Human Embryonic Stem Cells

Description

Our human ES cells are grown on a feeder layer of radiation inactivated mouse embryonic fibroblasts (MEF) to prevent differentiation and sustain pluripotency. The mouse feeder cell layer is seeded the day prior to propagating human ES cells. Two days after plating the human ES cells the first small colonies are visible.

For 6 to 7 days, the colonies of hES cells undergo exponential growth. To maintain the primitive state of hES cells, the colonies must be dispersed and replated on a fresh MEF feeder layer when they reach a critical size or before they make contact with adjacent hES colonies. The mechanical and enzymatic dispersion of mature ES colonies gives rise to small colony fragments which will develop into mature colonies following further propagation.

The strategy of this culture technique is to prevent differentiation of human ES cells and to maintain the cells under optimal conditions for growth.  Our Core will provide exponentially growing undifferentiated cells that have a  normal karyotype and are validated for expression of markers for the non-differentiated phenotype. These parameters for the primitive diploid state are monitored by the Core on a frequent schedule.

Replating of hES cells on tissue culture dishes without the mouse feeder layer or growth factor supplementation leads to differentiation, forming cell spheres called embryoid bodies. Embryoid bodies contain cells of meso-, endo- and ectodermal lineages. Further controlled differentiation of each embryoid body can generate various committed cell phenotypes.