Uniform downregulation of Oct4 protein levels during EB differentiation.
Undifferentiated ESCs colonies (day 0) and EBs at indicated stages of differentiation (day 4–12) were dissociated into single cells, stained with an Oct4 specific antibody and subjected to FACS analysis. Shaded curves depict Oct4 intracellular staining. Dashed lines indicate isotype controls and horizontal bars represent individual gates for positive Oct4 signals. The overlay shows exclusively Oct4 positive signals for each cell line. Numbers in red, blue and green indicate the percentages of Oct4 positive cells in wt, Dnmt1−/− and TKO ESCs/EBs, respectively, after subtraction of background signal from isotype controls.