Recombinant Anti-NADPH oxidase 4 antibody [UOTR1B492] (ab109225)
Key features and details
- Produced recombinantly (animal-free) for high batch-to-batch consistency and long term security of supply
- Rabbit monoclonal [UOTR1B492] to NADPH oxidase 4
- Suitable for: Flow Cyt (Intra), ICC/IF, WB, IP, IHC-P
- Reacts with: Human
Related conjugates and formulations
Overview
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Product name
Anti-NADPH oxidase 4 antibody [UOTR1B492]
See all NADPH oxidase 4 primary antibodies -
Description
Rabbit monoclonal [UOTR1B492] to NADPH oxidase 4 -
Host species
Rabbit -
Tested applications
Suitable for: Flow Cyt (Intra), ICC/IF, WB, IP, IHC-Pmore details -
Species reactivity
Reacts with: Human
Predicted to work with: Rat, Dog -
Immunogen
Synthetic peptide. This information is proprietary to Abcam and/or its suppliers.
(Peptide available asab179799) -
Positive control
- Fetal kidney, U87-MG, 293T, and JAR lysates Human kidney tissue
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General notes
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.
We are constantly working hard to ensure we provide our customers with best in class antibodies. As a result of this work we are pleased to now offer this antibody in purified format. We are in the process of updating our datasheets. The purified format is designated 'PUR' on our product labels. If you have any questions regarding this update, please contact our Scientific Support team.
Properties
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Form
Liquid -
Storage instructions
Shipped at 4°C. Store at +4°C short term (1-2 weeks). Store at -20°C. Avoid freeze / thaw cycle. -
Storage buffer
pH: 7.20
Preservative: 0.01% Sodium azide
Constituents: 40% Glycerol, 59% PBS, 0.05% BSA -
Concentration information loading...
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Purity
Protein A purified -
Clonality
Monoclonal -
Clone number
UOTR1B492 -
Isotype
IgG -
Research areas
Associated products
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Alternative Versions
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Compatible Secondaries
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Conjugation kits
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Immunizing Peptide (Blocking)
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Isotype control
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Recombinant Protein
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Related Products
Applications
The Abpromise guarantee
Our Abpromise guarantee covers the use of ab109225 in the following tested applications.
The application notes include recommended starting dilutions; optimal dilutions/concentrations should be determined by the end user.
Application | Abreviews | Notes |
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Flow Cyt (Intra) |
1/280.
ab172730 - Rabbit monoclonal IgG, is suitable for use as an isotype control with this antibody. |
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ICC/IF |
1/200.
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WB | (1) |
1/2000. Detects a band of approximately 63 kDa (predicted molecular weight: 67 kDa).Can be blocked with NADPH oxidase 4 peptide (ab179799).
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IP |
1/30.
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IHC-P | (1) |
1/250 - 1/500. Perform heat mediated antigen retrieval before commencing with IHC staining protocol.
See IHC antigen retrieval protocols. For unpurified, use 1/100 - 1/250. |
Notes |
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Flow Cyt (Intra)
1/280. ab172730 - Rabbit monoclonal IgG, is suitable for use as an isotype control with this antibody. |
ICC/IF
1/200. |
WB
1/2000. Detects a band of approximately 63 kDa (predicted molecular weight: 67 kDa).Can be blocked with NADPH oxidase 4 peptide (ab179799). |
IP
1/30. |
IHC-P
1/250 - 1/500. Perform heat mediated antigen retrieval before commencing with IHC staining protocol. See IHC antigen retrieval protocols. For unpurified, use 1/100 - 1/250. |
Target
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Function
Constitutive NADPH oxidase which generates superoxide intracellularly upon formation of a complex with CYBA/p22phox. Regulates signaling cascades probably through phosphatases inhibition. May function as an oxygen sensor regulating the KCNK3/TASK-1 potassium channel and HIF1A activity. May regulate insulin signaling cascade. May play a role in apoptosis, bone resorption and lipolysaccharide-mediated activation of NFKB. May produce superoxide in the nucleus and play a role in regulating gene expression upon cell stimulation. Isoform 3 is not functional. Isoform 4 displays an increased activity. Isoform 5 and isoform 6 display reduced activity. -
Tissue specificity
Expressed by distal tubular cells in kidney cortex and in endothelial cells (at protein level). Widely expressed. Strongly expressed in kidney and to a lower extent in heart, adipocytes, hepatoma, endothelial cells, skeletal muscle, brain, several brain tumor cell lines and airway epithelial cells. -
Sequence similarities
Contains 1 FAD-binding FR-type domain.
Contains 1 ferric oxidoreductase domain. -
Developmental stage
Expressed in fetal kidney and fetal liver. -
Post-translational
modificationsIsoform 3 and isoform 4 are N-glycosylated. Isoform 4 glycosylation is required for its proper function. -
Cellular localization
Endoplasmic reticulum membrane. Cell membrane. Cell junction > focal adhesion. Nucleus. May localize to plasma membrane and focal adhesions. According to PubMed:15927447, may also localize to the nucleus. - Information by UniProt
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Database links
- Entrez Gene: 50507 Human
- Entrez Gene: 85431 Rat
- Omim: 605261 Human
- SwissProt: Q9NPH5 Human
- SwissProt: Q924V1 Rat
- Unigene: 371036 Human
- Unigene: 14744 Rat
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Alternative names
- Kidney oxidase 1 antibody
- Kidney oxidase-1 antibody
- Kidney superoxide producing NADPH oxidase antibody
see all
Images
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Anti-NADPH oxidase 4 antibody [UOTR1B492] (ab109225) at 1/10000 dilution (purified) + JAR cell lysate at 10 µg
Secondary
HRP goat anti-rabbit (H+L) at 1/1000 dilution
Predicted band size: 67 kDa
Observed band size: 63 kDa why is the actual band size different from the predicted?Blocking buffer: 5% NFDM/TBST
Dilution buffer: 5% NFDM/TBST
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Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-NADPH oxidase 4 antibody [UOTR1B492] (ab109225)
Immunohistochemical analysis of paraffin-embedded Human kidney tissue using unpurified ab109225.
Perform heat mediated antigen retrieval before commencing with IHC staining protocol.
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ICC/IF image of unpurified ab109255 stained HeLa cells. The cells were 4% formaldehyde fixed (10 min) and then incubated in 1%BSA / 10% normal goat serum / 0.3M glycine in 0.1% PBS-Tween for 1h to permeabilise the cells and block non-specific protein-protein interactions. The cells were then incubated with the antibody (ab109225, 5µg/ml) overnight at +4°C. The secondary antibody (green) was ab96899, DyLight® 488 goat anti-rabbit IgG (H+L) used at a 1/250 dilution for 1h. Alexa Fluor® 594 WGA was used to label plasma membranes (red) at a 1/200 dilution for 1h. DAPI was used to stain the cell nuclei (blue) at a concentration of 1.43µM.
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Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-NADPH oxidase 4 antibody [UOTR1B492] (ab109225)
Immunohistochemical staining of paraffin embedded human stomach with purified ab109225 at a dilution of 1/500. A HRP goat anti-rabbit (ab97051) was used as the secondary antibody at a dilution of 1/500 and the sample was counter-stained with hematoxylin. Antigen retrieval was perfomed using Tris-EDTA buffer, pH 9.0. PBS was used instead of the primary antibody as the negative control, and is shown in the inset.
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Immunofluorescent staining of HeLa cells (fixed in 4% PFA, permeabilized with 0.1% Triton X 100) using purified ab109225 at a dilution of 1/200. An Alexa Fluor® 488 goat anti-rabbit antibody was used as the secondary at a dilution of 1/500 and the cells were counter stained with DAPI. The negative control is shown in the bottom right hand panel - for the negative control, Alex Fluor® 594 goat anti-mouse was used at a dilution of 1/500.
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Intracellular Flow Cytometry analysis of U87-MG (human glioblastoma) cells labeling NADPH oxidase 4 with purified ab109225 at 1/280 dilution (10ug/mL) (red). Cells were fixed with 4% paraformaldehyde and permeabilised with 90% methanol. A Goat anti rabbit IgG (Alexa Fluorr® 488) (1/2000 dilution) was used as the secondary antibody. Rabbit monoclonal IgG (Black) was used as the isotype control, cells without incubation with primary antibody and secondary antibody (Blue) were used as the unlabeled control.
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Anti-NADPH oxidase 4 antibody [UOTR1B492] (ab109225) at 1/2000 dilution (purified) + Human fetal kidney at 10 µg
Secondary
HRP anti-rabbit, specific to the non reducded form of IgG at 1/1000 dilution
Predicted band size: 67 kDa
Observed band size: 63 kDa why is the actual band size different from the predicted?Blocking buffer: 5% NFDM/TBST
Dilution buffer: 5% NFDM/TBST
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ab109225 (purified) at 1/30 immunoprecipitating NADPH oxidase 4 in HEK293. For western blotting, a HRP-conjugated anti-rabbit antibody was used as the secondary antibody (1/1000).
Blocking buffer and concentration: 5% NFDM/TBST.
Diluting buffer and concentration: 5% NFDM /TBST.
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All lanes : Anti-NADPH oxidase 4 antibody [UOTR1B492] (ab109225) at 1/1000 dilution (unpurified)
Lane 1 : Fetal kidney lysate
Lane 2 : U87-MG lysate
Lane 3 : 293T lysate
Lane 4 : JAR lysates
Lysates/proteins at 10 µg per lane.
Secondary
All lanes : Standard HRP labelled goat anti-rabbit at 1/2000 dilution
Predicted band size: 67 kDa
Observed band size: 63 kDa why is the actual band size different from the predicted? -
Unpurified ab109225 staining Nox4 in HeLa cells treated with (-)-cannabidiol (ab120448), by ICC/IF. Increase in Nox4 expression correlates with increased concentration of (-)-cannabidiol, as described in literature.
The cells were incubated at 37°C for 6h in media containing different concentrations of ab120448 ((-)-cannabidio) in DMSO, fixed with 4% formaldehyde for 10 minutes at room temperature and blocked with PBS containing 10% goat serum, 0.3 M glycine, 1% BSA and 0.1% tween for 2h at room temperature. Staining of the treated cells with ab109225 (5 µg/ml) was performed overnight at 4°C in PBS containing 1% BSA and 0.1% tween. A DyLight 488 goat anti-rabbit polyclonal antibody (ab96899) at 1/250 dilution was used as the secondary antibody.
Protocols
Datasheets and documents
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SDS download
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Datasheet download
References (73)
ab109225 has been referenced in 73 publications.
- Zhao HY et al. L-carnitine treatment attenuates renal tubulointerstitial fibrosis induced by unilateral ureteral obstruction. Korean J Intern Med 36:S180-S195 (2021). PubMed: 32942841
- Sun CY et al. Indoxyl sulfate caused behavioral abnormality and neurodegeneration in mice with unilateral nephrectomy. Aging (Albany NY) 13:6681-6701 (2021). PubMed: 33621199
- Liu N et al. NADPH and Mito-Apocynin Treatment Protects Against KA-Induced Excitotoxic Injury Through Autophagy Pathway. Front Cell Dev Biol 9:612554 (2021). PubMed: 33644049
- Cao JY et al. Autophagosome protects proximal tubular cells from aldosterone-induced senescence through improving oxidative stress. Ren Fail 43:556-565 (2021). PubMed: 33757397
- Zhou H et al. ASMq protects against early burn wound progression in rats by alleviating oxidative stress and secondary mitochondria-associated apoptosis via the Erk/p90RSK/Bad pathway. Mol Med Rep 23:N/A (2021). PubMed: 33760179