Insulin Secretion ELISA
Insulin secretion from hiPSC-derived pancreatic cells cultured on scaffolds was assessed by ultrasensitive insulin ELISA kit (ALPCO 80-INSHUU-E01.1) at stages 4, 5 and 6 of differentiation (days 1, 5 and 10 post seeding in 96-well plates and every other day in bioreactors). Following the manufacturer’s instructions, cell-culture supernatants, standard solutions, and controls were added to anti-insulin pre-coated microplate wells and incubated with detection antibody at room temperature with shaking for 1 hour. Subsequently, the microplate was washed six times with provided wash buffer and incubated with 3,3′,5,5′-Tetramethylbenzidine substrate for 30 minutes with shaking. After addition of the stop solution, the absorbance of each sample was measured at 450 nm.