Anti-ATP1A3 antibody [XVIF9-G10]

• IHC-P

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Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-ATP1A3 antibody [XVIF9-G10] (AB2826)

Immunohistochemistry was performed on both normal and cancer biopsies of deparaffinized Human prostate carcinoma tissues. To expose target proteins heat induced antigen retrieval was performed using 10mM sodium citrate (pH6.0) buffer microwaved for 8-15 minutes. Following antigen retrieval tissues were blocked in 3% BSA-PBS for 30 minutes at room temperature. Tissues were then probed at 1/50 dilution with ab2826 or without primary antibody (negative control) overnight at 4°C in a humidified chamber. Tissues were washed extensively with PBST and endogenous peroxidase activity was quenched with a peroxidase suppressor. Detection was performed using a biotin-conjugated secondary antibody and SA-HRP followed by colorimetric detection using DAB. Tissues were counterstained with hematoxylin and prepped for mounting.

• ICC/IF

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Immunocytochemistry/ Immunofluorescence - Anti-ATP1A3 antibody [XVIF9-G10] (AB2826)

Immunofluorescent analysis of Sodium/Potassium ATPase alpha-3 using ab2826 shows staining in U251 glioma cells. Sodium/Potassium ATPase alpha-3 staining (green) F-Actin staining with Phalloidin (red) and nuclei with DAPI (blue) is shown. Cells were grown on chamber slides and fixed with formaldehyde prior to staining. Cells were probed without (control) or with or an antibody recognizing Sodium/Potassium ATPase alpha-3 ab2826 at a dilution of 1 : 20 over night at 4 °C washed with PBS and incubated with a DyLight-488 conjugated secondary antibody. Images were taken at 60X magnification.

• Flow Cyt

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Flow Cytometry - Anti-ATP1A3 antibody [XVIF9-G10] (AB2826)

Overlay histogram showing SH-SY5Y cells stained with ab2826 (red line). The cells were fixed with 4% paraformaldehyde and incubated in 1x PBS / 10% normal goat serum / 0.3M glycine to block non-specific protein-protein interactions. The cells were then incubated with the antibody (ab2826, 1μg/1x10⁶ cells) for 30 min at 22°C. The secondary antibody used was DyLight® 488 goat anti-mouse IgG (H+L) (ab96879) at 1/500 dilution for 30 min at 22°C. Isotype control antibody (black line) was mouse IgG1 [ICIGG1] (ab91353, 2μg/1x10⁶ cells) used under the same conditions. Acquisition of >5,000 events was performed.

Please note that Abcam do not have any data for use of this antibody on non-fixed cells. We welcome any customer feedback.

• IHC-P

Unknown

Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-ATP1A3 antibody [XVIF9-G10] (AB2826)

Immunohistochemistry was performed on both normal and cancer biopsies of deparaffinized Human tonsil tissue tissues. To expose target proteins heat induced antigen retrieval was performed using 10mM sodium citrate (pH6.0) buffer microwaved for 8-15 minutes. Following antigen retrieval tissues were blocked in 3% BSA-PBS for 30 minutes at room temperature. Tissues were then probed at 1/200 dilution with ab2826 or without primary antibody (negative control) overnight at 4°C in a humidified chamber. Tissues were washed extensively with PBST and endogenous peroxidase activity was quenched with a peroxidase suppressor. Detection was performed using a biotin-conjugated secondary antibody and SA-HRP followed by colorimetric detection using DAB. Tissues were counterstained with hematoxylin and prepped for mounting.

• ICC/IF

Unknown

Immunocytochemistry/ Immunofluorescence - Anti-ATP1A3 antibody [XVIF9-G10] (AB2826)

Immunofluorescent analysis of Sodium/Potassium ATPase alpha-3 using ab2826 shows staining in HeLa cells. Sodium/Potassium ATPase alpha-3 staining (green) F-Actin staining with Phalloidin (red) and nuclei with DAPI (blue) is shown. Cells were grown on chamber slides and fixed with formaldehyde prior to staining. Cells were probed without (control) or with or an antibody recognizing Sodium/Potassium ATPase alpha-3 ab2826 at a dilution of 1 : 20 over night at 4°C washed with PBS and incubated with a DyLight-488 conjugated secondary antibody. Images were taken at 60X magnification.

• IHC-P

Unknown

Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-ATP1A3 antibody [XVIF9-G10] (AB2826)

Immunohistochemistry was performed on both normal and cancer biopsies of deparaffinized Human colon tissue tissues. To expose target proteins heat induced antigen retrieval was performed using 10mM sodium citrate (pH6.0) buffer microwaved for 8-15 minutes. Following antigen retrieval tissues were blocked in 3% BSA-PBS for 30 minutes at room temperature. Tissues were then probed at 1/200 dilution with ab2826 or without primary antibody (negative control) overnight at 4°C in a humidified chamber. Tissues were washed extensively with PBST and endogenous peroxidase activity was quenched with a peroxidase suppressor. Detection was performed using a biotin-conjugated secondary antibody and SA-HRP followed by colorimetric detection using DAB. Tissues were counterstained with hematoxylin and prepped for mounting.

• ICC/IF

Unknown

Immunocytochemistry/ Immunofluorescence - Anti-ATP1A3 antibody [XVIF9-G10] (AB2826)

Immunofluorescent analysis of Sodium/Potassium ATPase alpha-3 using ab2826 shows staining in C6 glioma cells. Sodium/Potassium ATPase alpha-3 staining (green) F-Actin staining with Phalloidin (red) and nuclei with DAPI (blue) is shown. Cells were grown on chamber slides and fixed with formaldehyde prior to staining. Cells were probed without (control) or with or an antibody recognizing Sodium/Potassium ATPase alpha-3 ab2826 at a dilution of 1 : 20 over night at 4 °C washed with PBS and incubated with a DyLight-488 conjugated secondary antibody. Images were taken at 60X magnification.

• WB

Supplier Data

Western blot - Anti-ATP1A3 antibody [XVIF9-G10] (AB2826)

Samples were electrophoresed using NuPAGE™ 4-12% Bis-Tris Protein Gel. Resolved proteins were then transferred onto a Nitrocellulose membrane by iBlot® 2 Dry Blotting System. Chemiluminescent detection was performed using Novex® ECL Chemiluminescent Substrate Reagent Kit.

All lanes:

Western blot - Anti-ATP1A3 antibody [XVIF9-G10] (ab2826) at 1 µg/mL

Lane 1:

SH-SY5Y (Human neuroblastoma cell line from bone marrow) whole cell lysate at 30 µg

Lane 2:

IMR-32 (Human brain neuroblast cell line) whole cell lysate at 30 µg

Lane 3:

SK-OV-3 (Human ovarian cancer cell line) whole cell lysate at 30 µg

Lane 4:

HeLa (Human cervix adenocarcinoma epithelial cell) whole cell lysate at 30 µg

Lane 5:

Mouse brain tissue lysate at 30 µg

Secondary

All lanes:

Goat anti-Mouse IgG (H+L) Superclonal™ Recombinant Secondary Antibody, HRP at 1/4000 dilution

Predicted band size: 111 kDa

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