Definitive Endoderm Differentiation of Human Embryonic Stem Cells Combined with Selective Elimination of Undifferentiated Cells by Methionine Deprivation

: Human embryonic stem cells (ESCs) show a characteristic feature in that they are highly dependent on methionine metabolism. Undifferentiated human ESCs cannot survive under condition that methionine is deprived from culture medium. We describe here a procedure for definitive endoderm differentiation from human ESCs, in which human ESCs are subject to 10 days' (d) differentiation combined with methionine deprivation between differentiation days (d) 8 to (d) 10. Methionine deprivation results in elimination of undifferentiated cells from the culture with no significant loss of definitive endoderm cells, as compared to those cultured under complete condition throughout the whole culture period.