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Hum. Reprod. Advance Access published online on September 2, 2005
Human Reproduction, doi:10.1093/humrep/dei275
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© The Author 2005. Published by Oxford University Press on behalf of the European Society of Human Reproduction and Embryology. All rights reserved.
Received December 5, 2004
Revised May 24, 2005
Accepted July 26, 2005

Article

Serum-free medium cultivation to improve efficacy in establishment of human embryonic stem cell lines

Chen Gui-an 1* and Peng Hong-mei 2

1 Department of Obstetrics and Gynecology, Peking University Third Hospital, Beijing 100083; Peking University Stem Cell Research Center, Beijing 100083, China
2 Department of Obstetrics and Gynecology, Peking University Third Hospital, Beijing 100083; Department of Obstetrics and Gynecology, General Hospital of PLA, Beijing 100853; Peking University Stem Cell Research Center, Beijing 100083, China

* To whom correspondence should be addressed.
Chen Gui-an, E-mail: chenguian{at}bjmu.edu.cn


  Abstract

BACKGROUND: Serum-containing and serum-free media were used to derive human embryonic stem (HES) cells from donated oocytes and embryos. METHODS and RESULTS: Inner cell masses (ICM) were isolated by immuno-surgery. The HES cells were found to be easily obtained and expanded in a serum-free medium. The efficacy in establishing human embryonic stem cell lines improved in a serum-free medium compared with that in serum-containing media. Four HES cell lines were derived from 13 isolated ICM on mouse embryonic fibroblast feeder layers. All four cell lines possess the same characteristics and differentiating potency: normal 46,XX or 46,XY karyotype; and expressing a series of surface markers such as APase, SSEA-3, SSEA-4, TRA-1-60, TRA-1-81, but not SSEA-1. They can form embryoid bodies in suspension culture and develop teratomas comprising derivatives of three embryonic germ layers when injected into severe combined immunodeficient mice. CONCLUSION: These preliminary results suggest that serum-free cultivation may be superior to serum-containing cultivation for deriving human embryonic stem cells.

Keywords: embryonic stem cell/derivation/passage/serum-free cultivation.
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