sheep anti cited2  (R&D Systems)

Journal: Oncotarget

Article Title: Down-regulation of CITED2 attenuates breast tumor growth, vessel formation and TGF-β-induced expression of VEGFA

doi: 10.18632/oncotarget.14048

Figure Lengend Snippet: ( A – B ) Wild-type MDA-MB-231 and MDA-MB-468 cells were treated with or without 2.5 ng/ml TGF-β for 24 hours. (A) mRNA expression of VEGFA was determined by qRT-PCR. (B) Protein expression of VEGFA was determined by western blot analysis performed on equal amounts of protein obtained from serum-free conditioned media. Protein bands obtained from low (5 seconds) and high (20 seconds) exposure times are presented. ( C ) Localization of CITED2 to the VEGFA promoter as assessed by ChIP assay using anti-CITED2 or IgG antibodies (control) in wild-type MDA-MB-231 cells upon treatment with or without 2.5 ng/ml of TGF-β for 24 hours. ( D ) scramble and shCITED2-expressing MDA-MB-231 cells were treated with or without 2.5 ng/ml TGF-β for 24 hours and western blot analysis of VEGFA isoform expression was performed on equal amounts of protein obtained from serum-free conditioned media. (* P < 0.05, ** P < 0.01).

Article Snippet: Chromatin immunoprecipitation (ChIP) was performed on nuclear cell lysates utilizing anti-sheep CITED2 (R&D Systems) and anti-sheep IgG (Jackson ImmunoResearch) antibodies and following the SimpleChIP Enyzmatic Chromatin IP Kit (Cell Signaling Technology) protocol based on manufacturer's instructions.

Techniques: Expressing, Quantitative RT-PCR, Western Blot, Control