The characteristics of the ES cells and their neural derivatives. (A) Schematic procedure of SFEB for neural induction of ES cells. Undifferentiated ESC 26 cells were characterized by pluripotent markers such as Oct4 (B), Nanog (C) and Sox2 (D). The 46C ES-derived GFP+ cells were coexpressed with neural markers, such as nestin (E), pax6 (F), N-cadherin (G), Zic1 (H), but not GFAP (I) on day 6. Nuclei of ES cells were stained with DAPI in blue (B-I). ESC 26 cells were treated with 20, 40, and 80 ng/ml FGF1 from day 1 through day 3 and the N-cadherin+ colonies were estimated under fluorescent microscope (J) on day 6 from three independent experiments. A cell cluster with over 50 μm was counted as a colony and a colony was N-cadherin positive if over half of the cells in the colony expressed N-cadherin. Scale bar, 10 μm in B.