Figure 4

Flow cytometry detection of P-gp transfers in tagged parental MCF-7. Parental sensitive MCF-7 were tagged with the persistent fluorescent probe CellTracker Blue (ctbMCF-7), prior to co-cultures. ctbMCF-7 alone, extemporaneous mixtures of 50:50 ctbMCF-7:MCF-7/Doxo or co-cultures at various times were analyzed after labelling with the PE-conjugated UIC2 monoclonal antibody. Scatter plots were obtained by quantifying CellTacker Blue content the FL1 channel and P-gp immunodetection in the FL2 channel. A, pure ctbMCF-7. B, pure MCF-7/Doxo. C, extemporaneous mixture of ctbMCF-7:MCF-7/Doxo. D-F, co-cultures obtained from 50:50 ctbMCF-7:MCF-7/Doxo, after 1 to 3 days. All analysis were performed with unchanged excitation light power and photomultipliers voltage settings. The quadrant limits were set in order to obtain less than 1% of double positive cells (Q2) in the analysis of the sample C. Despite a progressive loss of FL1 signal, due to Celltracker blue dilution within daughter ctbMCF-7, a population of dually labelled cells appeared in the upper right quadrant. Percentages indicate the fraction of cells having a double positive labelling in quadrant Q2.