All of the following statements about hybridomatechnology are true except aEUR'
Myeloma cells with mutation in salvage pathway grows well in HAT medium Hvbridomas Hybridomas are cells that have been engineered to produce a specific antibody in huge numbers To achieve this, qualities, of two types of cells have to be combined together i.e. i) Cells which can produce large amount of pure antibody. and ii) Cells which have the ability to grow continually. These two types of cells are then fused together to form hybridoma Procedure The pure antibody secreting cells are produced by injecting specific antigen in a mouse and obtaining the antigen specific plasma cells (antibody producing cell) from the mouse's spleen. Cell which can grow indefinitely in culture are myeloma cells (cancerous cells). These two cell lines are fused together. The hybrid cell which is thus produced can be cloned to produce large number of identical daughter clones. These daughter clone cells then produce antibodies. Since these antibodies come from only one type of cell (hybridoma cell) they are called monoclonal antibodies. HAT (hypoxanthine, Aminopterin and thymidine) medium is used for preparation of monoclonal antibodies because it allows only fused hybridonza cells to grow. It does not allow the unfused myeloma and !infused antibody cells to grow. So HAT medium is a selective medium which allows (which allow selective growth of fused hybridoma) How does this happen ? Before we move on with the discussion, remember these few points about purine synthesis. Purine synthesis is essential for the survival of cells. Purine can be synthesized in two way i.e. - De novo synthesis (dihydrofolate reductase enzyme is required for this pathway)- - Salvage pathway (an enzyme hypoxanthine - guanine phosphoribosyl transferase is required for salvage pathways) Myeloma cells lack HGPase enzyme therefore they cannot synthesize purine by salvage pathways. Antibody cells have HGPase enzyme so they can use the salvage pathways. Aminopterin inhibits dihydrofolate reductase an enzyme used in denovo synthesis of purine When two cell lines i.e. antibody producing cell and myeloma cells are grown in HAT medium only the fused hybridoma cells survive. -Myeloma cells die because they lack the enzyme HGPase so they cannot use the salvage pathways for purine synthesis. They also cannot use de-novo pathway, because Aminopterin present in the HAT medium inhibits dihydrofolate reductase (an enzyme essential ,for denovo synthesis of purine). The unfused antibody producing cells die as they cannot grow indefinitely because of their limited life span. Only fused hybridoma cells grow indefinitely because the antibody cell paner supplies HGPase and the myeloma paner gives it immoality. (as it is a cancer cell)
Specific antibody producing cells are integrated with myeloma cells
Myeloma cells with mutation salvage pathway grows well in HAT medium.
Aminopterin, a folate antagonist, inhibits denovo pathway
HGP ase and thymidylate synthetase are required for salvage pathway