Sample: Lane 1: Human HeLa cell lysates Lane 2: Human HT-1080 cell lysates Primary: Anti-TFRC (bsm-43165M) at 1/1000 dilution Secondary: IRDye800CW Goat Anti-Mouse IgG at 1/20000 dilution Predicted band size: 74/85 kDa Observed band size: 95 kDa

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 (TFRC) monoclonal Antibody, Unconjugated (bsm-43165M) 1:25, 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:Hela. Primary Antibody (green line): Mouse Anti-TFRC antibody (bsm-43165M) Dilution: 2ug/Test; Secondary Antibody (white blue line) : Goat anti-mouse IgG-FITC Dilution: 0.5ug/Test. Isotype control（orange line）：Normal Mouse IgG Protocol The cells were 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.