Anti-NQO1 antibody [A180] - BSA and Azide free

• Flow Cyt (Intra)

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Flow Cytometry (Intracellular) - Anti-NQO1 antibody [A180] - BSA and Azide free (AB264434)

Overlay histogram showing HeLa cells stained with ab28947 (red line). The cells were fixed with 80% methanol (5 min) and then permeabilized with 0.1% PBS-Tween for 20 min. The cells were then incubated in 1x PBS / 10% normal goat serum / 0.3M glycine to block non-specific protein-protein interactions followed by the antibody (ab28947 1μg/1x10⁶ cells) for 30 min at 22°C. The secondary antibody used was goat anti-mouse DyLight® 488 (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 >5000 events was performed.

This data was developed using the same antibody clone in a different buffer formulation containing PBS and Azide (ab28947).

• IHC-P

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Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-NQO1 antibody [A180] - BSA and Azide free (AB264434)

IHC image of NQO1 staining in human breast adenocarcinoma formalin fixed paraffin embedded tissue section, performed on a Leica Bond 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 ab28947, 0.1μg/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.

This data was developed using the same antibody clone in a different buffer formulation containing PBS and Azide (ab28947).

• IHC-P

Lab

Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-NQO1 antibody [A180] - BSA and Azide free (AB264434)

IHC image of NQO1 staining in sections of formalin fixed paraffin embedded normal human pancreas* (left) and human pancreas adenocarcinoma* (right), 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 ab28947, 0.1μg/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.

*Tissue obtained from the Human Research Tissue Bank, supported by the NIHR Cambridge Biomedical Research Centre

This data was developed using the same antibody clone in a different buffer formulation containing PBS and Azide (ab28947).

• ICC/IF

Lab

Immunocytochemistry/ Immunofluorescence - Anti-NQO1 antibody [A180] - BSA and Azide free (AB264434)

This data was developed using the same antibody clone in a different buffer formulation containing PBS and sodium azide (ab28947)

ab28947 staining NQO1 in HepG2 cells. The cells were fixed with 100% methanol (5 min), permeabilized with 0.1% PBS-Tween for 5 minutes and then blocked with 1% BSA/10% normal goat serum/0.3M glycine in 0.1%PBS-Tween for 1h. The cells were then incubated overnight at 4°C with ab28947 at 5µg/ml and ab6046, Rabbit polyclonal to beta Tubulin - Loading Control. Cells were then incubated with ab150117, Goat polyclonal Secondary Antibody to Mouse IgG H&L (Alexa Fluor^® 488) preadsorbed at 1/1000 dilution (shown in green) and ab150080, Goat polyclonal Secondary Antibody to Rabbit IgG - H&L (Alexa Fluor^® 594) at 1/1000 dilution (shown in pseudocolour red). Nuclear DNA was labelled with DAPI (shown in blue).

Image was acquired with a high-content analyser (Operetta CLS, Perkin Elmer) and a maximum intensity projection of confocal sections is shown.

• IHC-P

Unknown

Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-NQO1 antibody [A180] - BSA and Azide free (AB264434)

Human breast cancer tissue stained with ab28947 NQO1 antibody.

This data was developed using the same antibody clone in a different buffer formulation containing PBS and Azide (ab28947).

• sELISA

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Sandwich ELISA - Anti-NQO1 antibody [A180] - BSA and Azide free (AB264434)

Sandwich ELISA for the detection of NQO1, using ab28947 (1/500) as the capture antibody and ab34173 (1/1000) for the detection

This data was developed using the same antibody clone in a different buffer formulation containing PBS and Azide (ab28947).

• WB

Lab

Western blot - Anti-NQO1 antibody [A180] - BSA and Azide free (AB264434)

Lane 1 : Wild-type HAP1 whole cell lysate (20 μg)
Lane 2 : NQO1 knockout HAP1 whole cell lysate (20 μg)
Lane 3 : HepG2 whole cell lysate (20 μg)

Lanes 1 - 3 : Merged signal (red and green). Green - ab28947 observed at 31 kDa. Red - loading control, ab176560, observed at 50 kDa.

ab28947 was shown to specifically react with NQO1 in wild-type HAP1 cells as signal was lost in NQO1 knockout cells. Wild-type and NQO1 knockout samples were subjected to SDS-PAGE. ab28947 and ab176560 (Rabbit anti-alpha Tubulin loading control) were incubated overnight at 4°C at 1 μg/ml and 1/20000 dilution respectively. Blots were developed with Goat anti-Mouse IgG H&L (IRDye^® 800CW) preabsorbed ab216772 and Goat anti-Rabbit IgG H&L (IRDye^® 680RD) preabsorbed ab216777 secondary antibodies at 1/20000 dilution for 1 hour at room temperature before imaging.

This data was developed using the same antibody clone in a different buffer formulation containing PBS and Azide (ab28947).

All lanes:

Western blot - Anti-NQO1 antibody [A180] (<a href='/en-us/products/primary-antibodies/nqo1-antibody-a180-ab28947'>ab28947</a>)

Predicted band size: 31 kDa

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