Human iPSCs were differentiated into definitive endoderm (DE) as previously described [39 (link)]. Briefly, 0.25 × 106 single cells were seeded onto L7™ hPSC Matrix-coated 24-well plates on day 0 with L7™ TFO2 media containing L7™ hPSC medium supplement and 10 µM Y27632 (ReproCELL USA, Inc., 04-0012, Beltsville, MD, USA). On day 1, the STEMdiff™ Definitive Endoderm Kit (Stem Cell Technologies, 05110, Vancouver, Canada) was used to induce DE differentiation according to manufacturer’s protocol.
The cells were washed, fixed on day 5 and stained for DE-specific markers SOX17 (R&D systems, AF1924, Minneapolis, MN, USA) and FOXA2 (Abcam, Ab108422, Cambridge, UK).
The cells were washed, fixed on day 5 and stained for DE-specific markers SOX17 (R&D systems, AF1924, Minneapolis, MN, USA) and FOXA2 (Abcam, Ab108422, Cambridge, UK).
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