Mouse HIF-1 alpha ELISA Kit (ab275103)
Key features and details
- One-wash 90 minute protocol
- Sensitivity: 33.39 pg/ml
- Range: 117.19 pg/ml - 7500 pg/ml
- Sample type: Cell Lysate
- Detection method: Colorimetric
- Assay type: Sandwich (quantitative)
- Reacts with: Mouse
Overview
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Product name
Mouse HIF-1 alpha ELISA Kit
See all HIF-1 alpha kits -
Detection method
Colorimetric -
Precision
Intra-assay Sample n Mean SD CV% Extract 8 10% Inter-assay Sample n Mean SD CV% Extract 3 15% -
Sample type
Cell Lysate -
Assay type
Sandwich (quantitative) -
Sensitivity
33.39 pg/ml -
Range
117.19 pg/ml - 7500 pg/ml -
Recovery
Sample specific recovery Sample type Average % Range Cell Lysate 96 87% - 106% -
Assay time
1h 30m -
Assay duration
One step assay -
Species reactivity
Reacts with: Mouse -
Product overview
Human HIF-1 alpha ELISA kit (ab275103) is a single-wash 90 min sandwich ELISA designed for the quantitative measurement of Human HIF-1 alpha protein in cell and tissue extracts. It uses our proprietary SimpleStep ELISA® technology. Quantitate Human HIF-1 alpha with 33.39 pg/mL sensitivity.
SimpleStep ELISA® technology employs capture antibodies conjugated to an affinity tag that is recognized by the monoclonal antibody used to coat our SimpleStep ELISA® plates. This approach to sandwich ELISA allows the formation of the antibody-analyte sandwich complex in a single step, significantly reducing assay time. See the SimpleStep ELISA® protocol summary in the image section for further details. Our SimpleStep ELISA® technology provides several benefits:
-Single-wash protocol reduces assay time to 90 minutes or less
-High sensitivity, specificity and reproducibility from superior antibodies
-Fully validated in biological samples
-96-wells plate breakable into 12 x 8 wells strips
A 384-well SimpleStep ELISA® microplate (ab203359) is available to use as an alternative to the 96-well microplate provided with SimpleStep ELISA® kits.
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Notes
Hypoxia-inducible factor 1-alpha (HIF-1 alpha) is a constitutively expressed transcription factor that is degraded under normal oxygen tensions and stabilized when oxygen is limiting (hypoxia). Under hypoxic conditions, stabilized HIF-1 alpha translocates to the nucleus and promotes the transcription of genes that enable the cell to adapt to a low oxygen environment. The HIF-1 alpha mediated hypoxic response include promotion of angiogenesis and the switch from aerobic respiration to anaerobic glycolysis. Many of the HIF-1 alpha responsive genes encode proteins that promote glycolysis and/or inhibit oxidative phosphorylation included in the Warburg effect.
Abcam has not and does not intend to apply for the REACH Authorisation of customers’ uses of products that contain European Authorisation list (Annex XIV) substances.
It is the responsibility of our customers to check the necessity of application of REACH Authorisation, and any other relevant authorisations, for their intended uses. -
Platform
Pre-coated microplate (12 x 8 well strips)
Properties
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Storage instructions
Store at +4°C. Please refer to protocols. -
Components 1 x 96 tests 10X Mouse HIF-1 alpha Capture Antibody 1 x 600µl 10X Mouse HIF-1 alpha Detector Antibody 1 x 600µl 10X Wash Buffer PT (ab206977) 1 x 20ml 5X Cell Extraction Buffer PTR (ab193970) 1 x 10ml Antibody Diluent 5BR 1 x 6ml Mouse HIF-1 alpha Lyophilized Recombinant Protein 2 vials Plate Seals 1 unit Sample Diluent NS (ab193972) 1 x 12ml SimpleStep Pre-Coated 96-Well Microplate (ab206978) 1 unit Stop Solution 1 x 12ml TMB Development Solution 1 x 12ml -
Research areas
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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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Alternative names
- ARNT interacting protein
- ARNT-interacting protein
- Basic helix loop helix PAS protein MOP1
see all -
Database links
- Entrez Gene: 15251 Mouse
- SwissProt: Q61221 Mouse
- Unigene: 3879 Mouse
- Unigene: 446610 Mouse
Images
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SimpleStep ELISA Protocol Diagram
SimpleStep ELISA technology allows the formation of the antibody-antigen complex in one single step, reducing assay time to 90 minutes. Add samples or standards and antibody mix to wells all at once, incubate, wash, and add your final substrate. See protocol for a detailed step-by-step guide.
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Example of mouse HIF-1alpha standard curve in 1X Cell Extraction Buffer PTR.The HIF-1alpha standard curve was prepared as described in Section 10. Raw data values are shown in the table. Background-subtracted data values (mean +/- SD) are graphed.
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Interpolated concentrations of native HIF-1alpha in mouse NIH/3T3 cell extract either treated with deferoxamine or mock treatment based on a 100 µg/mL extract load.The concentrations of HIF-1alpha were measured in duplicate, interpolated from the HIF-1alpha standard curves and corrected for sample dilution. The interpolated dilution factor corrected values are plotted (mean +/- SD, n=2). The mean HIF-1alpha concentration was determined to be 1481 pg/mL.
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Interpolated concentrations of native HIF-1alpha in mouse deferoxamine treated NIH/3T3 cell extract based on a 10 µg/mL extract load.The concentrations of HIF-1alpha were measured in duplicate, interpolated from the HIF-1alpha standard curves and corrected for sample dilution. The interpolated dilution factor corrected values are plotted (mean +/- SD, n=2). The mean HIF-1alpha concentration was determined to be 485.55 pg/mL and 40.77 pg/ml in deferoxamine and mock treated cell extract, respectively.
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Sandwich ELISA - Mouse HIF-1 alpha ELISA Kit (ab275103)To learn more about the advantages of recombinant antibodies see here.
