Heterogeneity of the IFN response at the single cell level.

(A) A549-IFIT1deGFP cells stimulated with increasing amounts of IFN-α for 24 h were analyzed for IFIT1deGFP expression by flow cytometry. (B) A549-IFIT1deGFP cells stimulated with 10 or 100 IU/ml IFN-α for 24 h were subjected to FACS. IFIT1deGFP-positive and -negative cells were separated, seeded and re-stimulated 24 h later using IFN-α concentrations specified in the right. Twenty four hours later, cells were analyzed by flow cytometry. (C) Heterogeneity of IFN-α response kinetics at the single cell level. A549-IFIT1deGFP cells were treated with 10 or 100 IU/ml IFN-α (left and right panel, respectively) and single cells were tracked by time-lapse microscopy for a total observation period of 72 h. For each time point Z-stacks composed of 16 to 20 consecutive optical sections were acquired and total intensity of IFIT1deGFP in individual cells was extracted. Mean intensities of the IFIT1deGFP reporter per cell are plotted for selected cells (indicated by different colors). A more comprehensive data set is shown in S4 Fig. (D) Kinetics of expression onset of IFIT1deGFP in cells treated with 10 or 100 IU/ml IFN-α (left and right panel, respectively). Data are based on live cell imaging results displayed in panel C and in S4 Fig.