Sample:
Lane 1: Hela cell lysates
Lane 2: HL60 cell lysates
Lane 3: T47D cell lysates
Primary: Anti-EZH2 (bsm-51468M) at 1:2000 dilution
Secondary: IRDye800CW Goat Anti-Mouse IgG at 1/20000 dilution
Predicted band size: 82 kD
Observed band size: 100 kD

Paraformaldehyde-fixed, paraffin embedded (rat brain); Antigen retrieval by boiling in sodium citrate buffer (pH6.0) for 15min; Block endogenous peroxidase by 3% hydrogen peroxide for 20 minutes; Blocking buffer (normal goat serum) at 37°C for 30min; Incubation with (EZH2) Monoclonal Antibody, Unconjugated (bsm-51468M) at 1:200 overnight at 4°C, followed by operating according to SP Kit(Mouse)(sp-0024) instructionsand DAB staining.

Hela cell; 4% Paraformaldehyde-fixed; Triton X-100 at room temperature for 20 min; Blocking buffer (normal goat serum, C-0005) at 37°C for 20 min; Antibody incubation with (EZH2) monoclonal Antibody, Unconjugated (bsm-51468M) 1:25, 90 minutes at 37°C; followed by a conjugated Goat Anti-Mouse IgG antibody at 37°C for 90 minutes.Dylight® 554 Phalloidin was used to stain the cell Cytoplasmic .

U251 cell; 4% Paraformaldehyde-fixed; Triton X-100 at room temperature for 20 min; Blocking buffer (normal goat serum, C-0005) at 37°C for 20 min; Antibody incubation with (EZH2) monoclonal Antibody, Unconjugated (bsm-51468M) 1:25, 90 minutes at 37°C; followed by a conjugated Goat Anti-Mouse IgG antibody at 37°C for 90 minutes.Dylight® 554 Phalloidin was used to stain the cell Cytoplasmic .

Blank control（black line）:Jurkat.
Primary Antibody (green line): Mouse Anti-EZH2 antibody (bsm-51468M)
Dilution:1ug/Test;
Secondary Antibody（white blue line）: Goat anti-mouse IgG-FITC
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.