title: Traditional human embryonic stem cell culture.
abstract: Culturing human embryonic stem cells (hESCs) requires a significant commitment of time and resources. It takes weeks to establish a culture, and the cultures require daily attention. Once hESC cultures are established, they can, with skill and the methods described, be kept in continuous culture for many years. hESC lines were originally derived using very similar culture medium and conditions as those developed for the derivation and culture of mouse ESC lines. However, these methods were suboptimal for hESCs and have evolved considerably in the years since the first hESC lines were derived. Compared with mouse ESCs, hESCs are very difficult to culture - they grow slowly, and most importantly, since we have no equivalent assays for germline competence, we cannot assume that the cells that we have in our culture dishes are either stable or pluripotent. This makes it far more critical to assay the cells frequently using the characterization methods, such as karyotyping, immunocytochemistry, gene expression analysis, and flow cytometry, provided in this manual.
authors.affiliation: Neuroscience Laboratories, Centers for Neuroscience and Translational Research, Children's Hospital of Orange County Research Institute, Orange, CA, USA. pschwartz@choc.org
authors.forename: Philip H
authors.initials: PH
authors.lastname: Schwartz
authors.forename: David J
authors.initials: DJ
authors.lastname: Brick
authors.forename: Hubert E
authors.initials: HE
authors.lastname: Nethercott
authors.forename: Alexander E
authors.initials: AE
authors.lastname: Stover
mesh_terms: D000818:Animals; D002452:Cell Count; D018929:Cell Culture Techniques / Q000379:methods*; D002478:Cells, Cultured; D017364:Collagenases / Q000378:metabolism; D003114:Colony-Forming Units Assay; D015925:Cryopreservation; D004622:Embryo, Mammalian / Q000166:cytology; D053595:Embryonic Stem Cells / Q000166:cytology* / Q000378:metabolism; D005347:Fibroblasts / Q000166:cytology; D006801:Humans; D051379:Mice; D008858:Microscopy, Phase-Contrast; D039904:Pluripotent Stem Cells / Q000166:cytology / Q000378:metabolism
chemical_list: D017364:Collagenases