1. Clonogenic potential of Nanog:VNP subpopulations, under different culture conditions (serum/LIF and 2i/LIF).
Clonogenic potential of Nanog:VNP subpopulations, under different culture conditions (serum/LIF and 2i/LIF). (A) Representative histograms of FACS-sorted Nd subpopulations, grown in serum/LIF or 2i/LIF. VNPL and VNPH populations, as well as non-sorted bulk populations (All), were collected for posterior analysis. (B) Number and type of colonies (undifferentiated – AP positive, mixed or differentiated – AP negative; illustrative images in supplementary material Fig. S5) obtained from each mESC subpopulation (All, VNPL, VNPH) initially grown in serum/LIF (n=2). Subpopulations were plated at clonal density in serum/LIF or 2i/LIF media (600 cells per well of a six-well dish) and colony types analysed after 6 days. (C) Same as B for cells initially grown in 2i/LIF media (n=2), and replated in either serum/LIF or 2i/LIF. VNPL cells replated in serum/LIF generate predominantly mixed (71% and 92%) and differentiated colonies (27% and 6%), whereas VNPH cells generate fewer differentiated colonies and higher percentages of mixed and pure mESC colonies (more than 93% of total formed colonies). In addition, VNPH cells reveal higher clonogenic capacity than VNPL cells in 2i/LIF, although this difference is more attenuated when cells have been previously grown in 2i/LIF.
Elsa Abranches et al. Development 2014;141:
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