title: Chromosomal Modification in Human Embryonic Stem Cells Cultured in a Feeder-Free Condition after Single Cell Dissociation using Accutase.
abstract: Human embryonic stem (ES) cells are a potential source of cells for developmental studies and for a variety of applications in transplantation therapies and drug discovery. However, human ES cells are difficult to culture and maintain at a large scale, which is one of the most serious obstacles in human ES cell research. Culture of human ES cells on MEF cells after disassociation with accutase has previously been demonstrated by other research groups. Here, we confirmed that human ES cells (H9) can maintain stem cell properties when the cells are passaged as single cells under a feeder-free culture condition. Accutase-dissociated human ES cells showed normal karyotype, stem cell marker expression, and morphology. We prepared frozen stocks during the culture period, thawed two of the human ES cell stocks, and analyzed the cells after culture with the same method. Although the cells revealed normal expression of stem cell marker genes, they had abnormal karyotypes. Therefore, we suggest that accutase-dissociated single cells can be usefully expanded in a feeder-free condition but chromosomal modification should be considered in the culture after freeze-thawing.
authors.affiliation: Dept. of Biochemistry, College of Natural Sciences, Chungbuk National University, Cheongju 361-763, Korea ; Biotechnology Research Institute, Chungbuk National University, Cheongju 361-763, Korea.
authors.forename: Young-Eun
authors.initials: YE
authors.lastname: Kim
authors.affiliation: Dept. of Biochemistry, College of Natural Sciences, Chungbuk National University, Cheongju 361-763, Korea.
authors.forename: Jeong-A
authors.initials: JA
authors.lastname: Park
authors.affiliation: Dept. of Biochemistry, College of Natural Sciences, Chungbuk National University, Cheongju 361-763, Korea.
authors.forename: Yang-Wha
authors.initials: YW
authors.lastname: Ha
authors.affiliation: Dept. of Biochemistry, College of Natural Sciences, Chungbuk National University, Cheongju 361-763, Korea.
authors.forename: Sang-Kyu
authors.initials: SK
authors.lastname: Park
authors.affiliation: Institute of Reproductive Medicine and Population, Medical Research Center, Seoul National University, Seoul 110-799, Korea ; IVF Laboratory, Dept. of Obstetrics and Gynecology, Seoul National University Hospital, Seoul 110-744, Korea.
authors.forename: Hee Sun
authors.initials: HS
authors.lastname: Kim
authors.affiliation: Institute of Reproductive Medicine and Population, Medical Research Center, Seoul National University, Seoul 110-799, Korea ; IVF Laboratory, Dept. of Obstetrics and Gynecology, Seoul National University Hospital, Seoul 110-744, Korea.
authors.forename: Sun Kyung
authors.initials: SK
authors.lastname: Oh
authors.affiliation: Dept. of Biochemistry, College of Natural Sciences, Chungbuk National University, Cheongju 361-763, Korea ; Biotechnology Research Institute, Chungbuk National University, Cheongju 361-763, Korea.
authors.forename: Younghee
authors.initials: Y
authors.lastname: Lee
keywords: Accutase; Freeze-thawing; Human embryonic stem cell; Karyotype; Single cell dissociation