Both HSF1 and hiPS2-derived EBs were grown for 5–7 days in the presence of retinoic acid (RA) (1 μM, Sigma) and the Shh pathway agonist Purmorphamine (1.5 μM, Calbiochem) and generated EBs full of neural progenitors as judged by immunostaining for Brn2, Sox3, and Pax6 (a-f). Similar results were obtained using a different Shh pathway agonist (HhAg1.3, 500 nM, Curis, data not shown).