For differentiation into mesoderm following culture in pre-differentiation media, spent media was removed and replaced with fresh CDM-PVA media containing bone morphogenic protein 4 (BMP4, 10 ng/ml, R&D Systems Inc.), FGF2 (20 ng/ml; CSCR, University of Cambridge), recombinant Activin-A (10 ng/ml; CSCR, University of Cambridge), LY29004 (10 mM, Promega, UK.), CHIR99021 (5 mM, Selleckchem) and subsequently cultured for three days. Media was changed daily.
