Culturing Human Pluripotent Stem Cells on a Feeder Layer

Traditionally, hPSCs are cultured on a mitotically inactivated feeder layer such as mouse embryonic fibroblasts (MEFs). Feeder cells provide a suitable substrate for the hPSCs to attach to and help maintain the pluripotency of hPSCs by secreting various proteins into the growth medium. This chapter addresses the techniques necessary to culture hPSCs on a feeder layer and will help you gain a better understanding of the basic culture methods of hPSCs. The methods that we will discuss are ones that have been used reliably over time by many researchers in the field. Only time and experience will tell what methods will work best for you and your particular cells. The two fundamental methods that will be covered are manual or “mechanical” passaging and enzymatic dissociation. Generation of feeder cells and their mitotic inactivation are discussed in Chapter 2.