A NEW METHOD FOR ESTIMATING HIGH MUTATION-RATES IN CULTURED-CELLS

Fluctuation analysis allows for the determination of mutation rates in cell cultures in vitro. As originally described by Luria and Delbruck and extended by Lea and Coulson and by Capizzi and Jameson, this anal ysis has been useful in estimating mutation rates in cultured cells wh ere the frequency of mutational events is low. However, in cultures wh ere high mutation rates and multiple independent mutation events occur , leading to the accumulation of many mutant cells, these standard met hods may not apply. Here, we present a new method for the estimation o f mutation rates based on the assumption that multiple events may cont ribute to the accumulation of mutant cells. We compared mutation rates determined by Lea and Coulson's and by Capizzi and Jameson's methods with those determined by our method using experimental and simulated d ata from our studies of immunoglobulin gene mutation and isotype switc hing in B lymphocyte cultures. The three methods resulted in very diff erent calculated rates when many mutants were present in the culture, such as when mutation rates were high, while only small differences in calculated rates were found when mutants were ran. Unlike previous fl uctuation analysis calculations, our method is applicable for the esti mation of;both low and high rates.