Mutation myelomas

B lymphocytes will be eliminated during continuous culture because these cells have a short life span in culture. Commercially available myeloma cells for hybridoma production have mutations in one of the enzymes of the salvage pathway of purine nucleotide biosynthesis. Hybridoma cells are cultured in medium that forces the cells to utilize the salvage pathway for nucleotide synthesis. The mutated myeloma cells or hybridization products of two myeloma cells will die in this selection medium since they are incapable of nucleotide synthesis under these propagation conditions. However, myeloma cells that have fused to the B lymphocytes derived from the spleen of the immunized animal will have an intact salvage pathway and will survive in the selection medium. Thus, only the B lymphocytes-myeloma hybridomas will survive prolonged culture in the selection medium. 

Billadeau, D., Liu, P., Jelinek, D., Shah, N., LeBien, T. W. and van Ness, B. (1997). Activating mutations in the N- and K-ras oncogenes differentially affect the growth properties of the IL-6-dependent myeloma cell line ANBL6. Cancer Res. 57, 2268-2275. 

At this point, several questions concerning somatic mutation remained Was mutation peculiar to myelomas or did it occur in normal B cells Were VH genes diversified by nucleotide replacement What fraction of the antibody diversity expressed by an animal is the result of somatic mutation and how much of this diversity is functional The pursuit of these questions awaited the development of hybridoma technology by Kohler and Milstein [84] and, thus, the ability to immortalize B cells that are participating in an immune response in vivo. 

Smith et al. (8) believe that patterns of amino acid sequences in L chains of human myeloma proteins are difficult to reconcile with somatic mutation of a small number of germ line genes. Consider myeloma proteins from different individuals with L chains of the V, subgroup. The genealogy of amino acid sequences indicate that all are derived from some common ancestral gene. The existing sequences indicate the presence of mutations (from an ancestral gene) which are common to some sequences but not to others. To illustrate this point ... 

The observation that identical myeloma proteins are very rare in humans was ascribed to the fact that, typically, a human is about 50 years old, whereas a mouse is less than 1 year old when afflicted with multiple myeloma (11). During the 50-year period a large number of structural variants might arise as a result of sequential mutations. 

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