Western blotting analysis using creatine kinase B type antibody (Cat#63172). Total cell lysates (30 μg) from various cell lines were loaded and separated by SDS-PAGE. The blot was incubated with creatine kinase B type antibody (Cat#63172, 1:5,000) and HRP-conjugated goat anti-rabbit secondary antibody (Cat#201, 1:20,000) respectively. Image was developed using FeQ™ ECL Substrate Kit (Cat#226).
Western blotting analysis using creatine kinase B type antibody (Cat#63172). Creatine kinase B type expression in wild-type (WT) and creatine kinase B type (CKB) shRNA knockdown (KD) HeLa cells with 20 μg of total cell lysates. Hsp90 α serves as a loading control. The blot was incubated with creatine kinase B antibody (Cat#63172, 1:5,000) and HRP-conjugated goat anti-rabbit secondary antibody (Cat#201, 1:20,000) respectively. Image was developed using NaQ™ ECL Substrate Kit (Cat#716).
Immunohistochemistry was performed on paraffin-embedded human glioblastoma using creatine kinase B type antibody (Cat#63172, 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 creatine kinase B type antibody (Cat#63172, 1:1,000). Nuclei were stained blue with DAPI; Creatine kinase B type 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.
Immunocytochemical staining of HeLa cells using creatine kinase B type antibody (Cat#63172, 1:1,000), Top panel: wild-type (WT); Bottom panal: Creatine kinase B type shRNA knockdown (KD). Nuclei were stained blue with DAPI; Creatine kinase B type was stained magenta with Alexa Fluor® 647. 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 creatine kinase B type expression in HeLa cells using creatine kinase B type antibody (Cat#63172, 1:2,000). Fixative solution: 4% Paraformaldehyde in 1x PBS; Permeabilization solution: 0.5% Triton X-100 in 1x PBS. Green, isotype control; red, Creatine kinase B type.
Validation of creatine kinase B type knockdown using flow cytometry. Wild-type(WT, Blue) and knockdown(KD, Green) HeLa cells were stained with creatine kinase B type antibody (Cat#63172, 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.
