iPSCs were differentiated into kidney organoids following the previously published protocol by Freedman et al. (5 (link)) (Figure 1B ). Briefly, iPSCs were dissociated with 1:3 Accutase and plated onto 24-well plates precoated with 0.5% GelTrex in mTeSR1 supplemented with 10 μM Y-27632 ROCK Inhibitor (STEMCELL Technologies). After 24 hours, another layer of GelTrex at 1.5% was added in mTeSR1 media. At the end of day 4, the medium was replaced with Advanced RPMI (Gibco) supplemented with 12 μM CHIR-99021 and 10 ng/ml noggin (STEMCELL Technologies). Approximately 60 hours later, the medium was changed to Advanced RPMI with B27 (Gibco). Organoids were cultured in this medium until collection at day 25.
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Kidney Organoid Differentiation from iPSCs
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Variable analysis
independent variables
- CHIR-99021 concentration (12 μM)
- Noggin concentration (10 ng/ml)
- Formation of kidney organoids
- IPSCs as starting cell type
- Accutase for cell dissociation
- GelTrex coating of culture plates
- MTeSR1 media supplemented with Y-27632 ROCK Inhibitor
- Advanced RPMI media supplementation with B27
- Culture duration (25 days)
- Not explicitly mentioned
- Not explicitly mentioned
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