Development of an embryoid body-based screening strategy for assessing the hepatocyte differentiation potential of human embryonic stem cells following single-cell dissociation.

We have devised an embryoid body-based screening method for the selection of human embryonic stem cell (hESC) lines capable of forming functional hepatocyte-like cells (HLCs) after single-cell dissociation. The screening method highlighted one cell line from a panel of five that produced albumin-positive cells during embryoid body (EB) formation. Cell lines that did not produce albumin-positive cells during EB formation were shown to respond less well to directed differentiation following single-cell replating. Additionally, the seeding density of the pluripotent populations prior to differentiation was shown to exert a significant effect on the hepatic function of the final population of cells. In summary, we have developed a simple procedure that facilitates the identification of human hESC lines that tolerate single-cell replating and are capable of differentiating to HLCs. Although the hepatic function of cells produced by this method is ∼10-fold lower than our current gold standard stem cell-derived models, we believe that these findings represent an incremental step toward producing HLCs at scale.