Cultures derived from irradiated human mammary epithelial cells show more SA-βgal (-) and p16 (-) cells than those derived from unirradiated human mammary epithelial cells. Representative fields are shown stained for (A) SA-βgal activity (blue reaction product) or (C) total p16 protein (green immunofluorescence). In (C), cell nuclei were counterstained with DAPI (blue). Inset = minus 1°Ab control. Note the greater presence of SA-βgal (-) and p16 (-) cells in the irradiated cultures. In (B) and (D), cytometry was used to quantitatively compare the percentages of positively and negatively staining cells in the corresponding cultures. In (B), flow cytometry results of an experiment performed using B1450 cultures are summarized in a mean ± SE scatter plot (P < 0.001, non-parametric Wilcoxon test). In (D), the percentages of p16 (-) cells in unirradiated and irradiated B1400 and B1450 cultures were determined using a Cellomics Array Scan VTI automated fluorescence microscopic imaging system. For each condition, staining was measured for 460 fields per well and summarized in a mean ± SE scatter plot (P < 0.0001, non-parametric Wilcoxon test).