To label hiPSC NPCs with stable isotope amino acids, cells were cultured in NPC media for 10 days (~4–5 population doublings) with or without heavy isotope (13C15N)-enriched arginine and lysine (Cambridge Isotopes); heavy labeled cells were cultured in medium depleted of light arginine and lysine (Life Technologies) before harvest. Protein identification, quantification and analysis were done with Integrated Proteomics Pipeline—IP2 (Integrated Proteomics Applications, www.integratedproteomics.com/) using Pro-LuCID, DTASelect2, Census and QuantCompare.13
