Human primary fibroblasts were derived from biopsied skin tissue samples. The fibroblasts were established and expanded with DMEM containing 10% autologous human serum. Using these fibroblasts, iPS cells were generated as described previously17 (link). Briefly, following electroporation of reprogramming factors with episomal vectors using the Neon system (Life Technologies), the cells were plated on a non-coated cell culture plate. iPS cells were induced by changing the medium to StemFit™. Twenty to thirty days after plating, iPS cell colonies were observed.
Blood cell-derived iPS cells were generated as described previously16 (link). Briefly, mononuclear cells were prepared from peripheral blood using the Ficoll-Paque PREMIUM (GE Healthcare) separation method. The cells were electroporated with episomal vectors using a Nucleofector 4D system (with P3 Primary Cell Kit, Lonza) and plated on rLN511E8-coated cell culture plates. The iPSCs were induced by changing the medium to StemFit™. Twenty to thirty days after plating, iPS cell colonies were observed. A similar method was used to generate Ff-hiPSCs from human cord blood (provided by the RIKEN Bioresource Center Cell Bank). We generated several clones of Ff-hiPSCs from each experiment.
The experimental protocols dealing with human subjects were approved by the institutional review board at our institute (Kyoto University Graduate School and Faculty of Medicine, Ethics Committee). Written informed consent was provided by each donor.
Blood cell-derived iPS cells were generated as described previously16 (link). Briefly, mononuclear cells were prepared from peripheral blood using the Ficoll-Paque PREMIUM (GE Healthcare) separation method. The cells were electroporated with episomal vectors using a Nucleofector 4D system (with P3 Primary Cell Kit, Lonza) and plated on rLN511E8-coated cell culture plates. The iPSCs were induced by changing the medium to StemFit™. Twenty to thirty days after plating, iPS cell colonies were observed. A similar method was used to generate Ff-hiPSCs from human cord blood (provided by the RIKEN Bioresource Center Cell Bank). We generated several clones of Ff-hiPSCs from each experiment.
The experimental protocols dealing with human subjects were approved by the institutional review board at our institute (Kyoto University Graduate School and Faculty of Medicine, Ethics Committee). Written informed consent was provided by each donor.
