Anti-HIF-1 alpha antibody [ESEE122] (ab8366)
Key features and details
- Mouse monoclonal [ESEE122] to HIF-1 alpha
- Suitable for: ICC/IF, IHC-P
- Reacts with: Human
- Isotype: IgG1
Overview
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Product name
Anti-HIF-1 alpha antibody [ESEE122]
See all HIF-1 alpha primary antibodies -
Description
Mouse monoclonal [ESEE122] to HIF-1 alpha -
Host species
Mouse -
Tested applications
Suitable for: ICC/IF, IHC-Pmore details
Unsuitable for: WB -
Species reactivity
Reacts with: Human -
Immunogen
Recombinant fragment corresponding to Human HIF-1 alpha aa 300-550.
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Positive control
- IHC-P: Human colon tissue. ICC: HeLa BrdU cells.
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General notes
Under normoxic conditions HIF-1 alpha has a short half-life. It is largely undetectable in cells or tissues grown under normoxic conditions. It is stabilized only at O2 concentrations below 5% and upon stabilization under hypoxic conditions HIF-1 translocates to the nucleus. Therefore we recommend western blots using nuclear extracts and running Hypoxia treated samples as positive control (ab180880). Hypoxia can be induced with treatment using certain agents e.g. CoCl2 or DFO, etc. so proper sample preparation is critical.
This antibody clone is manufactured by Abcam. If you require a custom buffer formulation or conjugation for your experiments, please contact orders@abcam.com.
The Life Science industry has been in the grips of a reproducibility crisis for a number of years. Abcam is leading the way in addressing this with our range of recombinant monoclonal antibodies and knockout edited cell lines for gold-standard validation. Please check that this product meets your needs before purchasing.
If you have any questions, special requirements or concerns, please send us an inquiry and/or contact our Support team ahead of purchase. Recommended alternatives for this product can be found below, along with publications, customer reviews and Q&As
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. Avoid freeze / thaw cycle. -
Storage buffer
pH: 7.40
Preservative: 0.02% Sodium azide
Constituents: PBS, 6.97% L-Arginine -
Concentration information loading...
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Purity
Protein G purified -
Clonality
Monoclonal -
Clone number
ESEE122 -
Myeloma
NS1 -
Isotype
IgG1 -
Light chain type
unknown -
Research areas
Associated products
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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 ab8366 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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ICC/IF |
Use a concentration of 8 - 12 µg/ml.
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IHC-P | (2) |
1/1000 - 1/8000. Perform heat mediated antigen retrieval before commencing with IHC staining protocol.
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Notes |
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ICC/IF
Use a concentration of 8 - 12 µg/ml. |
IHC-P
1/1000 - 1/8000. Perform heat mediated antigen retrieval before commencing with IHC staining protocol. |
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: 3091 Human
- Omim: 603348 Human
- SwissProt: Q16665 Human
- Unigene: 597216 Human
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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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ab8366 staining HIF-1 alpha in Hela cells. Untreated and BrdU treated (10µM for 24 hours) cells. The cells were fixed with 4% paraformaldehyde (10 min), permeabilized with 0.1% PBS-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 overnight at 4°C with ab8366 at 10µg/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 confocal microscope (Leica-Microsystems TCS SP8) and a single confocal section is shown.
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IHC image of HIF-1-alpha staining in human normal colon 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 ab8366, 5µg/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
Protocols
Datasheets and documents
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SDS download
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Datasheet download
References (52)
ab8366 has been referenced in 52 publications.
- Lee TK et al. Hyperthermia accelerates neuronal loss differently between the hippocampal CA1 and CA2/3 through different HIF‑1α expression after transient ischemia in gerbils. Int J Mol Med 49:N/A (2022). PubMed: 35234273
- Nazon C et al. Together Intra-Tumor Hypoxia and Macrophagic Immunity Are Driven Worst Outcome in Pediatric High-Grade Osteosarcomas. Cancers (Basel) 14:N/A (2022). PubMed: 35326631
- Fu X et al. Qi-Long-Tian formula extract alleviates symptoms of acute high-altitude diseases via suppressing the inflammation responses in rat. Respir Res 22:52 (2021). PubMed: 33579290
- Liu R et al. A HIF1a-GPD1 feedforward loop inhibits the progression of renal clear cell carcinoma via mitochondrial function and lipid metabolism. J Exp Clin Cancer Res 40:188 (2021). PubMed: 34098990
- Thureau S et al. First Comparison between [18f]-FMISO and [18f]-Faza for Preoperative Pet Imaging of Hypoxia in Lung Cancer. Cancers (Basel) 13:N/A (2021). PubMed: 34439254