A simple approach for mouse embryonic stem cells isolation and differentiation inducing embryoid body formation.
Stem cells were derived from hatched blastocyst-stage mouse embryos of the C57BL/6 strain employing a knockout serum replacement instead of the traditional fetal calf serum, thereby avoiding the use of immunosurgery. Although fetal calf serum was not good for isolation of stem cells, a combination of this serum plus knockout serum increased the expansion rate of the cell culture. The derived cells were capable of maintaining an undifferentiated state during several passages, as demonstrated by the presence of alkaline phosphatase activity, stage-specific embryonic antigen 1 (SSEA-1), and octamer binding protein 4 (Oct-4). Suspension culture in bacteriological dishes gave better results than the hanging drop method for differentiation by means of embryoid body formation. Mouse embryonic stem cells showed spontaneous differentiation into derivatives of the 3 germ layers in culture media supplemented with fetal calf serum but not with knockout serum.
Fagundez
Carol B
CB
Instituto de Ciencias Básicas y Medicina Experimental, Hospital Italiano de Buenos Aires, Potosí 4240, C1199ACL Ciudad Autónoma de Buenos Aires, Argentina. carol.fagundez@hospitalitaliano.org.ar
Loresi
Mónica A
MA
Ojea Quintana
Marcos E
ME
Delcourt
Stella M
SM
Testa
Roberto
R
Gogorza
Sebastián J
SJ
Argibay
Pablo F
PF
D000818:Animals; D001755:Blastocyst / Q000166:cytology; D018929:Cell Culture Techniques; D002454:Cell Differentiation; D002469:Cell Separation / Q000379:methods*; D003470:Culture Media; D053595:Embryonic Stem Cells / Q000166:cytology* / Q000378:metabolism; D016256:Lewis X Antigen / Q000378:metabolism; D051379:Mice; D008810:Mice, Inbred C57BL; D009028:Morula; D050814:Octamer Transcription Factor-3 / Q000378:metabolism
D003470:Culture Media; D016256:Lewis X Antigen; D050814:Octamer Transcription Factor-3