Anti-HIF-1-alpha antibody - ChIP Grade (ab2185)
- Product nameAnti-HIF-1-alpha antibody - ChIP GradeSee all HIF-1-alpha primary antibodies ...
- DescriptionRabbit polyclonal to HIF-1-alpha - ChIP Grade
- Tested applicationsIHC-P, WB, IP, Gel supershift assays, IHC-Fr, ICC, ICC/IF, ChIP more details
- Species reactivityReacts with: Mouse, Rat, Guinea pig, Human, Xenopus laevis, Monkey
Predicted to work with: Cow
Fusion protein corresponding to Human HIF-1-alpha aa 432-528.
- Storage instructionsStore at +4°C short term (1-2 weeks). Upon delivery aliquot. Store at -20°C. Avoid freeze / thaw cycle.
- Storage bufferPreservative: 0.05% Sodium azide
Constituents: Tris glycine, 0.87% Sodium chloride
- PurityWhole antiserum
- Clonality Polyclonal
Our Abpromise guarantee covers the use of ab2185 in the following tested applications.
The application notes include recommended starting dilutions; optimal dilutions/concentrations should be determined by the end user.
|WB||WB: 1/500 - 1/1000.|
|Gel supershift assays||GSA: 1/1 - 1/100.|
|IHC-Fr||IHC-Fr: 1/10 - 1/2000.|
|ICC||ICC: Use at an assay dependent concentration.|
|ICC/IF||ICC/IF: 1/10 - 1/2000.|
|ChIP||ChIP: Use 25µl for 106 cells.|
- FunctionFunctions 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 specificityExpressed 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 similaritiesContains 1 basic helix-loop-helix (bHLH) domain.
Contains 1 PAC (PAS-associated C-terminal) domain.
Contains 2 PAS (PER-ARNT-SIM) domains.
- DomainContains two independent C-terminal transactivation domains, NTAD and CTAD, which function synergistically. Their transcriptional activity is repressed by an intervening inhibitory domain (ID).
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 localizationCytoplasm. Nucleus. Cytoplasmic in normoxia, nuclear translocation in response to hypoxia. Colocalizes with SUMO1 in the nucleus, under hypoxia.
- Entrez Gene: 281814 Cow
- Entrez Gene: 3091 Human
- Entrez Gene: 15251 Mouse
- Entrez Gene: 29560 Rat
- Entrez Gene: 100337573 Xenopus laevis
- Entrez Gene: 380141 Xenopus laevis
- Omim: 603348 Human
- SwissProt: Q9XTA5 Cow
- SwissProt: Q16665 Human
- SwissProt: Q61221 Mouse
- SwissProt: O35800 Rat
- SwissProt: Q9I8A9 Xenopus laevis
- Unigene: 597216 Human
- Unigene: 3879 Mouse
- Unigene: 446610 Mouse
- Unigene: 10852 Rat
- Unigene: 4589 Xenopus laevis
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Anti-HIF-1-alpha antibody - ChIP Grade images
ICC/IF image of ab2185 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 (ab2185, 1/1000 dilution) overnight at +4°C. The secondary antibody (green) was Alexa Fluor® 488 goat anti-rabbit IgG (H+L) used at a 1/1000 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.
Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) analysis of Human kidney tissue sections labeling HIF-1-alpha with ab2185.
All lanes : Anti-HIF-1-alpha antibody - ChIP Grade (ab2185)
Lane 1 : Rat nuclear extract lysate - normoxic
Lane 2 : Rat nuclear extract lysate - hypoxic
ChIP analysis of HIF-1-alpha genomic sequences from HeLa cells cultivated in normoxic (N) or hypoxic (Hx) conditions, using a HIF1-alpha polyclonal antibody (ab2185). For a negative control, IgG was used and the input as a positive control in the subsequent PCR. Primers for known target genes were used HIF1 alpha, A. EPO and B. VEGF.
References for Anti-HIF-1-alpha antibody - ChIP Grade (ab2185)
This product has been referenced in:
- Biswas S et al. Insulin regulates hypoxia-inducible factor-1a transcription by reactive oxygen species sensitive activation of Sp1 in 3T3-L1 preadipocyte. PLoS One 8:e62128 (2013). WB ; Mouse . Read more (PubMed: 23626778) »
- Yi T et al. Hypoxia-inducible factor-1a (HIF-1a) promotes cap-dependent translation of selective mRNAs through up-regulating initiation factor eIF4E1 in breast cancer cells under hypoxia conditions. J Biol Chem 288:18732-42 (2013). ChIP ; Human . Read more (PubMed: 23667251) »