Phospho-VEGF receptor 2 (Tyr1059) Rabbit Polyclonal Antibody

Blank control: Huvec. Primary Antibody (green line): Rabbit Anti-phospho-VEGF receptor 2 (Tyr1059) antibody (orb99143), dilution: 2 µg/10^6 cells, Isotype Control Antibody (orange line): Rabbit IgG. Secondary Antibody: Goat anti-rabbit IgG-AF647, dilution: 1 µg/Test. Protocol, The cells were fixed with 4% PFA (10 min at room temperature) and then permeabilized with 90% ice-cold methanol for 20 min at -20°C. The cells were then incubated in 5% BSA to block non-specific protein-protein interactions for 30 min at room temperature. Cells stained with Primary Antibody for 30 min at room temperature. The secondary antibody used for 40 min at room temperature. Acquisition of 20000 events was performed.

Sample: HUVEC (Hu) Lysate at 30 ug, Primary: Anti-phospho-VEGF receptor 2 (Tyr1059) (orb99143) at 1/300 dilution, Secondary: IRDye800CW Goat Anti-Rabbit IgG at 1/20000 dilution, Predicted band size: 147 kD, Observed band size: 147 kD.

Tissue/Cell: human kidney tissue, 4% Paraformaldehyde-fixed and paraffin-embedded, Antigen retrieval: citrate buffer (0.01M, pH6.0), Boiling bathing for 15 min, Block endogenous peroxidase by 3% Hydrogen peroxide for 30 min, Blocking buffer (normal goat serum) at 37°C for 20 min, Incubation: Anti-VEGFR Polyclonal Antibody, Unconjugated (orb99143) 1:200, overnight at 4°C, followed by conjugation to the secondary antibody and DAB staining.