FIG. 2.

(A) The incident of cystic EB formation (the number of EBs per total number of colonies) decreases in the presence of BMP4 and significantly increases with the addition of Noggin after 1 week in culture. Dose response in cystic EB formation following 1 week in culture with either increasing doses of BMP4 or equimolar concentrations of Noggin (50 ng/mL) plus BMP4 (20 ng/mL). EGC differentiation is enhanced in the presence of BMP4 inhibitor Noggin. Cystic EB formation is enhanced after (B) 24 h and (C) 7 days following exposure to 50 ng/mL Noggin in culture compared with controls (insets D and E, respectively). Inset figures D and E represent a single colony grown in the presence of 20 ng/mL BMP4 alone after 24 h and 7 days. Lower-case letters denote statistically significant differences among treatments (t-tests, P < 0.05; n = 8). Indirect immunofluorescence of cryosectioned EBs demonstrate a fluid-filled center surrounded by cells labeled by markers of the (F) ectoderm (Nestin: red) and (G) endoderm (AFP: green) lineage but not mesoderm. STO feeder cells surrounding EBs do not express these markers. DAPI (blue) stains the nuclei. Scale bars: 150 μM. EB, embryoid body. Color images available online at www.liebertonline.com/scd.