Figure 1. Characterization of hESC cultures.

For each hESC line used, culture stage was assessed by immunocytochemistry and culture morphology. As an example, the RG7 hESC line exhibited markers consistent with pluripotency (SSEA4) but not differentiation (SSEA1) in the ESC condition (panel A), expressed nestin but not β-III-tubulin (TuJ1) in the neural stem cell (NSC) stage (panel B), and expressed β-III-tubulin with lesser levels of nestin in the neural-restricted precursor (NRP) stage (panel C). In addition, the ESC cultures grew as colonies, the NSC grew as isolated groups of cells, and the NRP produced processes reminiscent of neurons. To confirm this interpretation, mRNA microarrays were run on single samples of H1 ESC and NSC cultures to compare with HSF1 and HSF6 culture samples as previously defined [26]. Hierarchical clustering of all detected genes shows a clear hierarchical clustering of samples by culture stage. Here, this grouping is highlighted using genes distinguishing ESC from NSC (Student's t-test by stage, 5% false discovery rate [FDR]; panel D).