Anti-ARF6 Antibody

• Catalog Number: orb371721
• Category: Antibodies
• Description: Anti-ARF6 Antibody
• Clonality: Polyclonal
• Species/Host: Rabbit
• Isotype: Rabbit IgG
• Conjugation: Unconjugated
• Reactivity: Human, Mouse, Rat
• Form/Appearance: Lyophilized
• Concentration: Adding 0.2 ml of distilled water will yield a concentration of 500 μg/ml.
• Immunogen: E.coli-derived human ARF6 recombinant protein (Position: G2-S175). Human ARF6 shares 100% amino acid (aa) sequence identity with both mouse and rat ARF6.
• UniProt ID: P62330
• MW: 20 kDa
• Tested applications: FC, ICC, IF, IHC, WB
• Application notes: Western blot, 0.1-0.5μg/ml, Human, Mouse, Rat Immunohistochemistry (Paraffin-embedded Section), 0.5-1μg/ml, Human, Rat Immunocytochemistry/Immunofluorescence, 2μg/ml, Human Flow Cytometry (Fixed), 1-3μg/1x106 cells, Human. Add 0.2ml of distilled water will yield a concentration of 500ug/ml
• Antibody Type: Primary Antibody
• Storage: Maintain refrigerated at 2-8°C for up to 2 weeks. For long term storage store at -20°C in small aliquots to prevent freeze-thaw cycles.
• Alternative names: ADP-ribosylation factor 6; ARF6
• Note: For research use only

IHC analysis of ARF6 using anti-ARF6 antibody. ARF6 was detected in paraffin-embedded section of human melanoma tissue. Heat mediated antigen retrieval was performed in EDTA buffer (pH8.0, epitope retrieval solution). The tissue section was blocked with 10% goat serum. The tissue section was then incubated with 1 µg/ml rabbit anti-ARF6 Antibody overnight at 4°C. Biotinylated goat anti-rabbit IgG was used as secondary antibody and incubated for 30 minutes at 37°C. The tissue section was developed using Strepavidin-Biotin-Complex (SABC) with DAB as the chromogen.

IHC analysis of ARF6 using anti-ARF6 antibody. ARF6 was detected in paraffin-embedded section of rat cardiac muscle tissue. Heat mediated antigen retrieval was performed in EDTA buffer (pH8.0, epitope retrieval solution). The tissue section was blocked with 10% goat serum. The tissue section was then incubated with 1 µg/ml rabbit anti-ARF6 Antibody overnight at 4°C. Biotinylated goat anti-rabbit IgG was used as secondary antibody and incubated for 30 minutes at 37°C. The tissue section was developed using Strepavidin-Biotin-Complex (SABC) with DAB as the chromogen.

Flow Cytometry analysis of SiHa cells using anti-ARF6 antibody. Overlay histogram showing SiHa cells (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-ARF6 Antibody (1 µg/1x10^6 cells) for 30 min at 20°C. DyLight®488 conjugated goat anti-rabbit IgG (5-10 µg/1x10^6 cells) was used as secondary antibody for 30 minutes at 20°C. Isotype control antibody (Green line) was rabbit IgG (1 µg/1x10^6) used under the same conditions. Unlabelled sample without incubation with primary antibody and secondary antibody (Red line) was used as a blank control.

IF analysis of ARF6 using anti-ARF6 antibody. ARF6 was detected in immunocytochemical section of A431 cells. 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 2 µg/mL rabbit anti-ARF6 Antibody overnight at 4°C. DyLight®594 Conjugated Goat Anti-Rabbit IgG was used as secondary antibody at 1:100 dilution and incubated for 30 minutes at 37°C. The section was counterstained with DAPI. Visualize using a fluorescence microscope and filter sets appropriate for the label used.

Western blot analysis of ARF6 using anti-ARF6 antibody. Electrophoresis was performed on a 5-20% SDS-PAGE gel at 70V (Stacking gel) / 90V (Resolving gel) for 2-3 hours. The sample well of each lane was loaded with 50 ug of sample under reducing conditions. Lane 1: rat kidney tissue lysates, Lane 2: mouse kidney tissue lysates, Lane 3: MCF-7 whole cell lysates, After Electrophoresis, proteins were transferred to a Nitrocellulose membrane at 150mA 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-ARF6 antigen affinity purified polyclonal antibody at 0.5 µg/mL overnight at 4°C, 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 1:10000 for 1.5 hour at RT. The signal is developed using an Enhanced Chemiluminescent detection (ECL) kit with Tanon 5200 system. A specific band was detected for ARF6 at approximately 20 KD. The expected band size for ARF6 is at 20D.