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. 2018 Jan 31;8:141–151. doi: 10.1016/j.omtm.2017.12.006

Figure 5.

Figure 5

Apoptotic Cell Death after Transfecting KB Cells with AA cmRNA

At 24 hr (A–C) or 48 hr (D and E) post-transfection of KB cells with AA (abrin-a, A-chain) or AAstop cmRNA, cells were examined for apoptosis. (A and B) Annexin V-AF488 and propidium iodide (PI) double staining flow cytometry assay. (A) Percentages of cells in early apoptotic stage (Annexin V-AF488 positive, PI negative). Data is shown as mean in % ± SEM and is compared to untransfected control cells (UT, dotted line). (B) Representative dot plots of cells transfected with 100 ng cmRNA. The percentages of the different populations are given in the respective corners. (C) Caspase-Glo 3/7 assay; activity was proportional to luminescence. Data is represented as mean in % ± SEM of untransfected control cells (UT, dotted line). (D and E) Hoechst33258 staining. (D) Representative phase contrast and fluorescence images of cells transfected with 100 ng cmRNA are displayed. Arrows depict cells showing “membrane blebbing” or nuclear fragmentation. (E) Number of Hoechst33258-positive cells. Data (mean ± SEM) is presented as multiple of untransfected control cells (UT, dotted line). Statistical significance versus AAstop cmRNA was assessed by two-way ANOVA adjusted for multiple comparisons, with **p < 0.01, ****p < 0.0001, and n = 3.