Simplified three-dimensional culture system for long-term expansion of embryonic stem cells

Figure 2 Effect of cell concentration on the rate of scaffold degradation and embryonic stem cells colony size. A: Swelling tests of floating scaffolds prepared with 7.5% thiol substitution were used to compare the rate of degradation of scaffolds prepared with various cell concentrations (0, 2 × 10⁶ cells/mL and 4 × 10⁶ cells/mL). Scaffolds with and without ESCs were incubated in ESC medium and weighed periodically. Data expressed as the mean swelling ratio (W_s/W_i) ± SE (n = 3); B: Representative composite confocal images of ESCs grown in floating scaffolds encapsulating 2 × 10⁶ cells/mL (B1 and B2) and 4 × 10⁶ cells/mL (B3 and B4) at day-4 and day-18, respectively. Scale bars = 100 μm; C: Quantification of ESC colony size in floating scaffolds encapsulating 2 × 10⁶ cells/mL or 4 × 10⁶ cells/mL at day-4 and day-18 of culture. Cell colony size was expressed as mean diameter (μm) ± SE (n = 3); D: Growth of low concentrations of ESCs in 3-D scaffolds. Light micrographs of the scaffolds encapsulating 1 × 10⁴ cells/mL at day-0 and day-7 (D1 and D2, respectively) and 1 × 10⁵ cells/mL at day-0 and day-7 (D3 and D4 respectively). Scale bars = 100 μm. ESCs: Embryonic stem cells; 3-D: Three-dimensional.