CLONAL VARIANTS OF HYBRIDOMA CELLS THAT SWITCH ISOTYPE AT A HIGH-FREQUENCY

As B cells differentiate under the influence of antigen and T cells, t hey frequently switch from the expression of IgM antibody to the expre ssion of other isotypes. This is accomplished by rearranging the expre ssed variable region gene to downstream constant region genes and dele ting the intervening sequences. Some B-cell lines that represent early stages in development switch constitutively in culture at frequencies that approach those of lipopolysaccharide- or lymphokine-stimulated n ormal B cells. Hybridoma cells represent a later stage of development and rarely switch in culture. In contrast to early B-cell lines, hybri domas produce large amounts of immunoglobulin, and single cells can be assayed easily for the expression of new isotypes. We have used the E LISA spot assay and fluctuation analysis to determine the rate of swit ching of two hybridoma cell lines. By identifying subclones that switc hed more frequently, we have progressively enriched for cells that swi tch spontaneously at higher rates. These cells, like normal cells, swi tch by rearrangement and deletion, and the frequency of switched cells in some of the clones is comparable to that which has been observed i n less differentiated B-cell lines and in normal B cells.