Anti-ATP1B1 antibody [M17-P5-F11]

• Flow Cyt

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Flow Cytometry - Anti-ATP1B1 antibody [M17-P5-F11] (AB2873)

Overlay histogram showing HEK293 cells stained with ab2873 (red line). The cells were fixed with 4% paraformaldehyde (10 min) 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 (ab2873, 1/100 dilution) 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 IgG2a [ICIGG2A] (ab91361, 1μg/1x10⁶ cells) used under the same conditions. Acquisition of >5,000 events was performed. This antibody gave a positive signal in HEK293 cells fixed with 80% methanol (5 min)/permeabilized with 0.1% PBS-Tween for 20 min used under the same conditions.

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

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Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-ATP1B1 antibody [M17-P5-F11] (AB2873)

Immunohistochemistry was performed on both normal and cancer biopsies of deparaffinized Human liver 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 a dilution of 1 : 200 with a mouse monoclonal antibody recognizing Sodium/Potassium ATPase beta ab2873 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.

• IHC-P

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Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-ATP1B1 antibody [M17-P5-F11] (AB2873)

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 a dilution of 1 : 200 with a mouse monoclonal antibody recognizing Sodium/Potassium ATPase beta ab2873 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.

• IHC-P

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Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-ATP1B1 antibody [M17-P5-F11] (AB2873)

IHC image of ab2873 staining in human normal liver formalin fixed paraffin embedded tissue section, performed on a Leica Bond^TM system using the standard protocol F. The section was pre-treated using heat mediated antigen retrieval with sodium citrate buffer (pH6, epitope retrieval solution 1) for 20 mins. The section was then incubated with ab2873, 1µg/ml, for 15 mins at room temperature and detected using an HRP conjugated compact polymer system. DAB was used as the chromogen. The section was then counterstained with haematoxylin and mounted with DPX.

For other IHC staining systems (automated and non-automated) customers should optimize variable parameters such as antigen retrieval conditions, primary antibody concentration and antibody incubation times.

• IHC-P

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Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-ATP1B1 antibody [M17-P5-F11] (AB2873)

Immunohistochemistry was performed on both normal and cancer biopsies of deparaffinized Human colon 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 a dilution of 1 : 200 with a mouse monoclonal antibody recognizing Sodium/Potassium ATPase beta ab2873 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

Supplier Data

Immunocytochemistry/ Immunofluorescence - Anti-ATP1B1 antibody [M17-P5-F11] (AB2873)

Immunocytochemistry/Immunofluorescence analysis of ATP1B1 shows staining in U251 cells. ATP1B1 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 incubated without (control) or with ab2873 (1 : 200) overnight at 4°C, washed with PBS and incubated with a DyLight-488 conjugated goat anti-mouse secondary antibody. Images were taken at 60X magnification.

• ICC/IF

Supplier Data

Immunocytochemistry/ Immunofluorescence - Anti-ATP1B1 antibody [M17-P5-F11] (AB2873)

Immunocytochemistry/Immunofluorescence analysis of ATP1B1 shows staining in MCF-7 cells. ATP1B1 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 incubated without (control) or with ab2873 (1 : 200) overnight at 4°C, washed with PBS and incubated with a DyLight-488 conjugated goat anti-mouse secondary antibody. Images were taken at 60X magnification.

• ICC/IF

Supplier Data

Immunocytochemistry/ Immunofluorescence - Anti-ATP1B1 antibody [M17-P5-F11] (AB2873)

Immunocytochemistry/Immunofluorescence analysis of ATP1B1 shows staining in HeLa cells. ATP1B1 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 incubated without (control) or with ab2873 (1 : 200) overnight at 4°C, washed with PBS and incubated with a DyLight-488 conjugated goat anti-mouse secondary antibody. Images were taken at 60X magnification.

• WB

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Western blot - Anti-ATP1B1 antibody [M17-P5-F11] (AB2873)

Chemiluminescent detection was performed using Pierce ECL Plus Western Blotting Substrate.

All lanes:

Western blot - Anti-ATP1B1 antibody [M17-P5-F11] (ab2873) at 1/5000 dilution

Lane 1:

Human brain lysates at 25 µg

Lane 2:

Human liver lysates at 25 µg

Lane 3:

Human kidney lysates at 25 µg

Lane 4:

Mouse kidney lysates at 25 µg

Secondary

All lanes:

HRP-conjugated secondary antibody

Predicted band size: 35 kDa

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