Identification of differentiation markers at each stage after differentiation induction of ES cells at each gelatin concentration. Mouse ES cells (OG2, 1 × 105 cells) were seeded and incubated on a 6-well culture plate coated with various concentrations of gelatin. For differentiation induction, ES cells were treated with a differentiation induction medium supplemented with 1 μM retinoic acid for 4 days. (A-H) Expression levels of pluripotency (A-C) and differentiation markers (D-H) were analyzed using prepared cDNAs by q-RT PCR. The graphs of each fold change were drawn by Prism 6 (GraphPad Software). Data were normalized to controls and presented as mean ± SEM of N = 3 experiments (*p < 0.05, **p < 0.01, compared with uncoating group; #p < 0.01, compared with undifferentiated mouse ES cell).