Generating iPSCs from Fibroblasts

Human fibroblasts were obtained from a 7-year-old male patient and were transduced with the CytoTune-iPS Reprogramming Kit (Life Technologies, Grand Island, NY, USA) according to the manufacturer’s instructions. Fibroblasts (2 × 10⁴ cells/cm2) were plated into 24-well culture plates, coated with gelatin containing Stempro 34 medium (Life Technologies), and supplemented with L-ascorbic acid (S4641, Sigma-Aldrich, St. Louis, MO, USA), stem cell factor (Peprotech, Rocky Hill, NJ, USA), and bFGF (Peprotech) 7 days before transduction. Transduced cells were seeded onto mouse embryonic fibroblast (MEF)-coated plates 1 week after transduction. After 3 weeks, colonies were selected and expanded in stem MACS medium (Miltenyi Biotec, Bergisch Gladbach, Germany) on plates coated with vitronectin (VTN; Life Technologies) [19 (link)].

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So S., Park Y., Kang S.S., Han J., Sunwoo J.H., Lee W., Kim J., Ye E.A., Kim J.Y., Tchah H., Kang E, & Lee H. (2022). Therapeutic Potency of Induced Pluripotent Stem-Cell-Derived Corneal Endothelial-like Cells for Corneal Endothelial Dysfunction. International Journal of Molecular Sciences, 24(1), 701.

Publication 2022

CytoTune-iPS Reprogramming Kit Stempro 34 medium L-ascorbic acid stem cell factor bFGF

Ascorbic acid Cells Embryonic Fibroblast Gelatin Human Male Mouse Patient Stem Stem cell factor Vitronectin

Corresponding Organization : University of Ulsan

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Variable analysis

independent variables

Transduction of human fibroblasts with the CytoTune-iPS Reprogramming Kit

dependent variables

Generation of induced pluripotent stem cell (iPSC) colonies

control variables

Fibroblasts (2 × 10^4 cells/cm2) were plated into 24-well culture plates, coated with gelatin containing Stempro 34 medium, and supplemented with L-ascorbic acid, stem cell factor, and bFGF 7 days before transduction
Transduced cells were seeded onto mouse embryonic fibroblast (MEF)-coated plates 1 week after transduction
IPSC colonies were selected and expanded in stem MACS medium on plates coated with vitronectin

controls

Positive control: Not explicitly mentioned.
Negative control: Not explicitly mentioned.

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