ARHGEF12 Antibody / Rho guanine nucleotide exchange factor 12 (FY12180)

Immunohistochemical staining of ARHGEF12L using anti-ARHGEF12L antibody. ARHGEF12L was detected in a paraffin-embedded section of human colorectal adenocarcinoma tissue. Heat mediated antigen retrieval was performed in EDTA buffer (pH 8.0, epitope retrieval solution). The tissue section was blocked with 10% goat serum. The tissue section was then incubated with 2 ug/ml rabbit anti-ARHGEF12L antibody overnight at 4oC. Peroxidase Conjugated Goat Anti-rabbit IgG was used as secondary antibody and incubated for 30 minutes at 37oC. The tissue section was developed using an HRP secondary and DAB substrate.

Western blot analysis of ARHGEF12 using anti-ARHGEF12 antibody. Lane 1: human Hela whole cell lysates, Lane 2: human HepG2 whole cell lysates, Lane 3: rat testis tissue lysates, Lane 4: rat brain tissue lysates, Lane 5: mouse testis tissue lysates, Lane 6: mouse brain tissue lysates. After electrophoresis, proteins were transferred to a nitrocellulose membrane at 150 mA for 50-90 minutes. Blocked the membrane with 5% non-fat milk/TBS for 1.5 hour at RT. The membrane was incubated with rabbit anti-ARHGEF12 antibody at 0.5 ug/ml overnight at 4oC, then washed with TBS-0.1%Tween 3 times with 5 minutes each and probed with a goat anti-rabbit IgG-HRP secondary antibody at a dilution of 1:5000 for 1.5 hour at RT. The signal was developed using enhanced chemiluminescent. The expected band size for ARHGEF12 is at 173 kDa but is commonly observed at 220-260 kDa due to extensive post-translational modification (especially phosphorylation) and anomalous SDS–PAGE migration due to its large, highly coiled-coil–rich, multidomain structure.

Immunohistochemical staining of ARHGEF12L using anti-ARHGEF12L antibody. ARHGEF12L was detected in a paraffin-embedded section of human liver cancer tissue. Heat mediated antigen retrieval was performed in EDTA buffer (pH 8.0, epitope retrieval solution). The tissue section was blocked with 10% goat serum. The tissue section was then incubated with 2 ug/ml rabbit anti-ARHGEF12L antibody overnight at 4oC. Peroxidase Conjugated Goat Anti-rabbit IgG was used as secondary antibody and incubated for 30 minutes at 37oC. The tissue section was developed using an HRP secondary and DAB substrate.

Immunohistochemical staining of ARHGEF12L using anti-ARHGEF12L antibody. ARHGEF12L was detected in a paraffin-embedded section of human parotid acinar cell carcinoma tissue. Heat mediated antigen retrieval was performed in EDTA buffer (pH 8.0, epitope retrieval solution). The tissue section was blocked with 10% goat serum. The tissue section was then incubated with 2 ug/ml rabbit anti-ARHGEF12L antibody overnight at 4oC. Peroxidase Conjugated Goat Anti-rabbit IgG was used as secondary antibody and incubated for 30 minutes at 37oC. The tissue section was developed using an HRP secondary and DAB substrate.

Immunohistochemical staining of ARHGEF12L using anti-ARHGEF12L antibody. ARHGEF12L was detected in a paraffin-embedded section of human squamous cell carcinoma of cervix tissue. Heat mediated antigen retrieval was performed in EDTA buffer (pH 8.0, epitope retrieval solution). The tissue section was blocked with 10% goat serum. The tissue section was then incubated with 2 ug/ml rabbit anti-ARHGEF12L antibody overnight at 4oC. Peroxidase Conjugated Goat Anti-rabbit IgG was used as secondary antibody and incubated for 30 minutes at 37oC. The tissue section was developed using an HRP secondary and DAB substrate.

Immunohistochemical staining of ARHGEF12L using anti-ARHGEF12L antibody. ARHGEF12L was detected in a paraffin-embedded section of rat brain tissue. Heat mediated antigen retrieval was performed in EDTA buffer (pH 8.0, epitope retrieval solution). The tissue section was blocked with 10% goat serum. The tissue section was then incubated with 2 ug/ml rabbit anti-ARHGEF12L antibody overnight at 4oC. Peroxidase Conjugated Goat Anti-rabbit IgG was used as secondary antibody and incubated for 30 minutes at 37oC. The tissue section was developed using an HRP secondary and DAB substrate.

Immunofluorescent staining of ARHGEF12 using anti-ARHGEF12 antibody (green) and anti-Beta Tubulin antibody (red). ARHGEF12 was detected in immunocytochemical section of cell. Enzyme antigen retrieval was performed using IHC enzyme antigen retrieval reagent for 15 mins. The cells were blocked with 10% goat serum. And then incubated with 5 ug/ml rabbit anti-ARHGEF12 antibody and mouse anti-Beta Tubulin antibody overnight at 4oC. DyLight 488 Conjugated Goat Anti-Rabbit IgG and DyLight 550 Conjugated Goat Anti-Mouse IgG were used as secondary antibody at 1:500 dilution and incubated for 30 minutes at 37oC. Visualize using a fluorescence microscope and filter sets appropriate for the label used.

Immunoprecipitating (IP) ARHGEF12 in HepG2 whole cell lysate. Western blot analysis of ARHGEF12 using anti-ARHGEF12 antibody; Lane 1: HepG2 whole cell lysates (30ug); Lane 2: Rabbit control IgG instead of anti-ARHGEF12 antibody in HepG2 whole cell lysate; Lane 3: anti-ARHGEF12 antibody (2ug) + HepG2 whole cell lysate (500ug). After electrophoresis, proteins were transferred to a membrane. Then the membrane was incubated with rabbit anti-ARHGEF12 antibody at a dilution of 0.5 ug/ml and probed with a goat anti-rabbit IgG-HRP secondary antibody. The signal is developed using ECL Plus Western Blotting Substrate. A specific band was detected for ARHGEF12 at approximately 220 kDa. The expected band size for ARHGEF12 is at 173 kDa.

Flow Cytometry analysis of HepG2 cells using anti-ARHGEF12 antibody. Overlay histogram showing HepG2 cells stained with (Blue line). To facilitate intracellular staining, cells were fixed with 4% paraformaldehyde and permeabilized with permeabilization buffer. The cells were blocked with 10% normal goat serum. And then incubated with rabbit anti-ARHGEF12 antibody (1 ug/million cells) for 30 min at 20oC. DyLight 488 conjugated goat anti-rabbit IgG (5-10 ug/million cells) was used as secondary antibody for 30 minutes at 20oC. Isotype control antibody (Green line) was rabbit IgG (1 ug/million cells) used under the same conditions. Unlabelled sample (Red line) was also used as a control.