FIG. 4.

In vitro differentiation of AF-iPS into the three germ layers. (a) Floating embryoid bodies (EB) derived from AF-iPS (AF-3 clone 10) at day 7. (b, c) After EB formation, aggregates were transferred onto gelatin-coated plates and allowed to spontaneously differentiate for 10 days. Resulting cells had differentiated into neural cells with typical morphology (b) and Tuj1-specific positive immunoreactivity (c). (d) After EB differentiation amniotic fluid cell-derived iPS cells (AF-iPS), iPS cells from adult skin (Skin-iPS), and hES (HES-2) were stained with anti-nestin (an ectodermal marker), anti-α-smooth muscle actin and anti-troponin I (mesodermal markers), and anti-α-fetoprotein (AFP) (an endodermal marker).