iPSC FUNDAMENTALS
Induced pluripotent stem cells enable patient-specific organoid generation from blood or skin samples. Quality iPSC cultures require feeder-free maintenance (mTeSR, E8), regular passaging at 70-80% confluence, and validation of pluripotency markers (Oct4, Sox2, Nanog). Karyotype verification ensures genomic stability before differentiation.
HEPATOCYTE DIFFERENTIATION
- Day 0-5: Definitive endoderm induction (Activin A, Wnt3a)
- Day 5-10: Hepatic specification (BMP4, FGF2)
- Day 10-15: Hepatoblast expansion (HGF, OSM)
- Day 15-25: Hepatocyte maturation (dexamethasone, oncostatin M)
- Validation: Albumin secretion, CYP450 activity, urea production
CARDIOMYOCYTE DIFFERENTIATION
- Day 0-2: Mesoderm induction (CHIR99021 Wnt activation)
- Day 2-4: Cardiac specification (IWP2 Wnt inhibition)
- Day 4-8: Cardiomyocyte commitment (spontaneous beating observed)
- Day 8-30: Maturation (metabolic selection, electrical pacing)
- Validation: Troponin expression, calcium transients, contractility
QUALITY CONTROL
Differentiation efficiency varies between iPSC lines—optimize protocols for each line. Flow cytometry quantifies differentiation efficiency (target >70% positive cells). Functional assays (albumin ELISA, calcium imaging, electrophysiology) validate mature phenotype. Gene expression profiling confirms lineage-specific markers.