Western blotting analysis using Bmi1 antibody (Cat#69531). Total cell lysates (30 μg) from various cell lines were loaded and separated by SDS-PAGE. The blot was incubated with Bmi1 antibody (Cat#69531,1:5,000) and HRP-conjugated goat anti rabbit secondary antibody (Cat#201,1:50,000) respectively. Image was developed using FeQ™ ECL Substrate Kit (Cat#226).
Western blotting analysis using Bmi1 antibody (Cat#69531). Bmi1 expression in wild type (WT) and Bmi1 shRNA knockdown (KD) 293T cells with 30 μg of total cell lysates. GAPDH serves as a loading control. The blot was incubated with Bmi1 antibody (Cat#69531,1:5,000) and HRP-conjugated goat anti rabbit secondary antibody (Cat#201,1:50,000) respectively. Image was developed using FeQ™ ECL Substrate Kit (Cat#226).
Immunohistochemistry was performed on paraffin-embedded human lymphoma using bmi1 antibody (Cat#69531, 1:200). Antigen retrieval was done in sodium citrate buffer (pH 6.0). DAB was used for detection, with hematoxylin counterstaining. Images were acquired using a Nikon Ci-L Plus microscope (40× objective). Scale bar: 25 μm.
Immunocytochemical staining of HeLa cells with Bmi1 antibody (Cat#69531, 1:1,000). Nuclei were stained blue with DAPI; Bmi1 was stained magenta with Alexa Fluor® 647. Images were taken using Leica stellaris 5. Protein abundance based on laser Intensity and smart gain: Medium. Fixative solution: 4% Paraformaldehyde in 1x PBS; Permeabilization solution: 0.5% Triton X-100 in 1x PBS. Scale bar: 20 μm.
Flow cytometric analysis of Bmi1 expression in HepG2 cells using Bmi1 antibody (Cat#69531, 1:2,000). Fixative solution: 4% Paraformaldehyde in 1x PBS; Permeabilization solution: 0.5% Triton X-100 in 1x PBS. Green, isotype control; red, Bmi1.
Validation of Bmi1 knockdown using flow cytometry. Wild-type(WT, Blue) and knockdown(KD, Green) HeLa cells were stained with Bmi1 antibody (Cat#69531, 1:2,000) and analyzed using BD flow cytometer. Fixative solution: 4% Paraformaldehyde in 1x PBS; Permeabilization solution: 0.5% Triton X-100 in 1x PBS.
