VALIDATION IMAGES
The Hel.92.1.7 (H) cells were fixed with 4% PFA (10 min at r.t.) and then permeabilized with 90% ice-cold methanol for 20 min at -20℃,the cells then were incubated in 5%BSA to block non-specific protein-protein interactions (30 min at r.t.).Primary Antibody (green):Rabbit Anti-GATA1 antibody (bsm-54013R,1:100); Secondary Antibody (white blue): Goat anti-Rabbit IgG-BF488 (bs-60295G-BF488): 1 μg/test. Isotype Control (orange): Rabbit IgG (bs-0295P). Blank control (black): PBS. Acquisition of 20,000 events was performed.
4% Paraformaldehyde-fixed Hel.92.1.7 (H) cell; Triton X-100 at r.t. for 20 min; Antibody incubation with (GATA1) monoclonal Antibody, unconjugated (bsm-54013R) 1:100, 90 min at 37°C; followed by conjugated Goat Anti-Rabbit IgG antibody (green, bs-60295G-BF488) at 37°C for 90 min, DAPI (blue, C02-04002) was used to stain the cell nuclei. PBS instead of the primary antibody was used as the blank control.
25 ug total protein per lane of various lysates (see on figure) probed with GATA1 monoclonal antibody, unconjugated (bsm-54013R) at 1:1000 dilution and 4°C overnight incubation. Followed by conjugated secondary antibody incubation at r.t. for 60 min.