HIF-1 alpha Antibody (ESEE122)

Immunohistochemistry: HIF-1 alpha Antibody (ESEE122) [NB100-131] - Immunohistochemical analysis in non-GIST STS representing negative, and score 1-3 of Carbonic Anhydrase IX/CA9, GLUT-1, HIF-1 alpha, and HIF-2 alpha/EPAS1. non-GIST STS: non-gastrointestinal stromal tumor soft-tissue sarcomas, Carbonic Anhydrase IX/CA9: carbonic anhydrase IX, GLUT-1: glucose transporter-1, and HIF-1/2alpha: hypoxia induced factor 1/2alpha. Image collected and cropped by CiteAb from the following publication (//www.hindawi.com/journals/sarcoma/2012/541650/), licensed under a CC-BY licence.

Immunohistochemistry: HIF-1 alpha Antibody (ESEE122) [NB100-131] - Histologically distinct cell types in hemangioblastomas do not arise from a common ancestral clone. Representative images of sample SH-0622 acquired at 400x of (a) H + E and IHC for (b) HIF1-alpha reveal heterogenous cell types in this tumor characterized by a rich vascular network. Arrowheads indicate that the stromal cells demonstrate increased cytoplasmic staining for HIF1-alpha and VEGF, whereas the double arrowheads highlight PDGFR-beta protein restricted to vascular endothelium. Scale bar is 25 um. Image collected and cropped by CiteAb from the following publication (//www.actaneurocomms.org/content/2/1/167), licensed under a CC-BY licence.

Simple Western: HIF-1 alpha Antibody (ESEE122) [NB100-131] - Image shows a specific band for HIF-1 alpha in 0.5 mg/mL of Hypoxic HeLa lysate. This experiment was performed under reducing conditions using the 12-230 kDa separation system.

Immunocytochemistry/Immunofluorescence: HIF-1 alpha Antibody (ESEE122) [NB100-131] - Detection of HIF-1 alpha (red dye 568) in a cultured raw mouse macrophage cell line, using NB100-131. Photos courtesy of Susan Alexander and Hattie Gresham, PhD.

Immunocytochemistry/Immunofluorescence: HIF-1 alpha Antibody (ESEE122) [NB100-131] - Detection of HIF-1 alpha (red dye) in a cell cytospin from a lavage of a murine skin pouch infected with S. aureus. 100X magnification. Blue: DAPI nuclear staining.

Immunohistochemistry-Paraffin: HIF-1 alpha Antibody (ESEE122) [NB100-131] - Analysis of a FFPE mouse kidney tissue section using HIF-1 alpha antibody clone ESEE122 at 1ug/mL concentration. The detection was performed using X-cell plus universal HRP polymer detection system with Vector SG chromagen substrate. Image courtesy of a product review by Steven Grover.

Immunohistochemistry: HIF-1 alpha Antibody (ESEE122) [NB100-131] - Immunohistochemical staining of HIF-1 alpha in normal renal tissue (A) and clear cell renal cell carcinoma (CCRCC) (D). A homogeneous cytoplasmic staining of tubular cells and weak staining in glomerules was observed with HIF-1 alpha (A). In CCRCC, HIF-1 alpha immmunoreactivity was nuclear and/or cytoplasmic (D), while it was perimembranous and/or diffuse cytoplasmic for VEGF-A and VEFG-C (E and F). (magnification x200). Image collected and cropped by CiteAb from the following publication (//www.jeccr.com/content/28/1/40), licensed under a CC-BY licence.

Immunohistochemistry-Paraffin: HIF-1 alpha Antibody (ESEE122) [NB100-131] - Analysis of HIF-1 alpha in paraffin-embedded mouse kidney tissue section using anti-HIF-1 alpha antibody. Image from verified customer review.

Immunohistochemistry-Paraffin: HIF-1 alpha Antibody (ESEE122) [NB100-131] - Representative immunohistochemical expression for HIF-1alpha, c-Met, CA9 and GLUT1. HIF-1alpha is stained in cytoplasm shown with no staining in normal cervix (A), weak staining intensity in high grade CIN (B), and strong staining intensity in squamous cell carcinoma (C). Scale bar: 50 um. Image collected and cropped by CiteAb from the following publication (//www.translational-medicine.com/content/11/1/185), licensed under a CC-BY licence.

Immunocytochemistry/Immunofluorescence: HIF-1 alpha Antibody (ESEE122) [NB100-131] - Detection of HIF-1 alpha (red dye) in a cell cytospin from a lavage of a murine skin pouch infected with S. aureus, using NB100-131. Blue: DAPI nuclear staining. Image courtesy of Susan Alexander and Hattie Gresham, PhD.

Immunocytochemistry/Immunofluorescence: HIF-1 alpha Antibody (ESEE122) [NB100-131] - Detection of HIF-1 alpha (red dye 568) in a cultured raw mouse macrophage cell line. 100X magnification. Image courtesy of Susan Alexander and Hattie Gresham, PhD.

Immunohistochemistry-Paraffin: HIF-1 alpha Antibody (ESEE122) [NB100-131] - Negative control stain of human placenta (from sea level) using mouse IgG at 1:100. 4uM paraffin-embedded section.

Immunohistochemistry-Paraffin: HIF-1 alpha Antibody (ESEE122) [NB100-131] - Analysis of a FFPE tissue section of human renal cancer xenograft using HIF-1 alpha antibody (NB100-131 Lot 83115) at 1:200 dilution. The antibody generated a strong cytoplasmic staining mainly in the cancer cells. Only a fraction of cells depicted nuclear staining, while weak to negligible positivity was seen in the tumor stromal cells.

Immunohistochemistry: HIF-1 alpha Antibody (ESEE122) [NB100-131] - HIF-1 alpha staining in hypoxia-induced human placenta.

• Reactivity: Hu, Mu, Rt, Bv, Ca
• Applications: WB, Simple Western, Flow, IB, ICC/IF, IHC, IHC-Fr, IHC-P, IP
• Clone: ESEE122
• Clonality: Monoclonal
• Host: Mouse
• Conjugate: Unconjugated
• Concentration: 1.0 mg/ml

HIF-1 alpha Antibody (ESEE122) Summary

• Immunogen: This HIF-1 alpha Antibody (ESEE122) was developed against Human HIF-1 alpha, corresponding to amino acids 329 - 530 [Uniprot# Q16665].
• Localization: Cytoplasm, Nucleus
• Isotype: IgG1
• Clonality: Monoclonal
• Host: Mouse
• Gene: HIF1A
• Purity: Protein G purified

Applications/Dilutions

• Dilutions:
  • Western Blot 1:500-1:1000
  • Simple Western 1:2000
  • Flow Cytometry
  • Immunoblotting
  • Immunocytochemistry/Immunofluorescence 1:100
  • Immunohistochemistry 1:100-1:5000
  • Immunohistochemistry-Frozen 1:100-1:5000
  • Immunohistochemistry-Paraffin 1:100-1:5000
  • Immunoprecipitation 1:10-1:500
• Application Notes: Variable results have been obtained in Western blot.
  
  In Simple Western only 10 - 15 uL of the recommended dilution is used per data point. Separated by Size-Wes, Sally Sue/Peggy Sue. Use in immunoprecipitation reported in scientific literature (PMID: 26757928; Fig 1G). Use in FLOW reported in scientific literature (Gestier S. et al).
• 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.
• Control:

  Human Kidney Whole Tissue Lysate (Adult Whole Normal)
  COS-7 Nuclear Hypoxic Induced Cell Lysate
  Human Placenta Whole Tissue Lysate (Adult Whole Normal)
  HeLa CoCl2 treated/Untreated Cell Lysate
  HepG2 CoCl2 treated/Untreated Cell Lysate
  HeLa Hypoxic / Normoxic Cell Lysate
  HepG2 Hypoxic / Normoxic Cell Lysate
  HIF-1 alpha Knockout HeLa Cell Lysate

Reactivity Notes

Please note that this antibody is reactive to Mouse and derived from the same host, Mouse. Additional Mouse on Mouse blocking steps may be required for IHC and ICC experiments. Please contact Technical Support for more information.

Packaging, Storage & Formulations

• Storage: Aliquot and store at -20C or -80C. Avoid freeze-thaw cycles.
• Buffer: Tris-Glycine, 0.15M NaCl
• Preservative: 0.05% Sodium Azide
• Concentration: 1.0 mg/ml
• Purity: Protein G purified

Alternate Names for HIF-1 alpha Antibody (ESEE122)

• AINT
• anti-HIF-1 alpha
• anti-HIF1A
• ARNT interacting protein
• ARNT-interacting protein
• Basic-helix-loop-helix-PAS protein MOP1
• BHLHE78
• Class E basic helix-loop-helix protein 78
• HIF 1A
• HIF1 alpha
• HIF-1 alpha
• HIF1
• HIF1A
• HIF-1a
• HIF-1alpha
• HIF-1-alpha
• HIF1-alpha
• hypoxia inducible factor 1 alpha subunit, hypoxia inducible factor 1 subunit alpha
• hypoxia inducible factor 1, alpha subunit (basic helix-loop-helix transcription factor)
• hypoxia-inducible factor 1-alpha
• Member of PAS protein 1
• member of PAS superfamily 1
• MOP1
• PAS domain-containing protein 8
• PASD8

Background

Hypoxia contributes to the pathophysiology of human disease, including myocardial and cerebral ischemia, cancer, pulmonary hypertension, congenital heart disease and chronic obstructive pulmonary disease (1). In cancer and particularly solid tumors, hypoxia plays a critical role in the regulation of genes involved in stem cell renewal, epithelial to mesenchymal transition (EMT), metastasis and angiogenesis. In the tumor microenvironment (TME), hypoxia influences the properties and function of stromal cells (e.g., fibroblasts, endothelial and immune cells) and is a strong determinant of tumor progression (2,3).

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

Limitations

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.

Publications for HIF-1 alpha Antibody (NB100-131)(76)

Publications using NB100-131

• Yoo SY, Yoo JY, Kim HB, et al. Neuregulin-1 Protects Neuronal Cells Against Damage due to CoCl2-Induced Hypoxia by Suppressing Hypoxia-Inducible Factor-1 alpha and P53 in SH-SY5Y Cells Int Neurourol J Nov 1 2019 [PMID: 31795610] (WB, Human)
• Hubbard WB, Lashof-Sullivan M, Greenberg S et al. Hemostatic nanoparticles increase survival, mitigate neuropathology and alleviate anxiety in a rodent blast trauma model. Sci Rep Jul 13 2018 [PMID: 30006635] (IHC, Rat)
• Kanakis G, Grimelius L, Papaioannou D et al. Can insulin-like growth factor 1 (IGF-1), IGF-1 receptor connective tissue growth factor and Ki-67 labelling index have a prognostic role in pulmonary carcinoids? Oncotarget Apr 27 2018 [PMID: 29854305] (IHC, Human)
• Koh J, Jang JY, Keam B et al. EML4-ALK enhances programmed cell death-ligand 1 expression in pulmonary adenocarcinoma via hypoxia-inducible factor (HIF)-1a and STAT3. OncoImmunology. 2015 Oct 29
• Gestier S. Methodology for the measurement of HIF-1alpha in bovine leukocytes and assessment of the molecule as a biomarker for bovine respiratory disease outcome. Thesis. 2014 (FLOW, Bovine)
• Otsuka S, Sakakima H, Terashi T et al. Preconditioning exercise reduces brain damage and neuronal apoptosis through enhanced endogenous 14-3-3g after focal brain ischemia in rats. Brain Struct Funct. Nov 26 2018 [PMID: 30478609] (IHC-P, Rat)
• Daskalakis K, Kaltsas G, Oberg K, Tsolakis AV. Lung Carcinoids: long-term surgical results and the lack of prognostic value of somatostatin receptors and other novel IHC markers. Neuroendocrinology Sep 23 2018 [PMID: 30244255] (IHC, Human)
• Song KH, Kim JH, Lee YH et al. Mitochondrial reprogramming via ATP5H loss promotes multimodal cancer therapy resistance J. Clin. Invest. Aug 31 2018 [PMID: 30124467] (IHC-P, Human)
• Rossow L, Veitl S, Vorlova S et al LOX-catalyzed collagen stabilization is a proximal cause for intrinsic resistance to chemotherapy. Oncogene. 2018 Sep [PMID: 29780168] (IHC, Mouse)
  
  Details:
  Citation using the HRP version of this antibody.
• Jayabal P, Houghton PJ, Shiio Y. EWS-FLI-1 creates a cell surface microenvironment conducive to IGF signaling by inducing pappalysin-1 Genes Cancer 2017 Nov 01 [PMID: 29321818] (WB, Human)
• Toblli Jorge E, Cao Gabriel, Giani Jorge F et al. Markers of oxidative/nitrosative stress and inflammation in lung tissue of rats exposed to different intravenous iron compounds. Drug Design, Development and Therapy 2017 Aug 1 [PMID: 28814833] (IHC-P, Rat)
• Gustafsson H, Kale A, Dasu A et al. EPR Oximetry of Cetuximab-Treated Head-and-Neck Tumours in a Mouse Model. Cell Biochem. Biophys. 2017 Jul 29 [PMID: 28756482] (Mouse)
• Fan Y, Li H, Ma X et al. Dicer suppresses the malignant phenotype in VHL-deficient clear cell renal cell carcinoma by inhibiting HIF-2alpha. Oncotarget Apr 5 2016 [PMID: 26943772] (WB, Human)
• Gestier S. Methodology for the measurement of HIF-1alpha in bovine leukocytes and assessment of the molecule as a biomarker for bovine respiratory disease outcome Thesis 2014 (FLOW, Bovine)
  
  Details:
  This reference used the DyLight 488 version of NB100-131.
• Boyer DF, McKelvie PA, de Leval L et al. Fibrin-associated EBV-positive Large B-Cell Lymphoma: An Indolent Neoplasm With Features Distinct From Diffuse Large B-Cell Lymphoma Associated With Chronic Inflammation. Am. J. Surg. Pathol. 2017 Mar 01 [PMID: 28195879] (IHC)
• Huang T, Chung S, Chua S et al. Effect of early administration of lower dose versus high dose of fresh mitochondria on reducing monocrotaline-induced pulmonary artery hypertension in rat. Am J Transl Res. Dec 30 2016 [PMID: 28077992] (ICC/IF, Rat)
• Hjelmgren O, Gellerman K, Kjelldahl J et al. Increased Vascularization in the Vulnerable Upstream Regions of Both Early and Advanced Human Carotid Atherosclerosis. PLoS ONE. Dec 15 2016 [PMID: 27973542] (IHC, Human)
• Siebenmann C, Keramidas M E, Rundqvist H et al. Cutaneous exposure to hypoxia does not affect skin perfusion in humans. Acta Physiologica (Oxford, England) 2019 Jun 25 [PMID: 27809413] (IHC-Fr, Human)
• Cahill LS, Gazdzinski LM, Tsui AK et al. Functional and anatomical evidence of cerebral tissue hypoxia in young sickle cell anemia mice. J. Cereb. Blood Flow Metab. May 10 2016 [PMID: 27165012] (IHC-P, Mouse)
• Koh J, Jang JY, Keam B et al. EML4-ALK enhances programmed cell death-ligand 1 expression in pulmonary adenocarcinoma via hypoxia-inducible factor (HIF)-1a and STAT3. Oncoimmunology 2015 Oct 29 [PMID: 27141364] (IHC, Human)
• Miller AN, Deal D, Green J et al. Use of the Reamer/Irrigator/Aspirator Decreases Carotid and Cranial Embolic Events in a Canine Model. J Bone Joint Surg Am. 2016 Apr 20 [PMID: 27098324] (IHC, Canine)
• Kim Y, Nam HJ, Lee J et al. Methylation-dependent regulation of HIF-1alpha stability restricts retinal and tumour angiogenesis. Nat Commun. 2016 Jan 13 [PMID: 26757928]
• Ochi F, Shiozaki A, Ichikawa D et al. Carbonic Anhydrase XII as an Independent Prognostic Factor in Advanced Esophageal Squamous Cell Carcinoma. J Cancer. 2015 Aug 28 [PMID: 26316888] (IHC-P, Human)
  
  Details:
  HIF-1 alpha antibody (clone ESEE122) was used for IHC-P analysis of tissue sections from human subjects with advanced esophageal squamous cell carcinoma (Fig 2).
• Shankar Ganesh M, Taylor-Weiner Amaro, Lelic Nina et al. Sporadic hemangioblastomas are characterized by cryptic VHL inactivation. Acta Neuropathol Commun. 2014 Dec 24 [PMID: 25589003] (IHC)
• Shi X, Guo L-W, Seedial S M et al. TGF-b/Smad3 inhibit vascular smooth muscle cell apoptosis through an autocrine signaling mechanism involving VEGF-A. Cell Death Dis 2014 [PMID: 25010983] (WB, Rat)
• Hempel Casper, Hoyer Nils, Kildemoes Anna et al. Systemic and Cerebral Vascular Endothelial Growth Factor Levels Increase in Murine Cerebral Malaria along with Increased Calpain and Caspase Activity and Can be Reduced by Erythropoietin Treatment. Frontiers in Immunology 2014 [PMID: 24995009] (WB, Mouse)
• Vasuri F, Fittipaldi S, Abualhin M et al. Biochemical and Immunomorphological Evaluation of Hepatocyte Growth Factor and c-Met Pathway in Patients with Critical Limb Ischemia Eur J Vasc Endovasc Surg. 2014 Jun 16 [PMID: 24947080] (IHC-P, Human)
  
  Details:
  HIF-1 alpha antibody (clone ESEE122) used in IHC-P (dilution 1:5500) on human skin samples from normal and Critical Limb Ischemia/CLI patients - FFPE sections, HIER with 50 mM Tris- 0.2 mM EDTA pH 9, endogenous peroxidase blocked with 3% H2O2 in CH3OH/10 min RT, stained with NovoLink Polymer Detection Kit - DAB - hematoxylin; Negative controls - no primary; Positive control - endothelial cells of dermal and hypodermic arterial vessels. Staining images - Figure 3B : RT-PCR showed 4X increase in HIF1A in CLI samples, but conversely, IHC showed decreased HIF1A protein in CLI samples compared to control skin.
• Schaible EV, Windschügl J, Bobkiewicz W et al. 2-Methoxyestradiol confers neuroprotection and inhibits a maladaptive HIF-1a response after traumatic brain injury in mice. J. Neurochem. 3/7/2014 [PMID: 24606183] (IHC-Fr, ICC/IF, WB, Mouse)
• Stenvold Helge, Donnem Tom, Andersen Sigve et al. High tumor cell expression of microRNA-21 in node positive non-small cell lung cancer predicts a favorable clinical outcome. BMC Clin Pathol. 2014 Feb 13 [PMID: 24524655] (IHC)
• Tajdini M, Mirbagheri SA, Nikooie R et al. Tissue hypoxia in pathogenesis of ulcerative colitis: should we change all our beliefs?. Scand J Gastroenterol. 2013 Oct 18 [PMID: 24134784] (IHC, Human)
• Kim BW, Cho H, Chung JY et al. Prognostic assessment of hypoxia and metabolic markers in cervical cancer using automated digital image analysis of immunohistochemistry. J Transl Med. 2013 Aug 8 [PMID: 23927384] (IHC-P, Human)
• Liu M, Guo X, Wang S et al. BOLD-MRI of breast invasive ductal carcinoma: correlation of R2* value and the expression of HIF-1alpha. Eur Radiol 2013 Jul 9 [PMID: 23835924] (IHC-P, Human)
• Gestier S. Methodology for the measurement of HIF-1alpha in bovine leukocytes and assessment of the molecule as a biomarker for bovine respiratory disease outcome. (FLOW, Bovine)
  
  Details:
  Using the DyLight 488 conjugated version of NB100-131, catalog number NB100-131G.
• Scott EM, Boursiquot N, Beltran WA, Dubielzig RR. Early histopathologic changes in the retina and optic nerve in canine primary angle-closure glaucoma. Vet Ophthalmol 2013 Jul [PMID: 23826772] (IHC, Canine)
• Finger EC, Cheng CF, Williams TR et al. CTGF is a therapeutic target for metastatic melanoma. Oncogene 2013 Feb 25 [PMID: 23435419] (WB, Human)
• Hu T, Beattie WS, Mazer CD et al. Treatment with a Highly Selective beta1 Antagonist Causes Dose-Dependent Impairment of Cerebral Perfusion After Hemodilution in Rats. Anesth Analg 2013 Mar [PMID: 23400988] (IHC, ICC/IF, Rat)
• Rahimi AS, Wilson DD, Saylor DK et al. p16, Cyclin D1, and HIF-1 alpha Predict Outcomes of Patients with Oropharyngeal Squamous Cell Carcinoma Treated with Definitive Intensity-Modulated Radiation Therapy Int J Otolaryngol 2012 [PMID: 22888357] (IHC, Human)
• SmelE, Kilvaer TK, Sorbye S,et al. Prognostic impacts of hypoxic markers in soft tissue sarcoma Sarcoma 2012 [PMID: 22454562] (IHC, Human)
• Rezvani HR, Ali N, Serrano-Sanchez M et al. Loss of epidermal hypoxia-inducible factor-1alpha accelerates epidermal aging and affects re-epithelialization in human and mouse. J Cell Sci 2011 Dec 15 [PMID: 22193962] (ICC/IF)
• Namas RA, Metukuri MR, Dhupar R et al. Hypoxia-induced overexpression of BNIP3 is not dependent on hypoxia-inducible factor 1? in mouse hepatocytes. Shock. 2011 Aug [PMID: 21558981] (WB, Mouse)
• Eckert AW, Lautner MHW, Schutze A et al. Coexpression of hypoxia-inducible factor-1[alpha] and glucose transporter-1 is associated with poor prognosis in oral squamous cell carcinoma patients. Histopathology. 2011 Mar 25. [PMID: 21438910]
• Halterman MW. An improved method for the study of apoptosis-related genes using the tet-on system J Biomol Screen 2011 Mar [PMID: 21335597] (WB, Mouse)
• Andersen S, Eilertsen M, Donnem T et al. Diverging prognostic impacts of hypoxic markers according to NSCLC histology. Lung Cancer. 2011 Jun. [PMID: 21075472]
• Park S, Ha SY, Cho HY, Chung DH, Kim NR, Hong J, Cho EK. Prognostic implications of hypoxia-inducible factor-1a in epidermal growth factor receptor-negative non-small cell lung cancer. Lung Cancer;72(1):100-7. 2011 Apr. [PMID: 20822827]
• Bennecke M, Kriegl L, Bajbouj M et al. Ink4a/Arf and oncogene-induced senescence prevent tumor progression during alternative colorectal tumorigenesis. Cancer Cell 2010 Aug 9 [PMID: 20708155] (IHC)
• Lerman OZ, Greives MR, Singh SP et al. Low-dose radiation augments vasculogenesis signaling through HIF-1-dependent and -independent SDF-1 induction. Blood. 2010 Nov 4 [PMID: 20631377] (ICC/IF, Human)
• Arvidsson Y, Bergstrom A, Arvidsson L et al. Hypoxia stimulates CXCR4 signalling in ileal carcinoids. Endocr Relat Cancer 2010 Jun 1 [PMID: 20071457] (IHC, Human)
• Lei Z, Li B, Yang Z et al. Regulation of HIF-1alpha and VEGF by miR-20b tunes tumor cells to adapt to the alteration of oxygen concentration. PLoS One 2009 [PMID: 19893619]
• Ragoonanan TE, Beattie WS, Mazer CD et al. Metoprolol reduces cerebral tissue oxygen tension after acute hemodilution in rats. Anesthesiology 2009 Nov [PMID: 19809291] (WB, Rat)
• Rasheed S, Harris AL, Tekkis PP et al. Hypoxia-inducible factor-1alpha and -2alpha are expressed in most rectal cancers but only hypoxia-inducible factor-1alpha is associated with prognosis. Br J Cancer 2009 May 19 [PMID: 19436307] (IHC-P, Human)
• Pina-Oviedo S, Khalili K, Del Valle L. Hypoxia inducible factor-1 alpha activation of the JCV promoter: role in the pathogenesis of progressive multifocal leukoencephalopathy. Acta Neuropathol 2009 Aug [PMID: 19360424] (IHC-P, ICC/IF, Human)
• Dordevic G, Matusan-Ilijas K, Babarovic E et al. Hypoxia inducible factor-1a correlates with vascular endothelial growth factor A and C indicating worse prognosis in clear cell renal cell carcinoma. Journal of Experimental & Clinical Cancer Research, 28:40. 2009 [PMID: 19302703] (IHC-P, Human)
• Gomes EM, Rodrigues MS, Phadke AP et al. Antitumor activity of an oncolytic adenoviral-CD40 ligand (CD154) transgene construct in human breast cancer cells. Clin Cancer Res;15(4):1317-25. 2009 Feb 15. [PMID: 19228733] (IHC, Human)
• Viola RJ, Provenzale JM, Li F et al. In vivo bioluminescence imaging monitoring of hypoxia-inducible factor 1alpha, a promoter that protects cells, in response to chemotherapy. AJR Am J Roentgenol 2008 Dec [PMID: 19020250] (IHC, Mouse)
• Sutton TA, Wilkinson J, Mang HE, Knipe NL et al. p53 regulates renal expression of HIF-1{alpha} and pVHL under physiological conditions and after ischemia-reperfusion injury. Am J Physiol Renal Physiol;295(6):F1666-77. 2008 Dec. [PMID: 18815219] (IHC-P, Rat)
• Reiser-Erkan C, Erkan M, Pan Z et al. Hypoxia-inducible proto-oncogene Pim-1 is a prognostic marker in pancreatic ductal adenocarcinoma. Cancer Biol Ther 2008 Sep [PMID: 18708761] (WB, Human)
• Shimogai R, Kigawa J, Itamochi H et al. Expression of hypoxia-inducible factor 1alpha gene affects the outcome in patients with ovarian cancer. Int J Gynecol Cancer 2008 May-Jun [PMID: 18476949] (IHC-P, Human)
• Kanasaki K, Palmsten K, Sugimoto H et al. Deficiency in catechol-O-methyltransferase and 2-methoxyoestradiol is associated with pre-eclampsia. Nature 2008 Jun 19 [PMID: 18469803]
• Hartmann H, Eltzschig HK, Wurz H, Hantke K, Rakin A, Yazdi AS, Matteoli G, Bohn E, Autenrieth IB, Karhausen J, Neumann D, Colgan SP, Kempf VA. Hypoxia-independent activation of HIF-1 by enterobacteriaceae and their siderophores. Gastroenterology;134(3):756-67. 2008 Mar. [PMID: 18325389] (IHC-P, WB, Mouse)
• Kubo T, Sugita T, Shimose S et al. Expression of hypoxia-inducible factor-1alpha and its relationship to tumour angiogenesis and cell proliferation in cartilage tumours. J Bone Joint Surg 2008 Mar. [PMID: 18310762]
• Piovan E, Tosello V, Indraccolo S et al. Differential regulation of hypoxia-induced CXCR4 triggering during B-cell development and lymphomagenesis. Cancer Res;67(18):8605-14. 2007 Sep 15. [PMID: 17875700] (IHC-P,IHC-Fr, Human)
• Monsef N, Helczynski L, Lundwall A et al. Localization of immunoreactive HIF-1alpha and HIF-2alpha in neuroendocrine cells of both benign and malignant prostate glands. Prostate;67(11):1219-29. 2007 Aug 1. [PMID: 17562539]
• Lee S, Garner EI, Welch WR, Berkowitz RS, Mok SC. Over-expression of hypoxia-inducible factor 1 alpha in ovarian clear cell carcinoma. Gynecol Oncol;106(2):311-7. 2007 Aug. [PMID: 17532031] (IHC-P, Human)
• Kyriacou HM, Steen KE, Raza A et al. In vitro inhibition of Plasmodium falciparum rosette formation by Curdlan sulfate. Antimicrob Agents Chemother 2007 Apr [PMID: 17283200] (IHC)
• Choke E, Cockerill GW, Dawson J et al. Hypoxia at the site of abdominal aortic aneurysm rupture is not associated with increased lactate. Ann N Y Acad Sci;1085:306-10. 2006 Nov. [PMID: 17182947]
• Khan Z, Michalopoulos GK, Stolz DB. Peroxisomal localization of hypoxia-inducible factors and hypoxia-inducible factor regulatory hydroxylases in primary rat hepatocytes exposed to hypoxia-reoxygenation. Am J Pathol;169(4):1251-69. 2006 Oct. [PMID: 17003483] (ICC/IF, IHC-P, Rat)
• Carroll, P A et al. Latent Kaposi's sarcoma-associated herpesvirus infection of endothelial cells activates hypoxia-induced factors. J Virol;80(21):10802-12. 2006 Nov. [PMID: 16956952] (IHC-P, Human)
• Ardyanto TD, Osaki M, Tokuyasu N et al. CoCl2-induced HIF-1alpha expression correlates with proliferation and apoptosis in MKN-1 cells: a possible role for the PI3K/Akt pathway. Int J Oncol 2006 Sep [PMID: 16865270] (WB, Human)
• Lidgren A, Hedberg Y, Grankvist K et al. Hypoxia-inducible factor 1alpha expression in renal cell carcinoma analyzed by tissue microarray. Eur Urol 2006 Dec [PMID: 16814458] (IHC, Human)
• Hirami Y, Aoe M, Tsukuda K, Hara F, Otani Y, Koshimune R, Hanabata T, Nagahiro I, Sano Y, Date H, Shimizu N. Relation of epidermal growth factor receptor, phosphorylated-Akt, and hypoxia-inducible factor-1alpha in non-small cell lung cancers. Cancer Lett;214(2):157-64. 2004 Oct 28. [PMID: 15363541]
• Abbate A, Santini D, Biondi-Zoccai GG et al. Cyclo-oxygenase-2 (COX-2) expression at the site of recent myocardial infarction: friend or foe?. Heart;90(4):440-3. 2004 Apr. [PMID: 15020525] (IHC-P, Human)
• Beasley NJ, Leek R, Alam M et al. Hypoxia-inducible factors HIF-1alpha and HIF-2alpha in head and neck cancer: relationship to tumor biology and treatment outcome in surgically resected patients. Cancer Res;62(9):2493-7. 2002 May 1. [PMID: 11980639] (IHC-P, Human)
• Giatromanolaki A, Koukourakis MI, Sivridis E et al. Expression of hypoxia-inducible carbonic anhydrase-9 relates to angiogenic pathways and independently to poor outcome in non-small cell lung cancer. Cancer Res;61(21):7992-8. 2001 Nov 1. [PMID: 11691824] (IHC-P, Human)
• Koukourakis MI, Giatromanolaki A, Skarlatos J et al. Hypoxia inducible factor (HIF-1a and HIF-2a) expression in early esophageal cancer and response to photodynamic therapy and radiotherapy. Cancer Res;61(5):1830-2. 2001 Mar 1. [PMID: 11280732] (IHC-P, Human)
• Talks KL, Turley H, Gatter KC et al. The expression and distribution of the hypoxia-inducible factors HIF-1alpha and HIF-2alpha in normal human tissues, cancers, and tumor-associated macrophages. Am J Pathol;157(2):411-21. 2000 Aug. [PMID: 10934146] (WB, IHC-P, Human)
• Wiesener MS, Turley H, Allen WE et al. Induction of endothelial PAS domain protein-1 by hypoxia: characterization and comparison with hypoxia-inducible factor-1alpha. Blood;92(7):2260-8. 1998 Oct 1. [PMID: 9746763]

Reviews for HIF-1 alpha Antibody (NB100-131) (3)

Average Rating: 4.3

(Based on 3 reviews)

We have 3 reviews tested in 2 species: Human, Mouse.

reviewed by: Anonymous Mouse 02/04/2015

Summary

• Species: Mouse
• File: View PDF

reviewed by: Anonymous IHC-P Mouse 01/31/2013

Summary

• Application: Immunohistochemistry-Paraffin
• Sample Tested: Murine Kidney
• Species: Mouse
• Lot: G5
• Comments: Excellent antibody providing strong positive staining with low levels of background, highly recommended for IHC-P, thanks Novus!

Blocking

• Blocking Details: Blocked with serum free protein block

Primary Anitbody

• Dilution Ratio: Incubated for 1 hour RT at a concentration of 1ug/ml

Secondary Antibody

• Secondary Description: X-cell plus universal HRP polymer detection system (Menarini Diagnostics)
• Secondary Manufacturer Cat#: MP-XCP-U25
• Secondary Concentration: According to manufacturers instructions

Details

• Detection Notes: Vector SG chromagen substrate (5 minute incubation)
• Fixation Details: Tissue fixed in formyl saline, heat mediated antigen retrieval performed at 100C in citrate (pH6) buffer
• Wash Description: 4 x 5 minute PBS washes

Comments

• Comments: Excellent antibody providing strong positive staining with low levels of background, highly recommended for IHC-P, thanks Novus!

reviewed by: Anonymous IHC-P Human 02/02/2009

Summary

• Application: Immunohistochemistry-Paraffin
• Sample Tested: human melanoma
• Species: Human
• Lot: E
• Comments: At this dilution there is still some background but the nuclear staining is very clear.

Blocking

• Blocking Details: Blocking Buffer: goat serum, Blocking Time: 30 minutes, Blocking Temp: room temperature

Primary Anitbody

• Dilution Ratio: Dilution Ratio: 1/1000, Incubation Dilution Buffer: blocking buffer, Incubation Time: overnight, Incubation Temp: 4 C

Secondary Antibody

• Secondary Description: Secondary Ab: goat anti rabbit from Santa Cruz, Secondary Ab Dilution: 1/200

Comments

• Comments: At this dilution there is still some background but the nuclear staining is very clear.

FAQs for HIF-1 alpha Antibody (NB100-131). (Showing 1 - 10 of 15 FAQs).

1. Why is there a difference between the theoretical MW for HIF1A and the observed MW for HIF-1 alpha?
   • HIF1A, like many other proteins, has post-translational modifications. Depending on the size, amount and nature of the post-translational modifications, it can cause subtle to very large changes in molecular weight.
2. Which antibody(ies) do you recommend for the detection of HIF-1a by immunohistochemistry in the sections of paraffin-embedded mouse liver samples? I would appreciate if you can give me several choices and rank them in the order of performance. My goal is to distinguish HIF upregulation by prolyl hydroxylase inhibitor in different liver cells.
   • All of our antibodies are of high quality and are well tested/validated in species/applications we list on the datasheet. However, we suggest the following four HIF-1 alpha antibodies based upon customer reviews, as well as the number of peer reviewed publications in which these products have been cited by researchers from reputed institutes. (1) HIF-1 alpha Antibody (H1alpha67) (cat# NB100-105) (cited in at least 218 peer reviewed publications) (2) HIF-1 alpha Antibody (cat# NB100-479) (cited in at least 51 peer reviewed publications) (3) HIF-1 alpha Antibody (H1alpha67) (cat# NB100-123 ) (cited in at least 38 peer reviewed publications) (4) HIF-1 alpha Antibody (cat# NB100-449) (cited in at least 31 peer reviewed publications).
3. I would like to know, does a path exist for detection of HIF 1 in venous blood before and after revascularization of the leg? 
   • We are not entirely sure if HIF-1 alpha will be present in the leg after revascularization. It may be present, but you may want to search the literature to see if this has been looked at before. If not, then this would certainly be an experiment worth doing.
4. What is the molecular weight (kDa) of protein HIF 1 alpha in western blot?
   • The theoretical molecular weight of HIF 1-alpha is ~93kDa. However, you will likely see a band between 100-120kDa due to phosphorylation.
5. We got the Hif1a (https://www.novusbio.com/products/hif-1-alpha-antibody-h1alpha67_nb100-105 ) antibody from you guys. I used the concentration that is mentioned on your website, but I am getting a band of a completely different size (~70kDa) and not the 120 kDa mentioned.
   • HIF-1 alpha is a notoriously difficult protein to work with due to its rapid degradation. Therefore, the ~70kDa bands are most likely degradation products. It is very important to lyse the cells in hypoxic conditions. We strongly recommend lysing the cells directly into the Laemmli buffer and doing that quickly, so that the exposure to oxygen is minimized.Please go through our hypoxia related FAQs, you should find them very informative:https://www.novusbio.com/support/hypoxia-and-hif-faqsAlso, running a positive control may help confirm the band specificity in your samples. You may prepare them yourself or choose some from our catalog, for example: 1) HeLa Hypoxic / Normoxic Cell Lysate (NBP2-36452)2) HeLa Hypoxic (CoCl2) / Normoxic Cell Lysate (NBP2-36450)
6. I performed several Western Blots of HIF-1 alpha with different lysis buffers, whole lysates, and cytoplasm/nuclei extractions. I can’t seem to get a good western blot (poor signal, band much lower than expected, etc.). Can someone suggest some technical considerations/tricks I should consider using?
   • A major issue that researchers working with HIF-1 alpha is degradation due to exposure to oxygen. In western blot, this results in a weaker band and/or the appearance of multiple low molecular weight bands (40-80 kDa). We recommend preparing the lysates after collection of cells/tissues as quickly as possible (on ice), preferably in a hypoxic chamber. We also recommend including a true hypoxia mimetic control (eg: cells treated with CoCl2, DMOG… etc.). The controls help distinguish your band of interest from potential degradation/dimer bands.For more troubleshooting tips and frequently asked questions regarding hypoxia/HIFs, you can refer to our hypoxia-related FAQs: https://www.novusbio.com/support/hypoxia-and-hif-faqs
7. I am doing HIF1 westerns in HIF-overexpressing mouse liver and adipose tissue using Novus antirabbit HIF1a antibody with overnight incubation. I am getting strong bands around 90kDa. I am aware that HIF theoretical molecular weight is 93kDa, but in westerns, the HIF band is usually around 120kDa according to my internet research. Can someone let me know if I’m getting the right HIF band or just some non-specific bands? Thanks.
   • (1)    HIF-1 alpha’s theoretical molecular weight is 93kDa. The post translationally modified/ubiquitinated form of HIF-1 alpha protein (fails to undergo proteasomal degradation) shows up as a band in the 110-130 kDa range on a Western blot.(2)    The dimeric protein may appear at a position above 200 kDa on non-reducing gels.(3)    Importantly, HIFs are among the most rapidly degradable proteins; therefore, sample preparation is highly important when analyzing HIF1 alpha or HIF2 alpha. When degraded, HIF-1 alpha may show up between 40-80 kDa position on Western blot. Degradation may be avoided by preparing the samples as soon as possible after collection of cells/tissues in hypoxic chamber. Notably, the tissues/cells should be kept on ice during lysate preparation and the lysates should be analyzed as soon as possible.(4)    For troubleshooting suggestions/feedback on more than 25 similar frequently asked questions, I would recommend visiting Novus page: FAQs - Hypoxia and HIFs https://www.novusbio.com/support/hypoxia-and-hif-faqs(5)    Last but not the least, Novus technical support team may be contacted at: technical@novusbio.com 
8. I have Hif1a nuclear protein extract at -80C. I am wondering if anyone knows how long it would be good for at that temperature since HIf1a is known to be degraded easily.Thank you!
   • You could try a few things to further inhibit the degradation.1) Use the protease inhibitors (if you are not already using them).2) Lyse cells into a buffer that contains SDS or LDS (eg: Laemmli's buffer), since SDS and LDS denature and inhibit proteases. Lysis may even be performed with reducing agents in the buffer (eg. DTT), but this will make your lysates unsuitable for BCA assay.3) Lysing samples rapidly ensures that the samples are instantly homogenized (it also shears DNA released by the SDS).5) Flash-freezing samples in liquid nitrogen rather than freezing at -80*C reduces the window of time for protease activity.6) Freeze samples in individual aliquots, instead of thawing the same vial multiple times.
9. I am curious to know the biochemical reactions of CoCl2 that mimic hypoxia. Is it that CoCl2 can bind any ubiquitin enzyme which regulates their degradation?
   • CoCl2 inhibits PHD enzymes (the body’s “oxygen sensors”) by replacing the Fe ion with Co, preventing these enzymes from marking HIF-1 alpha for degradation. CoCl2-based hypoxia mimetic samples are often used as positive control in HIF analysis. For more troubleshooting tips and frequently asked questions regarding hypoxia/HIFs, you can refer to our hypoxia-related FAQs: http://www.novusbio.com/support/hypoxia-and-hif-faqs
10. I am curious to know the biochemical reactions of CoCl2 that mimic hypoxia. Is it that CoCl2 can bind any ubiquitin enzyme which regulates their degradation?
    • CoCl2 inhibits PHD enzymes (the body’s “oxygen sensors”) by replacing the Fe ion with Co, preventing these enzymes from marking HIF-1 alpha for degradation. CoCl2-based hypoxia mimetic samples are often used as positive control in HIF analysis. For more troubleshooting tips and frequently asked questions regarding hypoxia/HIFs, you can refer to our hypoxia-related FAQs: https://www.novusbio.com/support/hypoxia-and-hif-faqs
11. Is cross-reactivity with HIF-2 alpha tested/predicted?
    • Although we don’t have cross-reactivity data with regards to HIF-2 alpha, we predict minimal cross-reactivity based on low sequence similarity observed from BLAST analysis between HIF-1 alpha and HIF-2 alpha.
12. Our customer wants to buy the peptide for your antibody NB100-131. Which catalog number does the peptide have?
    • This antibody was made to a recombinant protein containing amino acids 329-530 of HIF-1 alpha. Unfortunately we do not carry the recombinant protein as a sellable item.
13. Do you have any more protocol information regarding the cytospin staining images on your datasheet, credited to Susan Alexander and Hattie Gresham?
    • We do have the following notes:1) cultured raw cells (mouse macrophage cell line) were used2) cell cytospin from a lavage of a skin pouch (mouse) infected with staph aureus. 10 100X pic3) Also a cell cytospin from a lavage of a skin pouch of a mouse infected with staph aureus. 14. WT pic.4) cultured raw cells. 19. pic The blue staining is DAPI (nuclear stain). The stain was conjugated with a kit from molecular probes (A-20184) following their protocol. The final antibody concentration is uncertain but possibly around 25 ug/mL, used at 1:2 or 1:3 (about 8-12ug/mL).
14. Is your HIF-1 alpha Antibody able to work in Western Blot application?
    • Yes, this antibody can be used in Western Blot.
15. I've done colorations with an HIF-1alpha antibody and am now wondering if it is HIF-1alpha we see or it may be cross-reactivity with any other in the cytoplasm in mast cells. The image is reminiscent of the Immunofluorescence staining available on your site for HIF-1 alpha Antibody (ESEE122) [NB100-131] (Detection of HIF-1 alpha (red dye) in a cell cytospin from a lavage of a murine skin pouch infected with Staph Aureus, using NB100-131.) Gresham and Alexander have done that picture, but I cannot find it published. How to explain the appearance of HIF-1 staining. Our staining is localized to the nucleus, but is bead-like.
    • HIF-1 alpha can be found at very low levels in the cytoplasm under normoxic conditions. During hypoxia, HIF1a is stabilized and translocates to the nucleus to act as a transcription factor. You should expect to see nuclear staining in your hypoxic samples, but may also observe faint cytoplasmic staining corresponding to degraded HIF. We do not have a publication from Gresham and Alexander on our website, and I believe that their images were submitted as a customer review of the antibody. However we do have five other publications referring to immunofluorescent staining, which you may find helpful. You can see these at the bottom of the "Images, Reviews and Publications" tab by filtering on application and selecting "ICC/IF".
Show All 15 FAQs.

Bioinformatics

• Gene Symbol: HIF1A