Effect of BCR-ABL expression and chemical mutagenesis by ENU on kinase point mutation rates. (A) BA-transduced 32D cells were sorted by flow cytometry on the basis of GFP staining intensity into 32D-BA^low and 32D-BA^high cells (right). BCR-ABL protein expression was confirmed 72 hours after sorting by Western blotting using anti-Abl antibodies. The blot was reprobed with anti-actin antibodies as a loading control (left). (B) Cell-based BCR-ABL kinase mutation assay. 32D-BA^low and 32D-BA^high cells were exposed to ENU and 1 week later were exposed for 4 weeks to 2μM IM. The number of outgrowing resistant clones is given relative to the total input cell number per 96-well plate. Data represent means ± SEM of 3 independent experiments (***P < .001; *P < .05 according to 2-way ANOVA, with Bonferroni adjustment for multiple comparisons).