Western blotting analysis using Phospho-PKR (T451) antibody (Cat#69322). Total cell lysates (30 μg) from various cell lines were loaded and separated by SDS-PAGE. The blot was incubated with Phospho-PKR (T451) antibody (Cat#69322, 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 Phospho-PKR (T451) antibody (Cat#69322). Phospho-PKR (T451) expression in wild type (WT) and phospho-PKR (T451) shRNA knockdown (KD) HeLa cells with 30 μg of total cell lysates. β-Tubulin serves as a loading control. The blot was incubated with Phospho-PKR (T451) antibody (Cat#69322, 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).
Immunocytochemical staining of HT-1080 cells with Phospho-PKR (T451) antibody (Cat#69322, 1:1,000). Nuclei were stained blue with DAPI; Phospho-PKR (T451) 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 phospho-PKR (T451) expression in HT-1080 cells using phospho-PKR (T451) antibody (Cat#69322, 1:2,000). Fixative solution: 4% Paraformaldehyde in 1x PBS; Permeabilization solution: 0.5% Triton X-100 in 1x PBS. Green, isotype control; red, phospho-PKR (T451).
Validation of PKR knockdown using flow cytometry. Wild-type(WT, Blue) and knockdown(KD, Green) HeLa cells were stained with Phospho-PKR (T451) antibody (Cat#69322, 1:2,000) and analyzed using CytoFLEX. Fixative solution: 4% Paraformaldehyde in 1x PBS; Permeabilization solution: 0.5% Triton X-100 in 1x PBS.
