重组Anti-Nrf2抗体[EP1808Y] - BSA and Azide free (ab180845)
Key features and details
- Produced recombinantly (animal-free) for high batch-to-batch consistency and long term security of supply
- Rabbit monoclonal [EP1808Y] to Nrf2 - BSA and Azide free
- Suitable for: Flow Cyt (Intra), ICC/IF, WB
- Knockout validated
- Reacts with: Human
Related conjugates and formulations
概述
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产品名称
Anti-Nrf2抗体[EP1808Y] - BSA and Azide free
参阅全部 Nrf2 一抗 -
描述
兔单克隆抗体[EP1808Y] to Nrf2 - BSA and Azide free -
宿主
Rabbit -
特异性
The expression of Nrf2 is stimulated by oxidative stress, electrophiles and chemical activators (PMID: 25761198, PMID: 27638861 and PMID: 28587109). Nrf2 antibody (ab62352) detects no signal in most untreated samples in WB. Stimuli treated samples are recommended. We do not recommend using this product in western blot with tissue lysates, however some customers have used this antibody successfully using concentrated samples (see submitted abreviews).
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经测试应用
适用于: Flow Cyt (Intra), ICC/IF, WBmore details
不适用于: ChIP,IHC-P or IP -
种属反应性
与反应: Human -
免疫原
Synthetic peptide. This information is proprietary to Abcam and/or its suppliers.
(Peptide available asab167152) -
阳性对照
- WB: MG-132 treated HeLa whole cell lysate, THP-1, MG-132 treated HepG2 whole cell lysate, MG-132 treated HCT-116 and A549 whole cell lysate. ICC/IF: HepG2 and HeLa cells. Flow Cyt (intra): HeLa cells.
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常规说明
ab180845 is the carrier-free version of ab62352.
Our carrier-free antibodies are typically supplied in a PBS-only formulation, purified and free of BSA, sodium azide and glycerol. The carrier-free buffer and high concentration allow for increased conjugation efficiency.
This conjugation-ready format is designed for use with fluorochromes, metal isotopes, oligonucleotides, and enzymes, which makes them ideal for antibody labelling, functional and cell-based assays, flow-based assays (e.g. mass cytometry) and Multiplex Imaging applications.
Use our conjugation kits for antibody conjugates that are ready-to-use in as little as 20 minutes with <1 minute hands-on-time and 100% antibody recovery: available for fluorescent dyes, HRP, biotin and gold.
This product is compatible with the Maxpar® Antibody Labeling Kit from Fluidigm, without the need for antibody preparation. Maxpar® is a trademark of Fluidigm Canada Inc.
This product is a recombinant monoclonal antibody, which offers several advantages including:
- - High batch-to-batch consistency and reproducibility
- - Improved sensitivity and specificity
- - Long-term security of supply
- - Animal-free production
Our RabMAb® technology is a patented hybridoma-based technology for making rabbit monoclonal antibodies. For details on our patents, please refer to RabMAb® patents.
性能
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形式
Liquid -
存放说明
Shipped at 4°C. Store at +4°C. Do Not Freeze. -
存储溶液
Constituent: 100% PBS -
无载体
是 -
Concentration information loading...
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纯度
Protein A purified -
克隆
单克隆 -
克隆编号
EP1808Y -
同种型
IgG -
研究领域
相关产品
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Alternative Versions
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Conjugation kits
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Isotype control
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Recombinant Protein
应用
The Abpromise guarantee
Abpromise™承诺保证使用ab180845于以下的经测试应用
“应用说明”部分 下显示的仅为推荐的起始稀释度;实际最佳的稀释度/浓度应由使用者检定。
应用 | Ab评论 | 说明 |
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Flow Cyt (Intra) |
Use at an assay dependent concentration.
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ICC/IF |
Use at an assay dependent concentration.
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WB |
Use at an assay dependent concentration. Predicted molecular weight: 68 kDa.
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说明 |
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Flow Cyt (Intra)
Use at an assay dependent concentration. |
ICC/IF
Use at an assay dependent concentration. |
WB
Use at an assay dependent concentration. Predicted molecular weight: 68 kDa. |
靶标
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功能
Transcription activator that binds to antioxidant response (ARE) elements in the promoter regions of target genes. Important for the coordinated up-regulation of genes in response to oxidative stress. May be involved in the transcriptional activation of genes of the beta-globin cluster by mediating enhancer activity of hypersensitive site 2 of the beta-globin locus control region. -
组织特异性
Widely expressed. Highest expression in adult muscle, kidney, lung, liver and in fetal muscle. -
序列相似性
Belongs to the bZIP family. CNC subfamily.
Contains 1 bZIP domain. -
结构域
Acidic activation domain in the N-terminus, and DNA binding domain in the C-terminus. -
翻译后修饰
Phosphorylation of Ser-40 by PKC in response to oxidative stress dissociates NFE2L2 from its cytoplasmic inhibitor KEAP1, promoting its translocation into the nucleus. -
细胞定位
Cytoplasm > cytosol. Nucleus. Cytosolic under unstressed conditions, translocates into the nucleus upon induction by electrophilic agents. - Information by UniProt
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数据库链接
- Entrez Gene: 4780 Human
- Omim: 600492 Human
- SwissProt: Q16236 Human
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别名
- erythroid derived 2 antibody
- HEBP1 antibody
- like 2 antibody
see all
图片
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This data was developed using ab62352, the same antibody clone in a different buffer formulation.
ab62352 staining Nrf2 in untreated wild type A549 cells (left panel), treated wild type A549 cells (middle panel) and untreated NFE2L2 knockout A549 cells (right panel). Cells were treated with 2µM of MG-132 for 18 hours (ab141003). The cells were fixed with 4% paraformaldehyde (10 min) then permeabilized with 0.1% Triton X-100 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 with ab62352 at 0.2 µg/ml concentration and ab7291 (Mouse monoclonal to alpha Tubulin) at 1/1000 dilution overnight at 4°C followed by a further incubation at room temperature for 1h with a goat secondary antibody to rabbit IgG (Alexa Fluor® 488) (ab150081) at 2µg/ml (shown in green) and a goat secondary antibody to mouse IgG (Alexa Fluor® 594) (ab150120) at 2µg/ml (shown in magenta). Nuclear DNA was labelled in blue with DAPI.
Image was taken with a confocal microscope (Leica-Microsystems TCS SP8). -
All lanes : Anti-Nrf2 antibody [EP1808Y] (ab62352) at 1/500 dilution
Lane 1 : Wild-type HeLa control MG132 (0 uM, 18 h) cell lysate
Lane 2 : Wild-type HeLa treated MG132 (2 uM, 18 h) cell lysate
Lane 3 : NFE2L2 knockout HeLa control MG132 (0 uM, 18 h) cell lysate
Lane 4 : NFE2L2 knockout HeLa treated MG132 (2 uM, 18 h) cell lysate
Lane 5 : HCT 116 cell lysate
Lysates/proteins at 20 µg per lane.
Predicted band size: 68 kDa
Observed band size: 85 kDa why is the actual band size different from the predicted?False colour image of Western blot: Anti-Nrf2 antibody [EP1808Y] - ChIP Grade staining at 1/500 dilution, shown in green; Mouse anti-GAPDH antibody [6C5] (ab8245) loading control staining at 1/20000 dilution, shown in red. In Western blot, ab62352 was shown to bind specifically to Nrf2. A band was observed at 85 kDa in wild-type HeLa cell lysates with no signal observed at this size in NFE2L2 CRISPR-Cas9 edited cell line ab262507 (CRISPR-Cas9 edited cell lysate ab263934). The band observed in the CRISPR-Cas9 edited lysate lane below 85 kDa is likely to represent a truncated form of Nrf2. This has not been investigated further and the functional properties of the gene product have not been determined. To generate this image, wild-type and NFE2L2 CRISPR-Cas9 edited HeLa cell lysates were analysed. First, samples were run on an SDS-PAGE gel then transferred onto a nitrocellulose membrane. Membranes were blocked in 3 % milk in TBS-0.1 % Tween® 20 (TBS-T) before incubation with primary antibodies overnight at 4 °C. Blots were washed four times in TBS-T, incubated with secondary antibodies for 1 h at room temperature, washed again four times then imaged. Secondary antibodies used were Goat anti-Rabbit IgG H&L (IRDye® 800CW) preabsorbed (ab216773) and Goat anti-Mouse IgG H&L (IRDye® 680RD) preabsorbed (ab216776) at 1/20000 dilution.
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All lanes : Anti-Nrf2 antibody [EP1808Y] (ab62352) at 1/1000 dilution
Lane 1 : Wild-type A549 Vehicle control MG132 (0 uM, 18 h) cell lysate
Lane 2 : Wild-type A549 Treated MG132 (2 uM, 18 h) cell lysate
Lane 3 : NFE2L2 [21] knockout A549 Vehicle control MG132 (0 uM, 18 h) cell lysate
Lane 4 : NFE2L2 [21] knockout A549 Treated MG132 (2 uM, 18 h) cell lysate
Lysates/proteins at 20 µg per lane.
Performed under reducing conditions.
Predicted band size: 68 kDa
Observed band size: 85-90 kDa why is the actual band size different from the predicted?This data was developed using ab62352, the same antibody clone in a different buffer formulation.
False colour image of Western blot: Anti-Nrf2 antibody [EP1808Y] - ChIP Grade staining at 1/1000 dilution, shown in green; Mouse anti-GAPDH antibody [6C5] (ab8245) loading control staining at 1/20000 dilution, shown in red. In Western blot, ab62352 was shown to bind specifically to Nrf2. A band was observed at 85-90 kDa in wild-type A549 cell lysates with no signal observed at this size in NFE2L2 knockout cell line ab285359 (knockout cell lysate ab289682). To generate this image, wild-type and NFE2L2 knockout A549 cell lysates were analysed. Please note that MG132 treatment does not affect expression levels of Nrf2. First, samples were run on an SDS-PAGE gel then transferred onto a nitrocellulose membrane. Membranes were blocked in 3 % milk in TBS-0.1 % Tween® 20 (TBS-T) before incubation with primary antibodies overnight at 4 °C. Blots were washed four times in TBS-T, incubated with secondary antibodies for 1 h at room temperature, washed again four times then imaged. Secondary antibodies used were Goat anti-Rabbit IgG H&L 800CW and Goat anti-Mouse IgG H&L 680RD at 1/20000 dilution.
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All lanes : Anti-Nrf2 antibody [EP1808Y] (ab62352) at 1/1000 dilution
Lane 1 : HeLa (Human cervix adenocarcinoma epithelial cell) whole cell lysate at 20 µg
Lane 2 : HeLa (Human cervix adenocarcinoma epithelial cell) whole cell lysate treated with MG-132 2uM for 18h
Secondary
All lanes : Goat Anti-Rabbit IgG H&L (HRP) (ab97051) at 1/20000 dilution
Predicted band size: 68 kDa
Observed band size: 955-110 kDa why is the actual band size different from the predicted?
Exposure time: 10 secondsThis data was developed using ab62352, the same antibody clone in a different buffer formulation.
Blocking and diluting buffer and concnetration: 5% NFDM/TBST.
ab181602 was used as GAPDH loading control.
The two bands are different isoforms of Nrf2, the molecular weight observed is consistent with what has been described in the literature: PMID: 17512459, PMID: 22703241.
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Clone EP1808Y (ab180845) has been successfully conjugated by Abcam. This image was generated using Anti-Nrf2 antibody [EP1808Y] (Alexa Fluor® 647). Please refer to ab194985 for protocol details.
ab194985 staining Nrf2 in HeLa cells. The cells were fixed with 100% methanol (5 min), permeabilized in 0.1% Triton X-100 for 5 minutes and then blocked in 1% BSA/10% normal goat serum/0.3M glycine in 0.1%PBS-Tween for 1h. The cells were then incubated with ab194985 at a working dilution 1/100 (shown in red) and ab195887, Mouse monoclonal [DM1A] to alpha Tubulin (Alexa Fluor® 488, shown in green) at 2µg/ml overnight at +4°C. Nuclear DNA was labelled in blue with DAPI.
This product gave a positive signal in 4% formaldehyde (10 min) fixed HeLa cells under the same testing conditions.
Image was taken with a confocal microscope (Leica-Microsystems, TCS SP8).
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Clone EP1808Y (ab180845) has been successfully conjugated by Abcam. This image was generated using Anti-Nrf2 antibody [EP1808Y] (PE). Please refer to ab223926 for protocol details.
Overlay histogram showing HeLa (human epithelial cell line from cervix adenocarcinoma) cells stained with ab223926 (red line). The cells were fixed with 4% formaldehyde (10 minutes) and then permeabilized with 0.1% PBS-Triton X-100 for 15 minutes. The cells were then incubated in 1x PBS / 10% normal goat serum to block non-specific protein-protein interactions followed by the antibody (ab223926, 1/5000 dilution) for 30 minutes at 22°C.
Isotype control antibody (black line) was Rabbit IgG (monoclonal) Phycoerythrin (ab209478) used at the same concentration and conditions as the primary antibody. Unlabeled sample (blue line) was also used as a control.
Acquisition of >5,000 events were collected using a 50 mW Yellow/Green laser (561nm) and 586/15 bandpass filter.
This antibody gave a positive signal in HeLa cells fixed with 80% methanol (5 minutes)/permeabilized with 0.1% PBS-Triton X-100 for 15 minutes used under the same conditions.
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Clone EP1808Y (ab180845) has been successfully conjugated by Abcam. This image was generated using Anti-Nrf2 antibody [EP1808Y] (Alexa Fluor® 488). Please refer to ab194984 for protocol details.
ab194984 staining Nrf2 in HeLa cells. The cells were fixed with 4% formaldehyde (10 min), permeabilized in 0.1% Triton X-100 for 5 minutes and then blocked in 1% BSA/10% normal goat serum/0.3M glycine in 0.1%PBS-Tween for 1h. The cells were then incubated with ab194984 at a working dilution of 1/100 (shown in green) and ab195889, Mouse monoclonal [DM1A] to alpha Tubulin (Alexa Fluor® 594, shown in red) at 2µg/ml overnight at +4°C. Nuclear DNA was labelled in blue with DAPI.
Image was taken with a confocal microscope (Leica-Microsystems, TCS SP8).
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Immunocytochemistry/Immunofluorescence analysis of HepG2 cells labelling Nrf2 with purified ab62352 at 1/500. Cells were fixed with 4% paraformaldehyde and permeabilized with 0.1% Triton X-100. ab150077, an Alexa Fluor® 488-conjugated goat anti-rabbit IgG (1/500) was used as the secondary antibody. The cells were co-stained with ab195889, an Alexa Fluor® 594-conjugated mouse anti-alpha tubulin antibody (1/200). Nuclei counterstained with DAPI (blue).
Secondary antibody only control: PBS was used instead of the primary antibody as the negative control.
This data was developed using the same antibody clone in a different buffer formulation containing PBS, BSA, glycerol, and sodium azide (ab62352).
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Immunocytochemistry/Immunofluorescence analysis of HeLa cells labelling Nrf2 with purified ab62352 at 1/500. Cells were fixed with 4% paraformaldehyde and permeabilized with 0.1% Triton X-100. Goat Anti-Rabbit IgG H&L (Alexa Fluor® 488) (ab150077) secondary antibody (1/1000) was used as the secondary antibody. Cells were counterstained with ab195889, anti-alpha Tubulin antibody [DM1A] - Microtubule Marker (Alexa Fluor® 594). DAPI was used to stain the nuclei blue.
Secondary antibody only control: PBS was used instead of the primary antibody as the negative control.
This data was developed using the same antibody clone in a different buffer formulation containing PBS, BSA, glycerol, and sodium azide (ab62352).
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Intracellular Flow Cytometry analysis of HeLa cells labelling Nrf2 with purified ab62352 at a dilution of 1/60 (red). Cells were fixed with 4% paraformaldehyde. Goat Anti-Rabbit IgG H&L (Alexa Fluor® 488) (ab150077) secondary antibody (1/2000) was used as the secondary antibody. Black - Isotype control, rabbit monoclonal IgG (ab172730). Blue - Unlabelled control, cells without incubation with primary and secondary antibodies.
This data was developed using the same antibody clone in a different buffer formulation containing PBS, BSA, glycerol, and sodium azide (ab62352).
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Intracellular Flow Cytometry analysis of HeLa (human cervix adenocarcinoma) cells labelling Nrf2 with ab62352 at 1/40 (red). Cells were fixed with 4% paraformaldehyde and permeabilised with 90% methanol. An Alexa Fluor®488-conjugated goat anti-rabbit IgG (1/2000) was used as the secondary antibody. Black - Isotype control, rabbit monoclonal IgG. Blue - Unlabelled control, cells without incubation with primary and secondary antibodies.
This data was developed using the same antibody clone in a different buffer formulation containing PBS, BSA, glycerol, and sodium azide (ab62352).
实验方案
数据表及文件
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Datasheet download
Certificate of Compliance
文献 (13)
ab180845 被引用在 13 文献中.
- Chhunchha B et al. Switching of Redox Signaling by Prdx6 Expression Decides Cellular Fate by Hormetic Phenomena Involving Nrf2 and Reactive Oxygen Species. Cells 11:N/A (2022). PubMed: 35455944
- Bu X et al. CD147 confers temozolomide resistance of glioma cells via the regulation of β-TrCP/Nrf2 pathway. Int J Biol Sci 17:3013-3023 (2021). PubMed: 34421346
- Chhunchha B et al. Sulforaphane-Induced Klf9/Prdx6 Axis Acts as a Molecular Switch to Control Redox Signaling and Determines Fate of Cells. Cells 8:N/A (2019). PubMed: 31569690
- Hu T et al. Clinicopathologic significance of CXCR4 and Nrf2 in colorectal cancer. J Biomed Res 27:283-90 (2013). IHC ; Human . PubMed: 23885267
- Ma J et al. PALB2 interacts with KEAP1 to promote NRF2 nuclear accumulation and function. Mol Cell Biol 32:1506-17 (2012). WB . PubMed: 22331464