ATP1A2 Antibody / ATPase subunit alpha-2 (FY13046)

Immunohistochemical staining of ATP1A2 using anti-ATP1A2 antibody. ATP1A2 was detected in a paraffin-embedded section of human breast 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-ATP1A2 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 ATP1A2 using anti-ATP1A2 antibody. Lane 1: rat brain tissue lysates (100oC boiled for 5 minutes); Lane 2: rat brain tissue lysates (37oC incubated for 10 minutes); Lane 3: mouse brain tissue lysates (100oC boiled for 5 minutes); Lane 4: mouse brain tissue lysates (37oC incubated for 10 minutes). 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-ATP1A2 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. Western blot of ATP1A2 in rat and mouse brain lysates prepared at two denaturation temperatures. A strong band at ~112 kDa corresponds to ATP1A2. In samples incubated at 37°C, an additional higher-molecular-weight band is present, consistent with partially denatured alpha2 complexes (alpha2 with the glycosylated beta subunit and/or oligomeric species). Heating at 100°C for 5 min disrupts these membrane protein assemblies and reduces disulfide bonds, eliminating the upper complex band and yielding predominantly the ATP1A2 monomer.

Immunohistochemical staining of ATP1A2 using anti-ATP1A2 antibody. ATP1A2 was detected in a paraffin-embedded section of mouse cerebellum 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-ATP1A2 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 ATP1A2 using anti-ATP1A2 antibody. ATP1A2 was detected in a paraffin-embedded section of rat cerebellum 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-ATP1A2 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.