To further differentiate NPs into neurons, NPs were cultured on plates coated with 0.1% poly-l-ornithine (Sigma) and 10ng/ml of laminin (Invitrogen) in NDM without hLIF and bFGF at 25,000 ~ 50,000 cells/cm2. Media was replaced every 2 ~ 3 days. Differentiation was continued for up to 4 weeks in culture. Cells were harvested and RNA samples were obtained after two weeks to represent an early time point of neuronal differentiation (E) and four weeks to represent a later time point (L) of neuronal differentiation.
