPRT is not a problem for these cells unless the de novo purine synthesis pathway is also disrupted. Exposing cells to aminopterin (a folic acid analogue, which inhibits dihydrofolate reductase, DHFR), makes them unable to use the de novo pathway and become fully auxotrophic for nucleic acids, thus requiring supplementation to survive.
The selective culture medium is called HAT medium because it contains hypoxanthine, aminopterin and thymidine. This medium is selective for fused (hybridoma) cells. Unfused myeloma cells cannot grow because they lack HGPRT and thus cannot replicate their DNA. Unfused spleen cells cannot grow indefinitely because of their limited life span. Only fused hybrid cells referred to as hybridomas, are able to grow indefinitely in the medium because the spleen cell partner supplies HGPRT and the myeloma partner has traits that make it immortal (similar to a cancer cell).
