Flow Cytometry Detection of Anthracycline-Treated Breast Cancer Cells: An Optimized Protocol

Figures

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  Figure 1. Analysis of doxorubicin fluorescence emission via conventional flow cytometry (BD FACSVerse). Cells treated with 2.5 µM doxorubicin (Doxo) for 24 and 48 h were acquired using flow cytometry and shown as blue histograms on every channel of a conventional instrument (FACSVerse, BD Biosciences), equipped with three lasers (488 nm, 633 nm, and 405 nm). Overlayed red histograms show the profiles of matched untreated samples. Histograms are representative of three independent experiments.

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  Figure 2. Analysis of epirubicin fluorescence emission via conventional flow cytometry (BD FACSVerse). MDA-MB-231 cells treated for 24 and 48 h with 1 µM (A) or 2.5 µM (B) epirubicin were acquired using flow cytometry and shown as blue histograms on every channel of a conventional instrument (FACSVerse, BD Biosciences), equipped with three lasers (488 nm, 633 nm, and 405 nm). Overlayed red histograms show the related untreated samples. Histograms are representative of three independent experiments.

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  Figure 3. Analysis of doxorubicin fluorescence emission with ImageStream. MDA-MB-231 cells treated for 24 h with 1 µM doxorubicin were acquired using ImageStream: (A) representative images of treated cell brightfield and fluorescence detected in channel 1 (FITC and analogue fluorochromes) and channel 4 (propidium iodide and analogue fluorochromes) for representative cells treated with doxorubicin (Doxo) or untreated are shown; (B) histograms represent doxorubicin autofluorescence detected in channel 1 (FITC and analogue fluorochromes) and channel 4 (propidium iodide and analogue fluorochromes). Data are representative of three independent experiments.

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  Figure 4. Apoptosis analysis using flow cytometry after staining with Annexin V-BV450. Histograms show the percentage of apoptotic MDA-MB-231 cells after the treatment with doxorubicin at 2.5 μM for 24 h or 48 h. Data are presented as the means ± SD of triplicate experiments. ** p < 0.01 vs. control (ns, not significant).