A simple approach for mouse embryonic stem cells isolation and differentiation inducing embryoid body formation

Abstract

In the present study, we derived stem cells from hatched blastocyst-stage mouse embryos from C57BL/6 strain employing a knockout serum replacement instead of the traditional foetal calf serum, avoiding the use of immunosurgery. Although foetal 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 cells derived 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 the octamer binding protein 4 (Oct-4). Suspension culture in bacteriological dishes was better compared with hanging drop method for differentiation by means of embryoid body formation. The mouse embryonic stem cells obtained during this work showed spontaneous differentiation to derivatives of the three germ layers in culture media supplemented with foetal calf serum but not with knockout serum.