polyclonal goat anti paired box 2  (R&D Systems)

Journal: Antioxidants

Article Title: An In Vitro Oxidative Stress Model of the Human Inner Ear Using Human-Induced Pluripotent Stem Cell-Derived Otic Progenitor Cells

doi: 10.3390/antiox13111407

Figure Lengend Snippet: Derivation and characterization of OPCs from hiPSCs. ( a ) Scheme illustrating the generation of OPCs from hiPSCs and corresponding stages of human development. hiPSCs resemble the inner cell mass (ICM), while OPCs correspond to the otocyst stage. ( b ) Representative bright-field images of SK8-A hiPSCs on day 0 and derived OPCs on day 20. Scale bars, 100 µm. ( c ) qRT-PCR data showing fold changes in mRNA expression of otic-related markers ( PAX2 , PAX8 , and GATA3 ) in OPCs compared to hiPSCs. The expression levels in hiPSCs were set to 1. Mean ± SD; * p < 0.05; *** p < 0.001. ( d ) Representative immunocytochemistry images showing protein expression of otic lineage markers in OPCs. hiPSCs were used as negative controls to verify the absence of false-positive signals. Scale bars, 100 µm. Abbreviations: PAX2 , paired box 2; PAX8 , paired box 8; GATA3 , GATA binding protein 3. See “Statistical analysis and reproducibility” in for statistics and experimental information.

Article Snippet: The cells were blocked with 5% goat serum (Gibco, Grand Island, NY, USA, #PCN5000) or 5% normal horse serum (Abcam, Cambridge, UK, #ab7484) in PBST for 1 h. Subsequently, the cells were incubated overnight at 4 °C with the following primary antibodies diluted in 1% BSA in PBST: goat polyclonal IgG PAX2 (R&D Systems, Minneapolis, MN, USA; #AF3364; 1:50), rabbit polyclonal PAX8 (Abcam, Cambridge, UK; #AB97477; 1:200), rabbit monoclonal GATA3 (Cell Signaling Technology, Danvers, MA, USA; #5852; 1:800), and mouse monoclonal cytochrome c (Cell Signaling Technology, Danvers, MA, USA; #12963; 1:300).

Techniques: Derivative Assay, Quantitative RT-PCR, Expressing, Immunocytochemistry, Binding Assay