Stemmacstm ips brew xf medium

All of the cell lines, taken from 3 healthy donors, were previously characterized. The hiPSC-A (C2a in [12 (link)]) line was generated using the lentivirus method while the hiPSC-B (IRX5-Wt in [13 (link)]; RRID:CVCL_B5QD) and hiPSC-C (WT8288 in [14 (link)]; RRID:CVCL_B5Q5) lines were generated using the Sendai virus method. The hiPSC lines were maintained at 37 °C, 5% CO₂, 21% O₂ in StemMACS^TM iPS Brew XF Medium (Miltenyi Biotec, Bergisch Gladbach, Germany) on culture plates coated with Matrigel^® hESC-Qualified Matrix (0.05 mg/mL, Corning, NY, USA). At 75% confluency, the cells were passaged using Gentle Cell Dissociation Reagent (STEMCELL^TM Technologies, Vancouver, Canada).

For the experiments two cell lines have been used H9 and hiPSCs derived from BJ fibroblasts. Cell culture in microfluidics is similar to traditional multiwells, concerning the type of medium and reagents used; the unique variables is the volume handling. In the microfluidic platform used in this work, the effective channel volume is 5.4 μl. For the media change, a total volume of 12 μl was perfused to guarantee an extensive medium refreshment and waste products removal.
Prior to cell seeding, microfluidic channels within the chip were filled with 4°C cold Matrigel Reduced Factor® (MRF, BD) 0.5% v/v in DMEM, incubated at room temperature for at least 1 h and washed with StemMACSTM iPS-Brew XF medium (Miltenyi Biotech).
hPSCs were then dissociated in single cell using Tryple (Thermo Fisher Scientific) and injected at 700 cells/mm2 density into the channels in medium supplemented with 10 μM ROCK inhibitor (Y-27632, Miltenyi Biotech) to preserve cell viability.
Because of the reduced medium volume in the microchannels, to prevent a significant evaporation, the microfluidic platform was placed in a 100 mm Petri dish and maintained in an isotonic bath with PBS for an optimal humidity of the chamber.