Anti-HIF-1 alpha antibody (ab2185)
Key features and details
- Rabbit polyclonal to HIF-1 alpha
- Suitable for: IHC-P, WB, ICC/IF
- Reacts with: Rat, Human
- Isotype: IgG
Overview
-
Product name
Anti-HIF-1 alpha antibody
See all HIF-1 alpha primary antibodies -
Description
Rabbit polyclonal to HIF-1 alpha -
Host species
Rabbit -
Tested Applications & Species
See all applications and species dataApplication Species ICC/IF HumanIHC-P HumanWB Rat
-
Immunogen
Fusion protein corresponding to Human HIF-1 alpha aa 432-528.
Database link: Q16665 -
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.
Properties
-
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: 0.05% Sodium azide
Constituent: PBS -
Concentration information loading... -
Purity
Immunogen affinity purified -
Clonality
Polyclonal -
Isotype
IgG -
Research areas
Associated products
-
Assay kits
-
ChIP Related Products
-
Compatible Secondaries
-
Isotype control
-
Recombinant Protein
-
Related Products
Applications
The Abpromise guarantee
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.
Tested applications are guaranteed to work and covered by our Abpromise guarantee.
Predicted to work for this combination of applications and species but not guaranteed.
Does not work for this combination of applications and species.
| Application | Species |
|---|---|
| ICC/IF |
Human
|
| IHC-P |
Human
|
| WB |
Rat
|
| Application | Abreviews | Notes |
|---|---|---|
| IHC-P | (2) |
1/100 - 1/500.
|
| WB | (3) |
1/500 - 1/1000.
Ensure cell lysis occurs quickly if removed from hypoxia. Ultrasonic lysis is recommended to enrich more nuclear lysate. Positive Control: Hypoxic samples such as HeLa-DFO treated whole cell lysate ab116322. For a stronger signal, HeLa-DFO treated nuclear extracts are recommended ab180880. The cell fractionation kit can also be purchased separately ab109719. Negative Control: Most normal tissues or cells, other than kidney or heart. The observed band size of HIF-1 alpha is not exactly as predicted due to the different forms of HIF-1 alpha below: |
| ICC/IF | (2) |
1/100 - 1/500.
|
| Notes |
|---|
|
IHC-P
1/100 - 1/500. |
|
WB
1/500 - 1/1000. Ensure cell lysis occurs quickly if removed from hypoxia. Ultrasonic lysis is recommended to enrich more nuclear lysate. Positive Control: Hypoxic samples such as HeLa-DFO treated whole cell lysate ab116322. For a stronger signal, HeLa-DFO treated nuclear extracts are recommended ab180880. The cell fractionation kit can also be purchased separately ab109719. Negative Control: Most normal tissues or cells, other than kidney or heart. The observed band size of HIF-1 alpha is not exactly as predicted due to the different forms of HIF-1 alpha below: |
|
ICC/IF
1/100 - 1/500. |
Target
-
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
-
Database links
- Entrez Gene: 3091 Human
- Entrez Gene: 29560 Rat
- Omim: 603348 Human
- SwissProt: Q16665 Human
- SwissProt: O35800 Rat
- Unigene: 597216 Human
- Unigene: 10852 Rat
-
Alternative names
- ARNT interacting protein antibody
- ARNT-interacting protein antibody
- Basic helix loop helix PAS protein MOP1 antibody
see all
Images
-
Immunocytochemistry/ Immunofluorescence - Anti-HIF-1 alpha antibody (ab2185)
Immunocytochemical analysis of HeLa (Human epithelial cell line from cervix adenocarcinoma) cells labeling HIF-1 alpha with ab2185. 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) - Anti-HIF-1 alpha antibody (ab2185)Immunohistochemical analysis of AN3CA (human endometrium adenocarcinoma cell line) xenograft tissue labeling HIF-1 alpha with ab2185 at 1/300 dilution. The signal was developed using HRP-labelled secondary antibody and DAB reagent, and the section was further counterstained using hematoxylin. The tested section depicted mainly a diffused cytoplasmic staining but there were some cells which showed nuclear signal also (representing hypoxic cells).
-
Western blot - Anti-HIF-1 alpha antibody (ab2185)All lanes : Anti-HIF-1 alpha antibody (ab2185)
Lane 1 : Rat nuclear extract lysate - normoxic
Lane 2 : Rat nuclear extract lysate - hypoxic -
Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-HIF-1 alpha antibody (ab2185)Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) analysis of Human kidney tissue sections labeling HIF-1-alpha with ab2185.
References (147)
ab2185 has been referenced in 147 publications.
- Chen X et al. Copper promotes the migration of bone marrow mesenchymal stem cells via Rnd3-dependent cytoskeleton remodeling. J Cell Physiol 235:221-231 (2020). PubMed: 31187497
- Xie S et al. Andrographolide Protects Against Adverse Cardiac Remodeling After Myocardial Infarction through Enhancing Nrf2 Signaling Pathway. Int J Biol Sci 16:12-26 (2020). PubMed: 31892842
- Han X et al. Advanced glycation end products enhance macrophage polarization to the M1 phenotype via the HIF-1a/PDK4 pathway. Mol Cell Endocrinol 514:110878 (2020). PubMed: 32464167
- Kwon H et al. Hypoxia-Preconditioned Placenta-Derived Mesenchymal Stem Cells Rescue Optic Nerve Axons Via Differential Roles of Vascular Endothelial Growth Factor in an Optic Nerve Compression Animal Model. Mol Neurobiol N/A:N/A (2020). PubMed: 32524519
- Li J et al. Spatial heterogeneity of oxygenation and haemodynamics in breast cancer resolved in vivo by conical multispectral optoacoustic mesoscopy. Light Sci Appl 9:57 (2020). PubMed: 32337021
