Western blotting analysis using RHOA antibody (Cat#61270). Total cell lysates (30 μg) from various cell lines were loaded and separated by SDS-PAGE. The blot was incubated with RHOA antibody (Cat#61270, 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).
Western blotting analysis using RHOA antibody (Cat#61270). RHOA expression in wild-type (WT) and RHOA shRNA knockdown (KD) HT-1080 cells with 20 μg of total cell lysates. β-Tubulin serves as a loading control. The blot was incubated with RHOA antibody (Cat#61270, 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).
Immunohistochemistry was performed on paraffin-embedded mouse spleen using RHOA antibody (Cat#61270, 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.
Immunohistochemistry was performed on paraffin-embedded mouse kidney using RHOA antibody (Cat#61270, 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 C2C12 cells with RHOA antibody (Cat#61270, 1:1,000). Nuclei were stained blue with DAPI; RHOA 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 HT-1080 cells using RHOA antibody (Cat#61270, 1:1,000), Top panel: wild-type (WT); Bottom panal: RHOA shRNA knockdown (KD). Nuclei were stained blue with DAPI; RHOA 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 RHOA expression in C2C12 cells using RHOA antibody (Cat#61270, 1:2,000). Fixative solution: 4% Paraformaldehyde in 1x PBS; Permeabilization solution: 100% Methanol, chill on -20℃ before use. Green, isotype control; red, RHOA.
Validation of RHOA knockdown using flow cytometry. Wild-type(WT, Blue) and knockdown(KD, Green) HT-1080 cells were stained with RHOA antibody (Cat#61270, 1:2,000) and analyzed using BD flow cytometer. Fixative solution: 4% Paraformaldehyde in 1x PBS; Permeabilization solution: 100% Methanol, chill on -20℃ before use.
