Extra-embryonic endoderm cells (Mount Sinai Hospital)

Blastocysts were plated on embryonic fibroblasts (EMFIs) feeder cells and cultured in RPMI 1640 supplemented with fetal bovine serum (20%), sodium pyruvate (1 mM), L-glutamine (2 mM ), penicillin/streptomycin (50 mg/ml each), β-mercaptoethanol (100 μM), human recombinant FGF4 (25 ng/ml ) and heparin (1 μg/ml ). Medium was changed every 3 days and after 15 days, XEN cells, observed in half the cultures, were passaged 1:1 onto new EMFIs in four-well plates. Cells are routinely cultured were routinely cultured on EMFIs in medium without FGF4 and heparin or on gelatin (0.1%), supplemented with 70% EMFI-conditioned medium . The latter is the preferred culturing method.