Journal: Stem Cell Research & Therapy
Article Title: Targeted genome engineering in human induced pluripotent stem cells from patients with hemophilia B using the CRISPR-Cas9 system
Figure Lengend Snippet: Differentiation of iPSCs into hepatocytes and characterization of hepatocytic functions. a Schematic showing stepwise protocol for producing hepatocytes from iPSCs. b Immunofluorescence staining showed expression of SOX17, FOXA2, and GATA4 on day 5, HNF4α on day 10, AFP on day 15, and ALB on day 20. c Flow cytometry analysis showed expression of AFP and ALB > 90%. d qRT-PCR showed relative expression of HNF4α, AFP, ALB, TDO2, and TAT relative to GAPDH (n = 3 independent experiments for each sample). e Expression of pluripotent markers (OCT4, SOX2, and NANOG) and hepatic markers (AFP, ALB, HNF4α, TAT, TDO2, and CYP3A4) between iPSC-insertion and iPSC-insertion-Heps compared by qRT-PCR (n = 3 independent experiments for each sample). After differentiation, expression of pluripotent markers decreased, while expression of hepatic markers increased. All scale bars represent 100 μm. BMP bone morphogenetic protein, bFGF basic fibroblast growth factor, iPSC induced pluripotent cell, HGF, hepatocyte growth factor, DAPI 4′,6-diamidino-2-phenylindole, AFP alpha-fetoprotein, ALB albumin, GAPDH glyceraldehyde 3-phosphate dehydrogenase
Article Snippet: Quantitative real-time PCR (qRT-PCR) analysis was performed in triplicate, using a Step-One-Plus Real-Time PCR system (Applied Biosystems, Foster City, CA, USA).
Techniques: Immunofluorescence, Staining, Expressing, Flow Cytometry, Cytometry, Quantitative RT-PCR