Anti-Nrf2 antibody [EPR30100-592]

• ICC/IF

Supplier Data

Immunocytochemistry/ Immunofluorescence - Anti-Nrf2 antibody [EPR30100-592] (AB325240)

Immunofluorescent analysis of 100% methanol-fixed, 0.1% TritonX-100 permeabilized HeLa (human cervical adenocarcinoma epithelial cell) HeLa cells treated with MG-132 (10 μM) for 4 hr cells labelling Nrf2 with ab325240 at 1/50 (10.22 ug/ml) dilution, followed by ab150081 Goat Anti-Rabbit IgG H&L (Alexa Fluor® 488) preadsorbed antibody at 1/1000 2ug/ml dilution (Green).

Confocal image showing increased nuclear staining in HeLa cells (shown in green) treated with MG-132 (10 μM) for 4 hr. The counterstain was observed in magenta. Nuclear DNA was labelled with DAPI (shown in blue). Image was taken with a confocal microscope (Leica-Microsystems, TCS SP8).

ab7291 Anti-alpha Tubulin mouse monoclonal antibody was used to counterstain tubulin at 1/1000 1ug/ml dilution (Magenta). The Nuclear counterstain was DAPI (Blue).

Secondary antibody only control : Secondary antibody is ab150081 Goat Anti-Rabbit IgG H&L (Alexa Fluor® 488) preadsorbed at 1/1000 2ug/ml dilution.

• Flow Cyt (Intra)

Supplier Data

Flow Cytometry (Intracellular) - Anti-Nrf2 antibody [EPR30100-592] (AB325240)

Flow cytometric analysis of 4% paraformaldehyde fixed 90% methanol permeabilized HeLa (human cervical adenocarcinoma epithelial cell) treated with 10uM MG-132 for 4 hours (Green) and Untreated control (Magenta) cells labelling Nrf2 with ab325240 at 1/500 dilution (0.1ug) / Green and Red compared with a Rabbit monoclonal IgG (ab172730) (Black) and Grey isotype control and an unlabelled control (cells without incubation with primary antibody and secondary antibody).

Goat Anti-Rabbit IgG (Alexa Fluor® 488, ab150081) at 1/5000 dilution was used as the secondary antibody.

• ChIP-seq

Lab

ChIP-sequencing - Anti-Nrf2 antibody [EPR30100-592] (AB325240)

Chromatin was prepared from HeLa cells. Cells were fixed with 1% formaldehyde for 10 minutes. ChIP was performed with 5 x 10⁶ cells and 4 µg of ab325240 [EPR30100-592]. ChIP DNA was sequenced on the Illumina NovaSeq 6000 to a depth of 30 million reads. The Input control is also shown.

• ChIP-seq

Lab

ChIP-sequencing - Anti-Nrf2 antibody [EPR30100-592] (AB325240)

Chromatin was prepared from HeLa cells. Cells were fixed with 1% formaldehyde for 10 minutes. ChIP was performed with 5 x 10⁶ cells and 4 µg of ab325240 [EPR30100-592]. ChIP DNA was sequenced on the Illumina NovaSeq 6000 to a depth of 30 million reads. The Input control is also shown.

• ChIP-seq

Lab

ChIP-sequencing - Anti-Nrf2 antibody [EPR30100-592] (AB325240)

Chromatin was prepared from HeLa cells. Cells were fixed with 1% formaldehyde for 10 minutes. ChIP was performed with 5 x 10⁶ cells and 4 µg of ab325240 [EPR30100-592]. ChIP DNA was sequenced on the Illumina NovaSeq 6000 to a depth of 30 million reads. The Input control is also shown.

• IP

Supplier Data

Immunoprecipitation - Anti-Nrf2 antibody [EPR30100-592] (AB325240)

Nrf2 was immunoprecipitated from 0.35 mg HeLa (human cervical adenocarcinoma epithelial cell) whole cell lysate with ab325240 at 1/30 dilution (2ug in 0.35mg lysates). Western blot was performed on the immunoprecipitate using ab325240 at 1/1000 dilution.

Blocking and dilution buffer and concentration : 5% NFDM/TBST

To minimize protein degradation, cells were lysed immediately after harvest and then applied for Immunoprecipitation as soon as possible.

All lanes:

Immunoprecipitation - Anti-Nrf2 antibody [EPR30100-592] (ab325240) at 1/1000 dilution

Lane 1:

HeLa (human cervical adenocarcinoma epithelial cell) whole cell lysate at 25 µg

Lane 2:

HeLa (human cervical adenocarcinoma epithelial cell) whole cell lysate

Lane 3:

Rabbit monoclonal IgG (<a href='/ko/products/primary-antibodies/rabbit-igg-monoclonal-epr25a-isotype-control-ab172730'>ab172730</a>) instead of ab325240 in HeLa whole cell lysate

Secondary

All lanes:

Immunoprecipitation - VeriBlot for IP Detection Reagent (HRP) (<a href='/ko/products/reagents/veriblot-for-ip-detection-reagent-hrp-ab131366'>ab131366</a>) at 1/5000 dilution

Observed band size: 105 kDa

false

Exposure time: 50s

• ICC/IF

Supplier Data

Immunocytochemistry/ Immunofluorescence - Anti-Nrf2 antibody [EPR30100-592] (AB325240)

Immunofluorescent analysis of 100% methanol-fixed, 0.1% TritonX-100 permeabilized Hepa1-6 (mouse hepatoma epithelial cell) Hepa1-6 cells (shown in green) treated with Diethylmaleate (100 μM) for 4 hr cells labelling Nrf2 with ab325240 at 1/50 (10.22 ug/ml) dilution, followed by ab150081 Goat Anti-Rabbit IgG H&L (Alexa Fluor® 488) preadsorbed antibody at 1/1000 2ug/ml dilution (Green).

Confocal image showing increased nuclear staining in Hepa1-6 cells (shown in green) treated with Diethylmaleate (100 μM) for 4 hr. The counterstain was observed in magenta. Nuclear DNA was labelled with DAPI (shown in blue). Image was taken with a confocal microscope (Leica-Microsystems, TCS SP8).

ab7291 Anti-alpha Tubulin mouse monoclonal antibody was used to counterstain tubulin at 1/1000 1ug/ml dilution (Magenta). The Nuclear counterstain was DAPI (Blue).

Secondary antibody only control : Secondary antibody is ab150081 Goat Anti-Rabbit IgG H&L (Alexa Fluor® 488) preadsorbed at 1/1000 2ug/ml dilution.

• ICC/IF

Supplier Data

Immunocytochemistry/ Immunofluorescence - Anti-Nrf2 antibody [EPR30100-592] (AB325240)

Immunofluorescent analysis of 100% methanol-fixed, 0.1% TritonX-100 permeabilized C6 (rat glial tumor glial cell) C6 cells treated with MG-132 (10 μM) for 16 hr cells labelling Nrf2 with ab325240 at 1/50 (10.22 ug/ml) dilution, followed by ab150081 Goat Anti-Rabbit IgG H&L (Alexa Fluor® 488) preadsorbed antibody at 1/1000 2ug/ml dilution (Green).

Confocal image showing increased nuclear staining in C6 cells (shown in green) treated with MG-132 (10 μM) for 16 hr. The counterstain was observed in magenta. Nuclear DNA was labelled with DAPI (shown in blue). Image was taken with a confocal microscope (Leica-Microsystems, TCS SP8).

ab7291 Anti-alpha Tubulin mouse monoclonal antibody was used to counterstain tubulin at 1/1000 1ug/ml dilution (Magenta). The Nuclear counterstain was DAPI (Blue).

Secondary antibody only control : Secondary antibody is ab150081 Goat Anti-Rabbit IgG H&L (Alexa Fluor® 488) preadsorbed at 1/1000 2ug/ml dilution.

• Flow Cyt (Intra)

Supplier Data

Flow Cytometry (Intracellular) - Anti-Nrf2 antibody [EPR30100-592] (AB325240)

Flow cytometric analysis of 4% paraformaldehyde fixed 90% methanol permeabilized Hepa1-6 (mouse hepatoma epithelial cell) treated with 10uM MG-132 for 4 hours (Green) and Untreated control (Magenta) cells labelling Nrf2 with ab325240 at 1/500 dilution (0.1ug) / Green and Red compared with a Rabbit monoclonal IgG (ab172730) (Black) and Grey isotype control and an unlabelled control (cells without incubation with primary antibody and secondary antibody).

Goat Anti-Rabbit IgG (Alexa Fluor® 488, ab150081) at 1/5000 dilution was used as the secondary antibody.

• IP

Supplier Data

Immunoprecipitation - Anti-Nrf2 antibody [EPR30100-592] (AB325240)

Nrf2 was immunoprecipitated from 0.35 mg C6 (rat glial tumor glial cell) whole cell lysate with ab325240 at 1/30 dilution (2ug in 0.35mg lysates). Western blot was performed on the immunoprecipitate using ab325240 at 1/1000 dilution.

Blocking and dilution buffer and concentration : 5% NFDM/TBST

To minimize protein degradation, cells were lysed immediately after harvest and then applied for Immunoprecipitation as soon as possible.

All lanes:

Immunoprecipitation - Anti-Nrf2 antibody [EPR30100-592] (ab325240) at 1/30 dilution

Lane 1:

C6 (rat glial tumor glial cell) whole cell lysate at 15 µg

Lane 2:

C6 (rat glial tumor glial cell) whole cell lysate

Lane 3:

Rabbit monoclonal IgG (<a href='/ko/products/primary-antibodies/rabbit-igg-monoclonal-epr25a-isotype-control-ab172730'>ab172730</a>) instead of ab325240 in C6 whole cell lysate

Secondary

All lanes:

Immunoprecipitation - VeriBlot for IP Detection Reagent (HRP) (<a href='/ko/products/reagents/veriblot-for-ip-detection-reagent-hrp-ab131366'>ab131366</a>) at 1/5000 dilution

Observed band size: 105 kDa

false

Exposure time: 145s

• ChIP-seq

Lab

ChIP-sequencing - Anti-Nrf2 antibody [EPR30100-592] (AB325240)

Chromatin was prepared from Hepa1-6 cells. Cells were fixed with 1% formaldehyde for 10 minutes. ChIP was performed with 5 x 10⁶ cells and 4 µg of ab325240 [EPR30100-592]. ChIP DNA was sequenced on the Illumina NovaSeq 6000 to a depth of 30 million reads. The Input control is also shown.

• ChIP-seq

Lab

ChIP-sequencing - Anti-Nrf2 antibody [EPR30100-592] (AB325240)

Chromatin was prepared from Hepa1-6 cells. Cells were fixed with 1% formaldehyde for 10 minutes. ChIP was performed with 5 x 10⁶ cells and 4 µg of ab325240 [EPR30100-592]. ChIP DNA was sequenced on the Illumina NovaSeq 6000 to a depth of 30 million reads. The Input control is also shown.

• IP

Supplier Data

Immunoprecipitation - Anti-Nrf2 antibody [EPR30100-592] (AB325240)

Nrf2 was immunoprecipitated from 0.35 mg Hepa1-6 (mouse hepatoma epithelial cell) whole cell lysate with ab325240 at 1/30 dilution (2ug in 0.35mg lysates). Western blot was performed on the immunoprecipitate using ab325240 at 1/1000 dilution.

Blocking and dilution buffer and concentration : 5% NFDM/TBST

To minimize protein degradation, cells were lysed immediately after harvest and then applied for Immunoprecipitation as soon as possible.

All lanes:

Immunoprecipitation - Anti-Nrf2 antibody [EPR30100-592] (ab325240) at 1/1000 dilution

Lane 1:

Hepa1-6 (mouse hepatoma epithelial cell) whole cell lysate at 18 µg

Lane 2:

Hepa1-6 (mouse hepatoma epithelial cell) whole cell lysate

Lane 3:

Rabbit monoclonal IgG (<a href='/ko/products/primary-antibodies/rabbit-igg-monoclonal-epr25a-isotype-control-ab172730'>ab172730</a>) instead of ab325240 in Hepa1-6 whole cell lysate

Secondary

All lanes:

Immunoprecipitation - VeriBlot for IP Detection Reagent (HRP) (<a href='/ko/products/reagents/veriblot-for-ip-detection-reagent-hrp-ab131366'>ab131366</a>) at 1/5000 dilution

Observed band size: 105 kDa

false

Exposure time: 180s

• ChIP-seq

Lab

ChIP-sequencing - Anti-Nrf2 antibody [EPR30100-592] (AB325240)

Chromatin was prepared from Hepa1-6 cells. Cells were fixed with 1% formaldehyde for 10 minutes. ChIP was performed with 5 x 10⁶ cells and 4 µg of ab325240 [EPR30100-592]. ChIP DNA was sequenced on the Illumina NovaSeq 6000 to a depth of 30 million reads. The Input control is also shown.

• WB

Supplier Data

Western blot - Anti-Nrf2 antibody [EPR30100-592] (AB325240)

Blocking and diluting buffer and concentration : 5% NFDM/TBST.

The expression profile/molecular weight observed is consistent with what has been described in the literature (PMID : 3600148; PMID : 22932898; PMID : 38495294).

The expression of is upregulated in response to MG-132 treatment (PMID : 22932898).

In Western blot, Anti-GAPDH antibody [EPR16891] - Loading Control (ab181602) staining at 1/200000 dilution.

All lanes:

Western blot - Anti-Nrf2 antibody [EPR30100-592] (ab325240) at 1/1000 dilution

Lane 1:

Untreated U-2 OS (human bone osteosarcoma epithelial cell) whole cell lysate at 20 µg

Lane 2:

U-2 OS treated with 10uM MG-132 for 4 hours whole cell lysate at 20 µg

Lane 3:

Untreated MEF (mouse embryo fibroblast) whole cell lysate at 20 µg

Lane 4:

MEF treated with 20uM MG-132 for 4 hours whole cell lysate at 20 µg

Secondary

All lanes:

Western blot - Goat Anti-Rabbit IgG H&L (HRP) (<a href='/ko/products/secondary-antibodies/goat-rabbit-igg-h-l-hrp-ab97051'>ab97051</a>) at 1/20000 dilution

Observed band size: 105 kDa,36 kDa

false

Exposure time: 15s

• WB

Supplier Data

Western blot - Anti-Nrf2 antibody [EPR30100-592] (AB325240)

Blocking and diluting buffer and concentration : 5% NFDM/TBST.

The expression profile/molecular weight observed is consistent with what has been described in the literature (PMID : 3600148; PMID : 22932898; PMID : 38495294).

To minimize protein degradation, cells were lysed immediately after harvest and then applied to a gel and transfer membrane for Western blotting as soon as possible.

In Western blot, Anti-GAPDH antibody [EPR16891] - Loading Control (ab181602) staining at 1/200000 dilution.

All lanes:

Western blot - Anti-Nrf2 antibody [EPR30100-592] (ab325240) at 1/1000 dilution

Lane 1:

HeLa (human cervical adenocarcinoma epithelial cell) whole cell lysate at 20 µg

Lane 2:

NIH/3T3 (mouse embryonic fibroblast) whole cell lysate at 20 µg

Lane 3:

PC-12 (rat adrenal gland pheochromocytoma cell) whole cell lysate at 20 µg

Secondary

All lanes:

Western blot - Goat Anti-Rabbit IgG H&L (HRP) (<a href='/ko/products/secondary-antibodies/goat-rabbit-igg-h-l-hrp-ab97051'>ab97051</a>) at 1/20000 dilution

Observed band size: 105 kDa,36 kDa

false

Exposure time: 180s

• WB

Supplier Data

Western blot - Anti-Nrf2 antibody [EPR30100-592] (AB325240)

Blocking and diluting buffer and concentration : 5% NFDM/TBST.

The expression profile/molecular weight observed is consistent with what has been described in the literature (PMID : 3600148; PMID : 22932898; PMID : 38495294).

The expression of Nrf2 is upregulated in response to MG-132 treatment (PMID : 22932898; PMID : 38495294).

In Western blot, Anti-GAPDH antibody [EPR16891] - Loading Control (ab181602) staining at 1/200000 dilution.

All lanes:

Western blot - Anti-Nrf2 antibody [EPR30100-592] (ab325240) at 1/1000 dilution

Lane 1:

Untreated HeLa (human cervical adenocarcinoma epithelial cell) whole cell lysate at 20 µg

Lane 2:

HeLa treated with 10&micro;M MG-132 for 4 hours whole cell lysate at 20 µg

Lane 3:

Untreated Hepa1-6 (mouse hepatoma epithelial cell) whole cell lysate at 20 µg

Lane 4:

Hepa1-6 treated with 100&micro;M Diethylmaleate for 4 hours whole cell lysate at 20 µg

Lane 5:

Untreated C6 (rat glial tumor glial cell) whole cell lysate at 20 µg

Lane 6:

C6 (rat glial tumor glial cell) treated with 10&micro;M MG-132 for 16 hours whole cell lysate at 20 µg

Secondary

All lanes:

Western blot - Goat Anti-Rabbit IgG H&L (HRP) (<a href='/ko/products/secondary-antibodies/goat-rabbit-igg-h-l-hrp-ab97051'>ab97051</a>) at 1/20000 dilution

Observed band size: 105 kDa,36 kDa

false

Exposure time: 180s

• WB

Supplier Data

Western blot - Anti-Nrf2 antibody [EPR30100-592] (AB325240)

Blocking and diluting buffer and concentration : 5% NFDM/TBST.

The expression profile/molecular weight observed is consistent with what has been described in the literature (PMID : 3600148; PMID : 22932898; PMID : 38495294).

In Western blot, Anti-GAPDH antibody [EPR16891] - Loading Control (ab181602) staining at 1/200000 dilution.

All lanes:

Western blot - Anti-Nrf2 antibody [EPR30100-592] (ab325240) at 1/1000 dilution

Lane 1:

HeLa (human cervical adenocarcinoma epithelial cell) transfected with scrambled siRNA control whole cell lysate at 20 µg

Lane 2:

HeLa transfected with siRNA specifically targeting Nrf2 whole cell lysate at 20 µg

Secondary

All lanes:

Western blot - Goat Anti-Rabbit IgG H&L (HRP) (<a href='/ko/products/secondary-antibodies/goat-rabbit-igg-h-l-hrp-ab97051'>ab97051</a>) at 1/20000 dilution

Observed band size: 105 kDa,36 kDa

false

Exposure time: 180s

• Dot

Supplier Data

Dot Blot - Anti-Nrf2 antibody [EPR30100-592] (AB325240)

Dot blot analysis of Nrf2 using ab325240 at 1 : 1000 (0.511 ug/ml) followed by a Goat Anti-Rabbit IgG, (H+L), Peroxidase conjugated (ab97051) at 1 : 100,000 dilution.

Lane1 : 293T cells transfected with a mouse Nrf2 expression vector containing a myc-His-tag®, whole cell lysate
Lane2 : 293T cells transfected with a mouse NF2L1 expression vector containing a myc-His-tag®, whole cell lysate
Lane3 : 293T cells transfected with a mouse NF2L3 expression vector containing a myc-His-tag®, whole cell lysate

Exposure time : 180 seconds.

Blocking and diluting buffer and concentration : 5% NFDM/TBST.

This antibody does not cross-react with mouse NF2L1 and NF2L3

In Dot Blot, Anti-6X His tag® antibody [AD1.1.10] (ab15149) staining at 1/1000 dilution.

All lanes:

Dot Blot - Anti-Nrf2 antibody [EPR30100-592] (ab325240) at 1/1000 dilution

Lane 1:

293T cells transfected with a mouse Nrf2 expression vector containing a myc-His-tag®, whole cell lysate

Lane 2:

293T cells transfected with a mouse NF2L1 expression vector containing a myc-His-tag®, whole cell lysate

Lane 3:

293T cells transfected with a mouse NF2L3 expression vector containing a myc-His-tag®, whole cell lysate

Secondary

All lanes:

Dot Blot - Goat Anti-Rabbit IgG H&L (HRP) (<a href='/ko/products/secondary-antibodies/goat-rabbit-igg-h-l-hrp-ab97051'>ab97051</a>) at 1/100000 dilution

false

Exposure time: 180s