Fig. 2.

Overview of the experimental strategy (numbered 1–6) to identify antibody-expressing ES colonies that resist differentiation through blockade of FGF signaling. Antibody populations binding to FGF4, FGFR1β, and FGFR2β were selected from a phage display library and were cloned into the targeting vector pROSA-ic. Homologous recombination was used to direct integration into the ubiquitously expressed Rosa-26 genomic locus of individual Oct4–ΔPE-GFP ES cells. Puromycin-resistant colonies were selected, antibody expression was induced, and ES colonies were subjected to differentiation in semisolid medium for 2.5–4.5 d. Clones that resisted differentiation (judged by retention of round morphology and Oct4-ΔPE-GFP expression) were picked and the antibody genes retrieved by PCR for further use.