title: Efficient and scalable culture of single dissociated human pluripotent stem cells using recombinant E8 fragments of human laminin isoforms.
abstract: This unit describes a protocol for efficient expansion of human pluripotent stem cells (hPSCs). A key feature of this method is subculture of hPSCs by single-cell dissociation passaging on substrates coated with recombinant E8 fragments of human laminin isoforms (LM-E8s). LM-E8s, provide superior adhesion over intact laminin isoforms and Matrigel. Single hPSCs seeded on LM-E8s show accelerated migration and rapid reconstruction of clusters, resulting in robust survival and proliferation. This protocol yields 200-fold more hPSCs than conventional subculture methods in 1 month of culture. Furthermore, this protocol can be easily adapted to most hPSC lines in combination with the use of various xeno-free, defined culture media, and large-scale expansion of hPSCs is easily achievable to facilitate the practical applications of hPSCs.
authors: Department of Embryonic Stem Cell Research, Institute for Frontier Medical Sciences, Kyoto University, Kyoto, Japan.
authors: Takamichi
authors: T
authors: Miyazaki
authors: Department of Embryonic Stem Cell Research, Institute for Frontier Medical Sciences, Kyoto University, Kyoto, Japan.
authors: Eihachiro
authors: E
authors: Kawase
mesh_terms: D000818:Animals; D018929:Cell Culture Techniques / Q000379:methods*; D061252:Feeder Cells / Q000166:cytology; D006801:Humans; D007797:Laminin / Q000378:metabolism*; D051379:Mice; D039904:Pluripotent Stem Cells / Q000166:cytology*; D020033:Protein Isoforms / Q000378:metabolism; D011994:Recombinant Proteins / Q000378:metabolism*; D059010:Single-Cell Analysis
chemical_list: D007797:Laminin; D020033:Protein Isoforms; D011994:Recombinant Proteins
keywords: culture substrate; human pluripotent stem cells (hPSCs); laminin fragment; laminin isoform; single-cell dissociation culture
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