title: Adapting human pluripotent stem cells to high-throughput and high-content screening.
abstract: The increasing use of human pluripotent stem cells (hPSCs) as a source of cells for drug discovery, cytotoxicity assessment and disease modeling requires their adaptation to large-scale culture conditions and screening formats. Here, we describe a simple and robust protocol for the adaptation of human embryonic stem cells (hESCs) to high-throughput screening (HTS). This protocol can also be adapted to human induced pluripotent stem cells (hiPSCs) and high-content screening (HCS). We also describe a 7-d assay to identify compounds with an effect on hESC self-renewal and differentiation. This assay can be adapted to a variety of applications. The procedure involves the culture expansion of hESCs, their adaptation to 384-well plates, the addition of small molecules or other factors, and finally data acquisition and processing. In this protocol, the optimal number of hESCs plated in 384-well plates has been adapted to HTS/HCS assays of 7 d.
authors.affiliation: Center for Stem Cell Biology, Memorial Sloan-Kettering Cancer Center, New York, NY, USA.
authors.forename: Sabrina C
authors.initials: SC
authors.lastname: Desbordes
authors.forename: Lorenz
authors.initials: L
authors.lastname: Studer
mesh_terms: D000818:Animals; D018929:Cell Culture Techniques; D002454:Cell Differentiation; D049109:Cell Proliferation; D003094:Collagen / Q000737:chemistry; D004338:Drug Combinations; D053595:Embryonic Stem Cells / Q000502:physiology*; D005434:Flow Cytometry / Q000379:methods; D057166:High-Throughput Screening Assays / Q000379:methods*; D006801:Humans; D007621:Karyotyping / Q000379:methods; D007797:Laminin / Q000737:chemistry; D051379:Mice; D039904:Pluripotent Stem Cells / Q000502:physiology*; D011509:Proteoglycans / Q000737:chemistry; D054852:Small Molecule Libraries
chemical_list: D004338:Drug Combinations; D007797:Laminin; D011509:Proteoglycans; D054852:Small Molecule Libraries; C059485:matrigel; D003094:Collagen