Figure 4

ICM growth pattern as a function of media. Inner cell mass (ICM) growth patterns of intact blastocysts in standard DMEM medium (a-e) and in knockout culture system (f-j). Intact blastocysts plated on feeder cells (a, f). ICM attachment and trophoblast outgrowth (b, g). Multilayer differentiation of ICM on day 5 (D5) after plating, note endodermal "rind" (c). Immediately after first dissociation (d, i) blastocysts cultured in standard DMEM failed to produce ESC colonies (e) and generated epithelioid cells. In the knockout system, ICM growth showed negligible endodermal contamination (h) and after first dissociation with trypsin (red T bar) (i) generated ESC colonies (j). Scale = 50 μm.