HIF-1 alpha Antibody (JE75-33)
Novus Biologicals, part of Bio-Techne | Catalog # NBP3-32424
Recombinant Monoclonal Antibody
Key Product Details
Species Reactivity
Human, Mouse, Rat
Applications
Immunocytochemistry/ Immunofluorescence, Immunohistochemistry, Immunohistochemistry-Paraffin, Western Blot
Label
Unconjugated
Antibody Source
Recombinant Monoclonal Rabbit IgG Clone # JE75-33
Concentration
1 mg/ml
Product Specifications
Immunogen
Recombinant protein within Human HIF-1 alpha aa 251-550 / 826. (Uniprot: Q16665)
Localization
Cytoplasm, Nucleus, Nucleus speckle.
Clonality
Monoclonal
Host
Rabbit
Isotype
IgG
Theoretical MW
93 kDa.
Disclaimer note: The observed molecular weight of the protein may vary from the listed predicted molecular weight due to post translational modifications, post translation cleavages, relative charges, and other experimental factors.
Disclaimer note: The observed molecular weight of the protein may vary from the listed predicted molecular weight due to post translational modifications, post translation cleavages, relative charges, and other experimental factors.
Scientific Data Images for HIF-1 alpha Antibody (JE75-33)
Western Blot: HIF-1 alpha Antibody (JE75-33) [NBP3-32424]
Western Blot: HIF-1 alpha Antibody (JE75-33) [NBP3-32424] - Western blot analysis of HIF-1 alpha on different lysates with Rabbit anti-HIF-1 alpha antibody (NBP3-32424) at 1/1,000 dilution.Lane 1: HeLa cell lysate
Lane 2: HeLa treated with 250uM CoCl2 for 6 hours cell lysate
Lane 3: HepG2 cell lysate
Lane 4: HepG2 treated with 250uM CoCl2 for 6 hours cell lysate
Lysates/proteins at 20 ug/Lane.
Predicted band size: 93 kDa
Observed band size: 120 kDa
Exposure time: 1minute 2 seconds;
4-20% SDS-PAGE gel.
Proteins were transferred to a PVDF membrane and blocked with 5% NFDM/TBST for 1 hour at room temperature. The primary antibody (NBP3-32424) at 1/1,000 dilution was used in 5% NFDM/TBST at 4 overnight. Goat Anti-Rabbit IgG - HRP Secondary Antibody at 1/50,000 dilution was used for 1 hour at room temperature.
Immunohistochemistry: HIF-1 alpha Antibody (JE75-33) [NBP3-32424]
Immunohistochemistry: HIF-1 alpha Antibody (JE75-33) [NBP3-32424] - Immunohistochemical analysis of paraffin-embedded human kidney tissue with Rabbit anti-HIF-1 alpha antibody (NBP3-32424) at 1/200 dilution.The section was pre-treated using heat mediated antigen retrieval with Tris-EDTA buffer (pH 9.0) for 20 minutes. The tissues were blocked in 1% BSA for 20 minutes at room temperature, washed with ddH2O and PBS, and then probed with the primary antibody (NBP3-32424) at 1/200 dilution for 1 hour at room temperature. The detection was performed using an HRP conjugated compact polymer system. DAB was used as the chromogen. Tissues were counterstained with hematoxylin and mounted with DPX.
Immunocytochemistry/ Immunofluorescence: HIF-1 alpha Antibody (JE75-33) [NBP3-32424]
Immunocytochemistry/ Immunofluorescence: HIF-1 alpha Antibody (JE75-33) [NBP3-32424] - Immunocytochemistry analysis of HeLa cells treated with or without 500μM CoCl2 for 24 hours labeling HIF-1 alpha with Rabbit anti-HIF-1 alpha antibody (NBP3-32424) at 1/100 dilution.Cells were fixed in 4% paraformaldehyde for 20 minutes at room temperature, permeabilized with 0.1% Triton X-100 in PBS for 5 minutes at room temperature, then blocked with 1% BSA in 10% negative goat serum for 1 hour at room temperature. Cells were then incubated with Rabbit anti-HIF-1 alpha antibody (NBP3-32424) at 1/100 dilution in 1% BSA in PBST overnight at 4 ℃. Goat Anti-Rabbit IgG H&L (iFluor™ 488) was used as the secondary antibody at 1/1,000 dilution. PBS instead of the primary antibody was used as the secondary antibody only control. Nuclear DNA was labelled in blue with DAPI.
Applications for HIF-1 alpha Antibody (JE75-33)
Application
Recommended Usage
Immunocytochemistry/ Immunofluorescence
1:100
Immunohistochemistry-Paraffin
1:200-1:500
Western Blot
1:1000
Formulation, Preparation, and Storage
Purification
Protein A purified
Formulation
1*TBS (pH7.4), 0.05% BSA and 40% Glycerol
Preservative
0.05% Sodium Azide
Concentration
1 mg/ml
Shipping
The product is shipped with polar packs. Upon receipt, store it immediately at the temperature recommended below.
Stability & Storage
Store at 4C short term. Aliquot and store at -20C long term. Avoid freeze-thaw cycles.
Background: HIF-1 alpha/HIF1A
HIF-1 or hypoxia inducible factor 1 (predicted molecular weight 93kDa), is a transcription factor commonly referred to as a "master regulator of the hypoxic response" for its central role in the regulation of cellular adaptations to hypoxia. In its active form under hypoxic conditions, HIF-1 is stabilized by the formation of a heterodimer of HIF-1 alpha and ARNT/HIF-1 beta subunits. Nuclear HIF-1 engages p300/CBP for binding to hypoxic response elements (HREs). This process induces transcription and regulation of genes including EPO, VEGF, iNOS2, ANGPT1 and OCT4 (4,5).
Under normoxic conditions, the HIF-1 alpha subunit is rapidly targeted and degraded by the ubiquitin proteasome system. This process is mediated by prolyl hydroxylase domain enzymes (PHDs), which catalyze the hydroxylation of key proline residues (Pro-402 and Pro-564) within the oxygen-dependent degradation domain of HIF-1 alpha. Once hydroxylated, HIF-1 alpha binds the von Hippel-Lindau tumor suppressor protein (pVHL) for subsequent ubiquitination and proteasomal degradation (4). pVHL dependent regulation of HIF-1 alpha plays a role in normal physiology and disease states. Regulation of HIF-1 alpha by pVHL is critical for the suppressive function of FoxP3+ regulatory Tcells (6). Repression of pVHL expression in chronic lymphocytic leukemia (CLL) B cells leads to HIF-1 alpha stabilization and increased VEGF secretion (7).
References
1. Semenza, G. L., Agani, F., Feldser, D., Iyer, N., Kotch, L., Laughner, E., & Yu, A. (2000). Hypoxia, HIF-1, and the pathophysiology of common human diseases. Advances in Experimental Medicine and Biology.
2. Muz, B., de la Puente, P., Azab, F., & Azab, A. K. (2015). The role of hypoxia in cancer progression, angiogenesis, metastasis, and resistance to therapy. Hypoxia. https://doi.org/10.2147/hp.s93413
3. Huang, Y., Lin, D., & Taniguchi, C. M. (2017). Hypoxia inducible factor (HIF) in the tumor microenvironment: friend or foe? Science China Life Sciences. https://doi.org/10.1007/s11427-017-9178-y
4. Koyasu, S., Kobayashi, M., Goto, Y., Hiraoka, M., & Harada, H. (2018). Regulatory mechanisms of hypoxia-inducible factor 1 activity: Two decades of knowledge. Cancer Science. https://doi.org/10.1111/cas.13483
5. Dengler, V. L., Galbraith, M. D., & Espinosa, J. M. (2014). Transcriptional regulation by hypoxia inducible factors. Critical Reviews in Biochemistry and Molecular Biology. https://doi.org/10.3109/10409238.2013.838205
6. Lee, J. H., Elly, C., Park, Y., & Liu, Y. C. (2015). E3Ubiquitin Ligase VHL Regulates Hypoxia-Inducible Factor-1 alpha to Maintain Regulatory T Cell Stability and Suppressive Capacity. Immunity. https://doi.org/10.1016/j.immuni.2015.05.016
7. Ghosh, A. K., Shanafelt, T. D., Cimmino, A., Taccioli, C., Volinia, S., Liu, C. G., ... Kay, N. E. (2009). Aberrant regulation of pVHL levels by microRNA promotes the HIF/VEGF axis in CLL B cells. Blood. https://doi.org/10.1182/blood-2008-10-185686
Long Name
Hypoxia Inducible Factor 1 Subunit Alpha
Alternate Names
BHLHE78, HIF 1A, HIF-1a, HIF1 alpha, HIF1A, MOP1, PASD8
Gene Symbol
HIF1A
Additional HIF-1 alpha/HIF1A Products
Product Documents for HIF-1 alpha Antibody (JE75-33)
Product Specific Notices for HIF-1 alpha Antibody (JE75-33)
This product is for research use only and is not approved for use in humans or in clinical diagnosis. Primary Antibodies are guaranteed for 1 year from date of receipt.
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