Sample: Lane 1: Mouse NIH/3T3 cell lysates Lane 2: Human MCF-7 cell lysates Primary: Anti-PCGF4/BMI1 (PC2999) at 1/1000 dilution Secondary: Goat Anti-Rabbit IgG at 1/20000 dilution Predicted band size: 37 kDa Observed band size: 44 kDa

Blank control（black line）:NIH/3T3. Primary Antibody (green line): Rabbit Anti-PCGF4/BMI1 antibody (PC2999) Dilution:1ug/Test; Secondary Antibody（white blue line）: Goat anti-rabbit IgG-AF488 Dilution: 0.5ug/Test. Isotype control（orange line）: Normal Rabbit IgG Protocol The cells were fixed with 4% PFA (10min at room temperature)and then permeabilized with 90% ice-cold methanol for 20 min at -20℃, 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 20,000 events was performed.

Blank control（black line）:NIH/3T3. Primary Antibody (green line): Rabbit Anti-PCGF4/BMI1 antibody (PC2999) Dilution:1ug/Test; Secondary Antibody（white blue line）: Goat anti-rabbit IgG-AF488 Dilution: 0.5ug/Test. Isotype control（orange line）: Normal Rabbit IgG Protocol The cells were fixed with 4% PFA (10min at room temperature)and then permeabilized with 90% ice-cold methanol for 20 min at -20℃, 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 20,000 events was performed.