Aatf antibody

In brief, after baking at 65 °C for 2 h, 5‐μm‐thickness sections were deparaffinized in xylene, hydrated using a graded series of alcohol (100%, 95%, and 85%) and rinsed with PBS. Antigenic retrieval was performed by immersing in citric acid and microwaving. To block any non‐specific binding, the sections were treated using 0.3% hydrogen peroxide solution for 15 min. The sections were then incubated overnight at 4 °C with AATF antibody (Sigma‐Aldrich, St Louis, MO, USA, 1 : 100) and examined using HRP EnVision Systems (Dako, Shanghai, China). Finally, sections were counterstained with hematoxylin and visualized.

Endothelial cell growth medium (EGM), extracellular matrix gel (ECM), TRIzol, RIPA buffer, collagenase A, and AATF antibody were procured from Sigma-Aldrich (St. Louis, USA). Dulbecco’s modified eagle’s medium (DMEM), giemsa stain, hematoxylin, 8μm-transwell inserts, and 24- well plates were procured from HiMedia laboratories (India). Lipofectamine 3000, penicillin/streptomycin, glutamine, FBS were from Invitrogen (USA); WST-1 cell proliferation assay kit from Takara Bio Inc. (Shinga, Japan); antibodies to IgG, PEDF, pAkt, pErk1/2, pFAK, Akt, Erk, and control and AATF shRNA from Santa Cruz Biotechnology, Inc. (CA, USA); antibodies to β-actin from Cell Signaling Technologies (USA); verso cDNA synthesis kit, and DyNamo Colorflash SYBR green kit from Thermo Fisher Scientific (USA); western bright ECL HRP substrate from Advansta (USA); western blotting materials were from BioRad; primers from Integrated DNA Technologies (IDT) (Iowa, USA), and all other reagents were obtained from Thermo Fisher Scientific or Sigma.