Abstract: Provided herein are methods of perfusion culturing a mammalian cell that include: providing a vessel containing a mammalian cell disposed in a first liquid culture medium having an osmolality of about 270 mOsm/kg to about 380 mOsm/kg; incubating the mammalian cell for a period of time at about 32° C. to about 39° C.; and during the period of time, continuously or periodically removing a first volume of the first liquid culture medium and adding to the first liquid culture medium a second volume of a second liquid culture medium, wherein the first and second volumes are about equal and the osmolality of the first and second liquid culture medium in the vessel is maintained at about 270 mOsm/kg to about 380 mOsm/kg over the period of time.

Abstract: The invention provides methods for differentiating pluripotent stem cells such as ES cells with improved progenitor and differentiated cell yield using low oxygen conditions and optionally in the absence of exogenously added differentiation factors.