Renin receptor Antibody / ATP6AP2 (FY13351)

Immunofluorescent staining of Renin receptor/ATP6AP2 using anti-Renin receptor antibody (green). Renin receptor/ATP6AP2 was detected in an immunocytochemical section of human HeLa 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 5 ug/ml rabbit anti-Renin receptor antibody overnight at 4oC. DyLight 488 Conjugated Goat Anti-Rabbit IgG was used as secondary antibody at 1:500 dilution and incubated for 30 minutes at 37oC. The section was counterstained with DAPI nuclear stain (blue). Visualize using a fluorescence microscope and filter sets appropriate for the label used.

Western blot analysis of Renin receptor/ATP6AP2 using anti-Renin receptor antibody. 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 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. A predominant band is detected at approximately 45-50 kDa in all samples, running above the predicted ~39 kDa mass but consistent with the apparent molecular weight of the heavily glycosylated full length receptor. Doublets likely reflect distinct glycosylation or processing states of ATP6AP2.

Flow Cytometry analysis of SiHa cells using anti-Renin receptor antibody. Overlay histogram showing SiHa 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-Renin receptor 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 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 antibody; Lane 1: U251 whole cell lysates (30ug); Lane 2: Rabbit control IgG instead of anti-Renin receptor antibody in U251 whole cell lysate; Lane 3: anti-Renin receptor antibody (2ug) + U251 whole cell lysate (500ug). After electrophoresis, proteins were transferred to a membrane. Then the membrane was incubated with rabbit anti-Renin receptor 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 predominant band is detected at approximately 45-50 kDa in all samples, running above the predicted ~39 kDa mass but consistent with the apparent molecular weight of the heavily glycosylated full length receptor. Doublets likely reflect distinct glycosylation or processing states of ATP6AP2.