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Passaging and colony expansion of human pluripotent stem cells by enzyme-free dissociation in chemically defined culture conditions
Beers J.3; Gulbranson D.R.4; George N.2; Siniscalchi L.I.3; Jones J.2; Thomson J.A.4; Chen G.3
2012-11-01
Source PublicationNature Protocols
ISSN17542189 17502799
Volume7Issue:11Pages:2029-2040
Abstract

This protocol describes an EDTA-based passaging procedure to be used with chemically defined E8 medium that serves as a tool for basic and translational research into human pluripotent stem cells (PSCs). In this protocol, passaging one six-well or 10-cm plate of cells takes about 6-7 min. This enzyme-free protocol achieves maximum cell survival without enzyme neutralization, centrifugation or drug treatment. It also allows for higher throughput, requires minimal material and limits contamination. Here we describe how to produce a consistent E8 medium for routine maintenance and reprogramming and how to incorporate the EDTA-based passaging procedure into human induced PSC (iPSC) derivation, colony expansion, cryopreservation and teratoma formation. This protocol has been successful in routine cell expansion, and efficient for expanding large-volume cultures or a large number of cells with preferential dissociation of PSCs. Effective for all culture stages, this procedure provides a consistent and universal approach to passaging human PSCs in E8 medium. © 2012 Nature America, Inc. All rights reserved.

DOI10.1038/nprot.2012.130
URLView the original
Indexed BySCIE
Language英語English
WOS Research AreaBiochemistry & Molecular Biology
WOS SubjectBiochemical Research Methods
WOS IDWOS:000310585200011
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Cited Times [WOS]:197   [WOS Record]     [Related Records in WOS]
Document TypeJournal article
CollectionFaculty of Health Sciences
Affiliation1.University of Wisconsin School of Medicine and Public Health
2.WiCell Institute
3.National Heart, Lung, and Blood Institute
4.Morgridge Institute for Research
Recommended Citation
GB/T 7714
Beers J.,Gulbranson D.R.,George N.,et al. Passaging and colony expansion of human pluripotent stem cells by enzyme-free dissociation in chemically defined culture conditions[J]. Nature Protocols,2012,7(11):2029-2040.
APA Beers J.,Gulbranson D.R.,George N.,Siniscalchi L.I.,Jones J.,Thomson J.A.,&Chen G..(2012).Passaging and colony expansion of human pluripotent stem cells by enzyme-free dissociation in chemically defined culture conditions.Nature Protocols,7(11),2029-2040.
MLA Beers J.,et al."Passaging and colony expansion of human pluripotent stem cells by enzyme-free dissociation in chemically defined culture conditions".Nature Protocols 7.11(2012):2029-2040.
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