Material-dependent maintenance of hESC pluripotency. (A) Quantitative heat map of hESC NANOG expression relative to Matrigel in varying synthetic hydrogel-based culture conditions (n=3, n=5 in colony seeding conditions where ROCK inhibitor was removed, n=10 in single-cell seeding conditions where ROCK inhibitor was removed). Cells were cultured with either 0 or 5 µM ROCK inhibitor in maintenance culture, and hydrogels contained 0, 1, 2 or 4 mM cyclic RGD. The screen was performed once over the course of these studies. Conditions highlighted with a black arrowhead denote environments that maintained both hESC pluripotency and cell attachment over a 96-hour culture period without the use of ROCK Inhibitor. Conditions highlighted with a white arrowhead denote conditions that were further investigated for OCT3/4 expression in addition to NANOG expression. (B) Quantitative heat map of hESC attachment in varying culture conditions (n=3, n=5 in colony seeding conditions where ROCK inhibitor was removed, n=10 in single-cell seeding conditions where ROCK inhibitor was removed). The screen was performed once over the course of these studies.