Vera S. Brok-VolchanskayaDavid A. BenninKran SuknunthaLucas C. KlemmAnna HuttenlocherIgor SlukvinUniversity of Wisconsin-MadisonUniversity of Wisconsin School of Medicine and Public HealthMahidol UniversityWisconsin National Primate Research Center2020-01-272020-01-272019-12-10Stem Cell Reports. Vol.13, No.6 (2019), 1099-1110221367112-s2.0-85075885519https://repository.li.mahidol.ac.th/handle/20.500.14594/49997© 2019 The Authors Human induced pluripotent stem cells (hiPSCs) can serve as a versatile and scalable source of neutrophils for biomedical research and transfusion therapies. Here we describe a rapid efficient serum- and xenogen-free protocol for neutrophil generation, which is based on direct hematoendothelial programming of hiPSCs using ETV2-modified mRNA. Culture of ETV2-induced hematoendothelial progenitors in the presence of GM-CSF, FGF2, and UM171 led to continuous production of generous amounts of CD34+CD33+ myeloid progenitors which could be harvested every 8–10 days for up to 30 days of culture. Subsequently, myeloid progenitors were differentiated into neutrophils in the presence of G-CSF and the retinoic acid agonist Am580. Neutrophils obtained in these conditions displayed a typical somatic neutrophil morphology, produced reactive oxygen species, formed neutrophil extracellular traps and possessed phagocytic and chemotactic activities. Overall, this technology offers an opportunity to generate a significant number of neutrophils as soon as 14 days after initiation of differentiation.Mahidol UniversityBiochemistry, Genetics and Molecular BiologyArticleSCOPUS10.1016/j.stemcr.2019.10.007