While spontaneous differentiation is an undesired feature of expanding populations of embryonic stem cells, a variety of methods have been described for their intended differentiation into specialized cell types, such as the osteoblast or chondrocyte. Most commonly, differentiation initiation involves the aggregation of ESCs into a so-called embryoid body (EB), a sphere composed of approximately 15,000 differentiating cells. EB formation has been optimized through the years, for example through invention of the hanging drop protocol. Yet, it remains a highly laborious process. Here we describe the use of computer-controllable suspension bioreactors to form EBs in an automated and highly reproducible process and their subsequent differentiation along the osteoblast lineage. The development of the differentiating cells taken from bioreactor EBs to EBs formed in static control cultures through the hanging drop method will be compared.
