Fig. 1: The main concept, mechanism, and evaluation of simple dialysis culture with FP003 addition.

a The schematic concept of high-density hiPSCs expansion in a miniaturized dual-compartment dialysis system. The permeability of small molecules such as b glucose and c lactic acid was also confirmed during 12 h (n = 3 biologically independent experiments). There is no difference in the mass transfer of these molecules between the normal medium without FP003 (−FP003) and with FP003 (+FP003). The cell-free penetration assays of various large molecules: d 20 kDa FITC, e 10 kDa FITC, f 4 kDa FITC show a decent capability to accumulate the macromolecule (n = 3 biologically independent experiments). g The mechanism of the FP003 addition in the culture medium to prevent excess agglomeration and mechanical stress. h The mechanical stress-induced secondary apoptosis or necrosis measured by intracellular lactate dehydrogenase (LDH) release (n = 6 biologically independent experiments) and i Primary apoptosis was reduced during 24 h of culture in gellan gum addition (n = 6 biologically independent experiments). To obtain the approximate number of secondary apoptotic/necrotic and primary apoptotic cells, the resulting value of LDH or caspase 3/7 assays was calibrated to the value of the graded number of intentionally necrotic- or apoptotic induced hPSCs line. The percentage shows the approximate number of secondary apoptotic/necrotic and primary apoptotic cells from the total inoculation cell number. Mean ± standard deviation are indicated in each graph. Statistical significance: ****p < 0.0001.