Sample: Lane 1: NCI-H292 cell lysate Lane 2: A549 cell lysate Lane 3: HepG2 cell lysate Primary: Anti-ALDH2 (bsm-51466M) at 1/1000 dilution Secondary: IRDye800CW Goat Anti-Mouse IgG at 1/20000 dilution Predicted band size: 54 kD Observed band size: 54 kD

Paraformaldehyde-fixed, paraffin embedded (human 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; Antibody incubation with (ALDH2 ) Monoclonal Antibody, Unconjugated (bsm-51466M) at 1:200 overnight at 4°C, followed by operating according to SP Kit(Mouse)(sp-0024) instructionsand DAB staining.

HepG2 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 (ALDH2) monoclonal Antibody, Unconjugated (bsm-51466M) 1:50, 90 minutes at 37°C; followed by a conjugated Goat Anti-Mouse IgG antibody at 37°C for 90 minutes, DAPI (blue, C02-04002) was used to stain the cell nuclei.

Blank control:HepG2. Primary Antibody (green line): Mouse Anti-ALDH2 antibody (bsm-51466M) Dilution: 1:50; Secondary Antibody : Goat anti-mouse IgG-PE 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.