In vitro and in vivo differentiation potentials of the protein-iPS cells. (A) EB formation by suspension culture (left), followed by the attachment of EB onto gelatin-coated dishes for the induction of spontaneous differentiation induction (right). (B-C) Seven to 14 days after commencing spontaneous differentiation, cells were harvested, and RT-PCR (B) and immunocytochemistry (C) were performed. UD, undifferentiated; Diff, spontaneous differentiation; GFAP, glial fibrillary acidic protein; SMA, α-smooth muscle actin; AFP, α-fetoprotein. DAPI (4′,6-Diamidino-2-phenylindole; blue) for nuclei. (D) cFB-protein-iPS cells were injected into NOD/SID mice. Adult fibroblasts and ES cells served as controls. Four weeks later, tumors were harvested (arrows). Gross morphologies showed well-demarcated tumors. (E) Hematoxylin and eosin (H&E) staining of a tumor from cFB-protein-iPS cells showing a well-differentiated teratoma.