Recombinant Anti-HIF-1 alpha antibody [EPR16897] (ab179483)
![Western blot - Anti-HIF-1 alpha antibody [EPR16897] (ab179483)](https://www.abcam.com/ps/products/179/ab179483/Images/ab179483-320321-anti-hif-1-alpha-antibody-epr16897-western-blot.jpg "Western blot - Anti-HIF-1 alpha antibody [EPR16897] (ab179483)")
Key features and details
- Produced recombinantly (animal-free) for high batch-to-batch consistency and long term security of supply
- Rabbit monoclonal [EPR16897] to HIF-1 alpha
- Suitable for: WB, ICC/IF
- Knockout validated
- Reacts with: Mouse, Rat, Human
Related conjugates and formulations
Overview
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Product name
Anti-HIF-1 alpha antibody [EPR16897]
See all HIF-1 alpha primary antibodies -
Description
Rabbit monoclonal [EPR16897] to HIF-1 alpha -
Host species
Rabbit -
Specificity
Stimulation is required for the detection of HIF-1 alpha in most samples. For better results in WB, please try 1% SDS Hot lysis method presented in applications section.
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Tested applications
Suitable for: WB, ICC/IFmore details -
Species reactivity
Reacts with: Mouse, Rat, Human
Predicted to work with: Sheep, Rabbit, Cow, Ferret, Non human primates
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Immunogen
Recombinant fragment. This information is proprietary to Abcam and/or its suppliers.
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Positive control
- WB: HAP1 treated with DMOD, HeLa treated with DMOG, HeLa treated with CoCl2; NIH/3T3 treated with CoCl2, and C6 treated with CoCl2 and MG-132 (ab141003) cell lysates. ICC/IF: HeLa cells treated with DFO (1 mM, 24 h).
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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.
Properties
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Form
Liquid -
Storage instructions
Shipped at 4°C. Store at +4°C short term (1-2 weeks). Upon delivery aliquot. Store at -20°C long term. Avoid freeze / thaw cycle. -
Storage buffer
Preservative: 0.01% Sodium azide
Constituents: 59% PBS, 40% Glycerol, 0.05% BSA -
Concentration information loading... -
Purity
Protein A purified -
Clonality
Monoclonal -
Clone number
EPR16897 -
Isotype
IgG -
Research areas
Associated products
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Alternative Versions
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Assay kits
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Compatible Secondaries
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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 ab179483 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 |
|---|---|---|
| WB | (5) |
1/1000. Detects a band of approximately 110 kDa (predicted molecular weight: 92 kDa).
This antibody (ab179483) has been confirmed for Mouse sample in WB. This antibody works better in 1% SDS Hot Lysates in WB. For Lysate preparation protocol, please refer to the protocol book in the protocol section and/or here (downloadable copy). |
| ICC/IF | (1) |
1/500.
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| Notes |
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WB
1/1000. Detects a band of approximately 110 kDa (predicted molecular weight: 92 kDa). This antibody (ab179483) has been confirmed for Mouse sample in WB. This antibody works better in 1% SDS Hot Lysates in WB. For Lysate preparation protocol, please refer to the protocol book in the protocol section and/or here (downloadable copy). |
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ICC/IF
1/500. |
Target
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Function
Functions as a master transcriptional regulator of the adaptive response to hypoxia. Under hypoxic conditions activates the transcription of over 40 genes, including, erythropoietin, glucose transporters, glycolytic enzymes, vascular endothelial growth factor, and other genes whose protein products increase oxygen delivery or facilitate metabolic adaptation to hypoxia. Plays an essential role in embryonic vascularization, tumor angiogenesis and pathophysiology of ischemic disease. Binds to core DNA sequence 5'-[AG]CGTG-3' within the hypoxia response element (HRE) of target gene promoters. Activation requires recruitment of transcriptional coactivators such as CREBPB and EP300. Activity is enhanced by interaction with both, NCOA1 or NCOA2. Interaction with redox regulatory protein APEX seems to activate CTAD and potentiates activation by NCOA1 and CREBBP. -
Tissue specificity
Expressed in most tissues with highest levels in kidney and heart. Overexpressed in the majority of common human cancers and their metastases, due to the presence of intratumoral hypoxia and as a result of mutations in genes encoding oncoproteins and tumor suppressors. -
Sequence similarities
Contains 1 basic helix-loop-helix (bHLH) domain.
Contains 1 PAC (PAS-associated C-terminal) domain.
Contains 2 PAS (PER-ARNT-SIM) domains. -
Domain
Contains two independent C-terminal transactivation domains, NTAD and CTAD, which function synergistically. Their transcriptional activity is repressed by an intervening inhibitory domain (ID). -
Post-translational
modificationsIn normoxia, is hydroxylated on Pro-402 and Pro-564 in the oxygen-dependent degradation domain (ODD) by EGLN1/PHD1 and EGLN2/PHD2. EGLN3/PHD3 has also been shown to hydroxylate Pro-564. The hydroxylated prolines promote interaction with VHL, initiating rapid ubiquitination and subsequent proteasomal degradation. Deubiquitinated by USP20. Under hypoxia, proline hydroxylation is impaired and ubiquitination is attenuated, resulting in stabilization.
In normoxia, is hydroxylated on Asn-803 by HIF1AN, thus abrogating interaction with CREBBP and EP300 and preventing transcriptional activation. This hydroxylation is inhibited by the Cu/Zn-chelator, Clioquinol.
S-nitrosylation of Cys-800 may be responsible for increased recruitment of p300 coactivator necessary for transcriptional activity of HIF-1 complex.
Requires phosphorylation for DNA-binding.
Sumoylated; by SUMO1 under hypoxia. Sumoylation is enhanced through interaction with RWDD3. Desumoylation by SENP1 leads to increased HIF1A stability and transriptional activity.
Ubiquitinated; in normoxia, following hydroxylation and interaction with VHL. Lys-532 appears to be the principal site of ubiquitination. Clioquinol, the Cu/Zn-chelator, inhibits ubiquitination through preventing hydroxylation at Asn-803.
The iron and 2-oxoglutarate dependent 3-hydroxylation of asparagine is (S) stereospecific within HIF CTAD domains. -
Cellular localization
Cytoplasm. Nucleus. Cytoplasmic in normoxia, nuclear translocation in response to hypoxia. Colocalizes with SUMO1 in the nucleus, under hypoxia. - Information by UniProt
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Database links
- Entrez Gene: 281814 Cow
- Entrez Gene: 3091 Human
- Entrez Gene: 15251 Mouse
- Entrez Gene: 100009579 Rabbit
- Entrez Gene: 29560 Rat
- Entrez Gene: 443519 Sheep
- Omim: 603348 Human
- SwissProt: Q16665 Human
see all -
Alternative names
- ARNT interacting protein antibody
- ARNT-interacting protein antibody
- Basic helix loop helix PAS protein MOP1 antibody
see all
Images
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Western blot - Anti-HIF-1 alpha antibody [EPR16897] (ab179483)All lanes : Anti-HIF-1 alpha antibody [EPR16897] (ab179483) at 1/1000 dilution
Lane 1 : Wild-type HAP1 whole cell lysate
Lane 2 : Wild type HAP1 treated with DMOG (0.5mM 18hr) whole cell lysate
Lane 3 : HIF1A knockout HAP1 whole cell lysate
Lane 4 : HIF1A knockout HAP1 treated with DMOG (0.5mM 18hr) whole cell lysate
Lane 5 : HeLa whole cell lysate
Lane 6 : HeLa treated with DMOG (0.5mM 18hr) whole cell lysate
Lysates/proteins at 40 µg per lane.
Predicted band size: 92 kDa
Observed band size: 105 kDa why is the actual band size different from the predicted?Lanes 1 - 6: Merged signal (red and green). Green - ab179483 observed at 105 kDa. Red - loading control, ab24834, observed at 50 kDa.
ab179483 was shown to specifically react with HIF-1 alpha in wild-type HAP1 treated DMOG (0.5mM 18hr) cells as signal was lost in HAP1 knockout treated DMOG (0.5 mM 18hr) knockout cells. Wild-type and HAP1 knockout samples were subjected to SDS-PAGE. ab179483 and ab24834 (Mouse anti-Histone H3 loading control) were incubated overnight at 4°C at 1/1000 dilution and 1/10000 dilution respectively. Blots were developed with Goat anti-Rabbit IgG H&L (IRDye® 800CW) preabsorbed ab216773 and Goat anti-Mouse IgG H&L (IRDye® 680RD) preabsorbed ab216776 secondary antibodies at 1/20000 dilution for 1 hour at room temperature before imaging.
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![Immunocytochemistry/ Immunofluorescence - Anti-HIF-1 alpha antibody [EPR16897] (ab179483)](https://www.abcam.com/ps/products/179/ab179483/Images/ab179483-388548-anti-hif-1-alpha-antibody-epr16897-immunocytochemistry-dfo-treated-hela.jpg)
Immunocytochemistry/ Immunofluorescence - Anti-HIF-1 alpha antibody [EPR16897] (ab179483)Immunofluorescent analysis of 4% paraformaldehyde-fixed, 0.1% Triton X-100 permeabilized HeLa (Human epithelial cells from cervix adenocarcinoma) cells labeling HIF-1-alpha with ab179483 at 1/500 dilution, followed by Goat anti-rabbit IgG (Alexa Fluor® 488) (ab150077) secondary antibody at 1/1000 dilution (green). Confocal image showing nuclear staining in HeLa cells treated with DFO (1 mM, 24 h). The nuclear counter stain is DAPI (blue). Tubulin is detected with ab195889 (anti-alpha Tubulin mouse mAb) (Alexa Fluor® 594) at 1/200 dilution (red).
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![Western blot - Anti-HIF-1 alpha antibody [EPR16897] (ab179483)](https://www.abcam.com/ps/products/179/ab179483/Images/ab179483-285921-anti-hif-1-alpha-antibody-epr16897-western-blot.jpg)
Western blot - Anti-HIF-1 alpha antibody [EPR16897] (ab179483)All lanes : Anti-HIF-1 alpha antibody [EPR16897] (ab179483) at 1/5000 dilution
Lane 1 : HeLa (Human cervix adenocarcinoma epithelial cell) whole cell lysates
Lane 2 : HeLa (Human cervix adenocarcinoma epithelial cell) treated with 0.5 mM CoCl2 for 6 hours whole cell lysates
Lysates/proteins at 15 µg per lane.
Secondary
All lanes : Goat Anti-Rabbit IgG H&L (HRP) (ab97051) at 1/20000 dilution
Predicted band size: 92 kDa
Observed band size: 110 kDa why is the actual band size different from the predicted?
Exposure time: 1 secondBlocking and diluting buffer: 5% NFDM/TBST
The different result is due to the lysates preparation method. The lysate in the left image is prepared by RIPA lysis method. The lysate in the right image is prepared by 1%SDS Hot lysis method. This antibody works better in 1%SDS Hot Lysates in WB.
For Lysate preparation protocol, please refer to the protocol book in the protocol section and/or here (downloadable copy).
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![Western blot - Anti-HIF-1 alpha antibody [EPR16897] (ab179483)](https://www.abcam.com/ps/products/179/ab179483/Images/ab179483-313313-anti-hif-1-alpha-antibody-epr16897-western-blot.jpg)
Western blot - Anti-HIF-1 alpha antibody [EPR16897] (ab179483)All lanes : Anti-HIF-1 alpha antibody [EPR16897] (ab179483) at 0.163 µg/ml
Lane 1 : Untreated C6 (rat glial tumor glial cell), whole cell lysate
Lane 2 : C6 treated with 400 µM CoCl2 and 20 µM MG-132 (ab141003) for 4 hours
Lysates/proteins at 10 µg per lane.
Secondary
All lanes : Goat Anti-Rabbit IgG H&L (HRP) (ab97051) at 1/20000 dilution
Predicted band size: 92 kDa
Observed band size: 110 kDa why is the actual band size different from the predicted?
Exposure time: 26 secondsBlocking and diluting buffer: 5% NFDM/TBST.
The expression of HIF-1 alpha is induced by CoCl2 and maintained by MG-132 (PMID: 15836611).
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![Western blot - Anti-HIF-1 alpha antibody [EPR16897] (ab179483)](https://www.abcam.com/ps/products/179/ab179483/Images/ab179483-237957-anti-hif-1-alpha-antibody-epr16897-western-blot.jpg)
Western blot - Anti-HIF-1 alpha antibody [EPR16897] (ab179483)All lanes : Anti-HIF-1 alpha antibody [EPR16897] (ab179483) at 1/1000 dilution
Lane 1 : HeLa (Human epithelial cells from cervix adenocarcinoma) cell lysate treated with 0.5mM CoCl2 (Cobalt(II) chloride) for 6 hours.
Lane 2 : Untreated HeLa cell lysate
Lysates/proteins at 10 µg per lane.
Secondary
All lanes : Goat Anti-Rabbit IgG, (H+L),Peroxidase conjugated at 1/1000 dilution
Predicted band size: 92 kDa
Observed band size: 110 kDa why is the actual band size different from the predicted?
Exposure time: 15 secondsBlocking/Dilution buffer: 5% NFDM/TBST.
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![Western blot - Anti-HIF-1 alpha antibody [EPR16897] (ab179483)](https://www.abcam.com/ps/products/179/ab179483/Images/ab179483-237956-anti-hif-1-alpha-antibody-epr16897-western-blot.jpg)
Western blot - Anti-HIF-1 alpha antibody [EPR16897] (ab179483)All lanes : Anti-HIF-1 alpha antibody [EPR16897] (ab179483) at 1/1000 dilution
Lane 1 : NIH/3T3 (Mouse embyro fibroblast cells) cell lysate treated with 0.1mM CoCl2 for 24 hours.
Lane 2 : untreated NIH/3T3 cell lysate
Lysates/proteins at 10 µg per lane.
Secondary
All lanes : Goat Anti-Rabbit IgG, (H+L),Peroxidase conjugated at 1/1000 dilution
Predicted band size: 92 kDa
Observed band size: 110 kDa why is the actual band size different from the predicted?
Exposure time: 30 secondsBlocking/Dilution buffer: 5% NFDM/TBST.
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![Anti-HIF-1 alpha antibody [EPR16897] (ab179483)](https://www.abcam.com/ps/products/179/ab179483/Images/ab179483-9-benefits-of-recombinant-antibodies.png)
Anti-HIF-1 alpha antibody [EPR16897] (ab179483)
Protocols
Datasheets and documents
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SDS download
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Datasheet download
Certificate of Compliance
References (80)
ab179483 has been referenced in 80 publications.
- Ye J et al. The effect of hypoxia on the proliferation capacity of dermal papilla cell by regulating lactate dehydrogenase. J Cosmet Dermatol 20:684-690 (2021). PubMed: 32599679
- Lee LL et al. Cardiac pericytes function as key vasoactive cells to regulate homeostasis and disease. FEBS Open Bio 11:207-225 (2021). PubMed: 33135334
- Sun J et al. Hypoxia-sensitive long noncoding RNA CASC15 promotes lung tumorigenesis by regulating the SOX4/ß-catenin axis. J Exp Clin Cancer Res 40:12 (2021). PubMed: 33407675
- Dong L et al. Hypoxic hUCMSC-derived extracellular vesicles attenuate allergic airway inflammation and airway remodeling in chronic asthma mice. Stem Cell Res Ther 12:4 (2021). PubMed: 33407872
- Jing L et al. Norwogonin attenuates hypoxia-induced oxidative stress and apoptosis in PC12 cells. BMC Complement Med Ther 21:18 (2021). PubMed: 33413359