Figure 2.

seBC demonstrate functional maturity. A: Representative perifusion analysis of imBC, seBC, and hIslets; 20–25 clusters were analyzed per sample, and data are presented as % of total insulin in cluster pellet recovered. Total insulin content of pellet recovered after perifusion (B) and relative insulin secretion during perifusion of imBC (n = 3 independent differentiation experiments), seBC (n = 5 independent differentiation experiments), and hIslet (n = 3 independent human islets prep) (C, D, and E, respectively) (data normalized to basal [0.5 mmol/L glucose] secretion). F: Representative images of imBC (left) and seBC clusters (right) displaying pINSGFP, Ca²⁺ indicator (Rhod2 AM) labeling, and overlay; scale bar is 10 µm. G: Color map (top) of islets displayed in F showing magnitude and extent of Ca²⁺ elevations at 2 mmol/L and 11 mmol/L glucose; scale bar is 10 µm, and white arrows point to cells chosen for time courses (bottom). Representative time courses (bottom) of individual cells from the imBC and seBC clusters displayed in F at 2 mmol/L and 11 mmol/L glucose; scale bar is 20% change from mean. H: Fraction of area within intact cluster showing elevations in Ca²⁺ activity at 2 mmol/L and 11 mmol/L glucose (n = 3 independent differentiation experiments with >10 clusters measured per condition). Human islet data quantified in the same manner is included for reference (21). I: Fold change in Ca²⁺ activity when glucose is elevated from 2 mmol/L to 11 mmol/L glucose (n = 3 independent differentiation experiments with >10 clusters measured per condition). *P < 0.05; **P < 0.01; ***P < 0.001. B–E: Error bars are representative of the mean ± SD. H and I: Error bars are representative of the mean ± SEM. AU, arbitrary units; Gluc, glucose; ns, not significant; sec, seconds.