Serum replacement with a growth factor-free synthetic substance in culture medium contributes to effective establishment of mouse embryonic stem cells of various origins.
OBJECTIVE
To evaluate whether serum replacement with growth factor-free synthetic substances contributed to the effective establishment of embryonic stem (ES) cells.


METHODS
Randomized, prospective model study.


METHODS
Gamete and stem cell biotechnology laboratory at Seoul National University in Korea.


METHODS
F1 (C57BL6 x DBA2) mice.


METHODS
Blastocysts of different origins were cultured in serum-replaced media.


METHODS
Embryonic stem cell establishment.


RESULTS
Eight batches of ES cells were established from colony-forming inner cell mass cells after the replacement of fetal bovine serum (FBS) with synthetic knockout serum replacement (KSR) in mkDMEM. The established cells were positive for ES cell markers and formed both embryoid bodies in vitro and teratomas in vivo, but the established cell batches and control (transformed) ES cells responded differently to the culture media. Higher levels of cell viability were detected after the replacement with the 75:25 FBS-KSR mixture than with any other mixtures, and a gradual decrease in viability was detected as the KSR volume ratio was increased. The 75:25 FBS-KSR mixture-containing medium supported ES cell establishment of outbred ICR, F1, and F2 of C57BL6/DBA2; F1 parthenogenetic and ES cell-complemented tetraploid blastocysts; and single ES-cell cultures.


CONCLUSIONS
A serum-replaced medium could be used for effective ES-cell establishment of various origins.
Lee
Seung Tae
ST
Department of Food and Animal Biotechnology, Seoul National University, Seoul, Korea.
Oh
Se Woong
SW
Kim
Dae Yong
DY
Han
Jae Yong
JY
Moon
Shin Yong
SY
Lim
Jeong Mook
JM
D000818:Animals; D021461:Animals, Outbred Strains; D015415:Biomarkers / Q000378:metabolism; D001755:Blastocyst / Q000166:cytology / Q000187:drug effects; D002417:Cattle / Q000097:blood; D002470:Cell Survival; D002478:Cells, Cultured; D003470:Culture Media / Q000138:chemical synthesis* / Q000737:chemistry / Q000494:pharmacology*; D004622:Embryo, Mammalian / Q000166:cytology*; D005260:Female; D005312:Fetal Blood; D006133:Growth Substances / Q000032:analysis; D008297:Male; D051379:Mice; D008815:Mice, Inbred Strains; D013234:Stem Cells / Q000166:cytology* / Q000187:drug effects* / Q000378:metabolism / Q000502:physiology
D015415:Biomarkers; D003470:Culture Media; D006133:Growth Substances