(Tris-Glycine gel) Discontinuous SDS-PAGE (reduced) with 5% enrichment gel and 15% separation gel.

Western Blot
Positive WB detected in A375 whole cell lysate
All lanes CD146 antibody at 0.6μg/ml
Secondary
Goat polyclonal to rabbit IgG at 1/50000 dilution
Predicted band size: 72 KDa
Observed band size: 120 KDa

IHC image of CSB-RA013563A0HU diluted at 1:100 and staining in paraffin-embedded human breast cancer performed on a Leica BondTM system. After dewaxing and hydration, antigen retrieval was mediated by high pressure in a citrate buffer (pH 6.0). Section was blocked with 10% normal goat serum 30min at RT. Then primary antibody (1% BSA) was incubated at 4℃ overnight. The primary is detected by a biotinylated secondary antibody and visualized using an HRP conjugated SP system.

Immunofluorescence staining of U251 cells with CSB-RA013563A0HU at 1:37,counter-stained with DAPI. The cells were fixed in 4% formaldehyde, permeabilized using 0.2% Triton X-100 and blocked in 10% normal Goat Serum. The cells were then incubated with the antibody overnight at 4℃.The secondary antibody was Alexa Fluor 488-congugated AffiniPure Goat Anti-Rabbit IgG (H+L).

Overlay histogram showing A375 cells stained with CSB-RA013563A0HU (red line) at 1:50. The cells were fixed with 70% Ethylalcohol (18h) and then permeabilized with 0.3% Triton X-100 for 2 min. The cells were then incubated in 1x PBS /10% normal goat serum to block non-specific protein-protein interactions followed by primary antibody for 1 h at 4℃.The secondary antibody used was FITC goat anti-rabbit IgG (H+L) at 1/200 dilution for 1 h at 4℃. Control antibody (green line) was used under the same conditions. Acquisition of >10,000 events was performed.