Anti-Renin receptor/ATP6AP2 Antibody Picoband®

Western blot analysis of Renin receptor/ATP6AP2 using anti-Renin receptor/ATP6AP2 antibody (A02665-3).
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 30 ug of sample under reducing conditions.
Lane 1: human SiHa whole cell lysates,
Lane 2: human HUVEC whole cell lysates,
Lane 3: human U251 whole cell lysates,
Lane 4: rat brain tissue lysates,
Lane 5: rat heart tissue lysates,
Lane 6: mouse brain tissue lysates,
Lane 7: mouse heart 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-Renin receptor/ATP6AP2 antigen affinity purified polyclonal antibody (Catalog # A02665-3) 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 1:5000 for 1.5 hour at RT. The signal is developed using an Enhanced Chemiluminescent detection (ECL) kit (Catalog # EK1002) with Tanon 5200 system. A specific band was detected for Renin receptor/ATP6AP2 at approximately 49 kDa. The expected band size for Renin receptor/ATP6AP2 is at 39 kDa.

IF analysis of Renin receptor/ATP6AP2 using anti-Renin receptor/ATP6AP2 antibody (A02665-3).
Renin receptor/ATP6AP2 was detected in an immunocytochemical section of HELA cells. Enzyme antigen retrieval was performed using IHC enzyme antigen retrieval reagent (AR0022) for 15 mins. The cells were blocked with 10% goat serum. And then incubated with 5 μg/mL rabbit anti-Renin receptor/ATP6AP2 Antibody (A02665-3) overnight at 4°C. DyLight®488 Conjugated Goat Anti-Rabbit IgG (BA1127) was used as secondary antibody at 1:500 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.

Flow Cytometry analysis of SiHa cells using anti-Renin receptor/ATP6AP2 antibody (A02665-3).
Overlay histogram showing SiHa cells stained with A02665-3 (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-Renin receptor/ATP6AP2 Antibody (A02665-3, 1 μg/1x10⁶ cells) for 30 min at 20°C. DyLight®488 conjugated goat anti-rabbit IgG (BA1127, 5-10 μg/1x10⁶ cells) was used as secondary antibody for 30 minutes at 20°C. Isotype control antibody (Green line) was rabbit IgG (1 μg/1x10⁶) used under the same conditions. Unlabelled sample without incubation with primary antibody and secondary antibody (Red line) was used as a blank control.

Immunoprecipitating Renin receptor/ATP6AP2 in U251 whole cell lysate.
Western blot analysis of Renin receptor/ATP6AP2 using anti-Renin receptor/ATP6AP2 antibody (A02665-3);
Lane 1: U251 whole cell lysates (30ug);
Lane 2: Rabbit control IgG instead of anti-Renin receptor/ATP6AP2 antibody in U251 whole cell lysate;
Lane 3: anti-Renin receptor/ATP6AP2 antibody (2μg) + U251 whole cell lysate (500μg).
After electrophoresis, proteins were transferred to a membrane. Then the membrane was incubated with rabbit anti-Renin receptor/ATP6AP2 antigen affinity purified polyclonal antibody (A02665-3) at a dilution of 0.5 μg/mL and probed with a goat anti-rabbit IgG-HRP secondary antibody (Catalog # BA1054). The signal is developed using ECL Plus Western Blotting Substrate (Catalog # AR1196-200). A specific band was detected for Renin receptor/ATP6AP2 at approximately 45 kDa. The expected band size for Renin receptor/ATP6AP2 is at 39 kDa.