Direct Comparison of Four Hematopoietic Differentiation Methods from Human Induced Pluripotent Stem Cells

Induced pluripotent stem cells (iPSCs) are an invaluable resource for the study of human disease. However, there are no standardized methods for differentiation into hematopoietic cells, and there is a lack of robust, direct comparisons of different methodologies. In the current study we improved a feeder-free, serum-free method for generation of hematopoietic cells from iPSCs, and directly compared this with three other commonly used strategies with respect to efficiency, repeatability, hands-on time, and cost. We also investigated their capability and sensitivity to model genetic hematopoietic disorders in cells derived from Down syndrome and β-thalassemia patients. Of these methods, a multistep monolayer-based method incorporating aryl hydrocarbon receptor hyperactivation ("2D-multistep") was the most efficient, generating significantly higher numbers of CD34⁺ progenitor cells and functional hematopoietic progenitors, while being the most time- and cost-effective and most accurately recapitulating phenotypes of Down syndrome and β-thalassemia.