Fig. 4

Results of the expansion of TCLab hiPSCs with mTeSR1 and mTeSR3D supplemented with dextran sulfate (T1 + DS and T3D + DS, respectively) in the PBS MINI 0.1 bioreactor. At day 0, the reactors were inoculated with 1.5 × 10⁷ cells (2.5 × 10⁵ cells∙mL^− 1) and starting from 48 h post-inoculation, medium changes were perfomed according to the corresponding feeding regime. a-d Representative images of cell aggregates harvested from the bioreactor, analysed through brightfield microscopy at days 1 and 7 of culture in a T1 + DS and c T3D + DS (scale bars = 250 μm) and through fluorescence microscopy at day 7 of culture in b T1 + DS and d T3D + DS following staining with calcein AM (scale bars = 100 μm). e Growth kinetics of the cells, in terms of total cell number and fold increase. f-i Aggregate size dynamics of the cells, in terms of f average aggregate diameter, g coefficient of variation, and aggregate size distribution profiles of representative experiments at days 1, 4 and 7 post-inoculation for h T1 + DS and i T3D + DS. j-l Supernatant analysis of j glucose consumption and k lactate production, and l yield of lactate from glucose. mTeSR1 (T1) and mTeSR3D (T3D) data are shown for comparison