Generating Neural Inducible Stem Cells

Neural induced differentiation Media (NGD) and neural induction medium (NIM) were prepared according to previous reports [43 (link), 44 (link)]. Compound C (Medchem Express LLC, Monmouth, UK, 2.5 μM), FGF2 (Sino Biological Inc., Beijing, China, 20 ng/mL), EGF (Sino Biological Inc., 20 ng/mL), SB431542 (Medchem Express LLC., 10 μM), DMH-1 (Medchem Express LLC., 2 μM), LIF (Sino Biological Inc., 10 ng/mL), LDN193189 (Medchem Express LLC., 0.25 μM) and insulin (Beijing Solarbio Science & Technology Co., Ltd., Beijing, China, 10 ng/mL) were added to medium as described previously [6 , 43 (link)–48 (link)]. iPSCs^TLX were seeded onto a matrigel-coated 6-well plate at a density of 2.5 × 10⁴ cells/cm². The medium was changed to a differentiation medium on the next day. On day 6, cells were digested by Accutase™ for 7 min at 37 °C. Then iNSCs^TLX were seeded into a vitronectin-coated 6-well plate at a density of 2.5 × 10⁴ cells/cm² and passaged every five days.