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    products/primary-antibodies/hif-1-alpha-antibody-ep1215y-ab51608.pdf

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RecombinantRabMAb

Recombinant Anti-HIF-1 alpha antibody [EP1215Y] (ab51608)

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Western blot - Anti-HIF-1 alpha antibody [EP1215Y] (ab51608)
  • Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-HIF-1 alpha antibody [EP1215Y] (ab51608)
  • Immunocytochemistry/ Immunofluorescence - Anti-HIF-1 alpha antibody [EP1215Y] (ab51608)
  • ChIC/CUT&RUN sequencing - Anti-HIF-1 alpha antibody [EP1215Y] (ab51608)
  • Flow Cytometry (Intracellular) - Anti-HIF-1 alpha antibody [EP1215Y] (ab51608)
  • Immunoprecipitation - Anti-HIF-1 alpha antibody [EP1215Y] (ab51608)
  • Immunocytochemistry/ Immunofluorescence - Anti-HIF-1 alpha antibody [EP1215Y] (ab51608)
  • ChIC/CUT&RUN sequencing - Anti-HIF-1 alpha antibody [EP1215Y] (ab51608)
  • Western blot - Anti-HIF-1 alpha antibody [EP1215Y] (ab51608)
  • Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-HIF-1 alpha antibody [EP1215Y] (ab51608)
  • Immunocytochemistry/ Immunofluorescence - Anti-HIF-1 alpha antibody [EP1215Y] (ab51608)
  • Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-HIF-1 alpha antibody [EP1215Y] (ab51608)
  • Immunocytochemistry/ Immunofluorescence - Anti-HIF-1 alpha antibody [EP1215Y] (ab51608)
  • Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-HIF-1 alpha antibody [EP1215Y] (ab51608)
  • Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-HIF-1 alpha antibody [EP1215Y] (ab51608)
  • Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-HIF-1 alpha antibody [EP1215Y] (ab51608)
  • Western blot - Anti-HIF-1 alpha antibody [EP1215Y] (ab51608)
  • Western blot - Anti-HIF-1 alpha antibody [EP1215Y] (ab51608)
  • Western blot - Anti-HIF-1 alpha antibody [EP1215Y] (ab51608)
  • OI-RD Scanning - Anti-HIF-1 alpha antibody [EP1215Y] (ab51608)
  • Anti-HIF-1 alpha antibody [EP1215Y] (ab51608)

Key features and details

  • Produced recombinantly (animal-free) for high batch-to-batch consistency and long term security of supply
  • Rabbit monoclonal [EP1215Y] to HIF-1 alpha
  • Suitable for: ICC/IF, Flow Cyt (Intra), IP, WB, IHC-P, ChIC/CUT&RUN-seq
  • Reacts with: Human

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Overview

  • Product name

    Anti-HIF-1 alpha antibody [EP1215Y]
    See all HIF-1 alpha primary antibodies
  • Description

    Rabbit monoclonal [EP1215Y] to HIF-1 alpha
  • Host species

    Rabbit
  • Specificity

    ab51608 recognizes HIF-1-alpha. For mouse specific Hif-1-alpha rabbit monoclonal antibody, please see ab179483 (clone ID: EPR16897). ab179483 has been confirmed for mouse samples in WB.
  • Tested applications

    Suitable for: ICC/IF, Flow Cyt (Intra), IP, WB, IHC-P, ChIC/CUT&RUN-seqmore details
  • Species reactivity

    Reacts with: Human
  • Immunogen

    Synthetic peptide. This information is proprietary to Abcam and/or its suppliers.

  • Positive control

    • WB: DFO treated HeLa nuclear lysate (ab180880), Ramos cell lysate treated with Cocl2. IHC-P: Human ovarian and breast carcinoma, colonic adenocarcinoma and squamous cell cervical carcinoma tissues. Human gastric cancer and CRC tumour tissue ICC/IF: DFO treated Hela cells, Cocl2 treated HeLa cells and baicalein treated HepG2 cells Flow Cyt (intra): DFO treated HeLa cells IP: DFO treated HeLa nuclear lysate ChIC/CUT&RUN-seq: HeLa cells, HeLa cells treated with Cocl2 (500 µM 20h+4h) and MG-132(10µM 4h).
  • General notes

    For Mouse specific Hif-1-alpha rabbit monoclonal antibody, please see ab179483 (clone ID: EPR16897).

    ab179483 has been confirmed for Mouse sample in WB.

    We have mixed customer feedback towards the rat specificity so we are unable to confirm and guarantee its performance with rat samples. Please contact technical team for more information.

    This product is a recombinant monoclonal antibody, which offers several advantages including:

    • - High batch-to-batch consistency and reproducibility
    • - Improved sensitivity and specificity
    • - Long-term security of supply
    • - Animal-free production
    For more information see here.

    Our RabMAb® technology is a patented hybridoma-based technology for making rabbit monoclonal antibodies. For details on our patents, please refer to RabMAb® patents.

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.
  • Dissociation constant (KD)

    KD = 2.24 x 10 -10 M
    Learn more about KD
  • Storage buffer

    pH: 7.20
    Preservative: 0.01% Sodium azide
    Constituents: 59% PBS, 40% Glycerol, 0.05% BSA
  • Concentration information loading...
  • Purity

    Protein A purified
  • Clonality

    Monoclonal
  • Clone number

    EP1215Y
  • Isotype

    IgG
  • Research areas

    • Cardiovascular
    • Hypoxia
    • HIF
    • Epigenetics and Nuclear Signaling
    • Transcription
    • Domain Families
    • HLH / Leucine Zipper
    • HLH
    • Epigenetics and Nuclear Signaling
    • Cardiovascular/Immune
    • Hypoxia
    • HIF
    • Cancer
    • Invasion/microenvironment
    • Hypoxia
    • HIF
    • Cancer
    • Cancer Metabolism
    • Response to hypoxia
    • Metabolism
    • Pathways and Processes
    • Metabolic signaling pathways
    • Nucleotide metabolism
    • Metabolism
    • Pathways and Processes
    • Metabolism processes
    • Hypoxia
    • Metabolism
    • Types of disease
    • Cancer
    • Neuroscience
    • Processes

Associated products

  • Alternative Versions

    • Alexa Fluor® 647 Anti-HIF-1 alpha antibody [EP1215Y] (ab190569)
    • HRP Anti-HIF-1 alpha antibody [EP1215Y] (ab205833)
    • Anti-HIF-1 alpha antibody [EP1215Y] - BSA and Azide free (ab210073)
  • Assay kits

    • HIF-1 alpha Transcription Factor Assay Kit (ab133104)
  • Compatible Secondaries

    • Goat Anti-Rabbit IgG H&L (Alexa Fluor® 488) (ab150077)
    • Goat Anti-Rabbit IgG H&L (Alexa Fluor® 488) preadsorbed (ab150081)
    • Goat Anti-Rabbit IgG H&L (HRP) (ab205718)
    • Goat Anti-Rabbit IgG H&L (DyLight® 488) preadsorbed (ab96899)
  • Conjugation kits

    • PE / R-Phycoerythrin Conjugation Kit - Lightning-Link® (ab102918)
    • APC Conjugation Kit - Lightning-Link® (ab201807)
  • Isotype control

    • Rabbit IgG, monoclonal [EPR25A] - Isotype Control (ab172730)
  • Positive Controls

    • Hela-DFO treated (0.5mM, 24h) Nuclear Lysate (ab180880)
  • Recombinant Protein

    • Recombinant Human HIF-1 alpha protein (ab154478)
    • Recombinant Human HIF-1 alpha protein (ab48734)
  • Related Products

    • Prestained Protein Ladder – Broad molecular weight (10-245 kDa) (ab116028)
    • Baicalein, Lipoxygenase inhibitor. (ab120723)
    • HeLa nuclear extract lysate (ab150036)
    • Human HIF-1 alpha ELISA Kit (ab171577)
    • HIF-1 alpha (H1alpha67, EP1215Y, ESEE122, EPR16897) Antibody Panel (ab263460)

Applications

The Abpromise guarantee

Our Abpromise guarantee covers the use of ab51608 in the following tested applications.

The application notes include recommended starting dilutions; optimal dilutions/concentrations should be determined by the end user.

Application Abreviews Notes
ICC/IF
1/500.
Flow Cyt (Intra)
1/10000.

ab172730 - Rabbit monoclonal IgG, is suitable for use as an isotype control with this antibody.

IP
Use a concentration of 5 µg/ml.
WB (10)
1/100 - 1/1000. Predicted molecular weight: 93 kDa.

The antibody only works in hypoxic cell and tissue lysates.

For Mouse specific Hif-1-alpha rabbit monoclonal antibody, please see ab179483 (clone ID: EPR16897).

ab179483 has been confirmed for mouse samples in WB.

IHC-P (3)
1/100. Perform heat mediated antigen retrieval with Tris/EDTA buffer pH 9.0 before commencing with IHC staining protocol.

For IHC antigen retrieval – See protocols IHC Antigen Retrieval Protocol.

ChIC/CUT&RUN-seq
Use at an assay dependent concentration.
Notes
ICC/IF
1/500.
Flow Cyt (Intra)
1/10000.

ab172730 - Rabbit monoclonal IgG, is suitable for use as an isotype control with this antibody.

IP
Use a concentration of 5 µg/ml.
WB
1/100 - 1/1000. Predicted molecular weight: 93 kDa.

The antibody only works in hypoxic cell and tissue lysates.

For Mouse specific Hif-1-alpha rabbit monoclonal antibody, please see ab179483 (clone ID: EPR16897).

ab179483 has been confirmed for mouse samples in WB.

IHC-P
1/100. Perform heat mediated antigen retrieval with Tris/EDTA buffer pH 9.0 before commencing with IHC staining protocol.

For IHC antigen retrieval – See protocols IHC Antigen Retrieval Protocol.

ChIC/CUT&RUN-seq
Use at an assay dependent concentration.

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
    modifications

    In 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.
  • Target information above from: UniProt accession Q16665 The UniProt Consortium
    The Universal Protein Resource (UniProt) in 2010
    Nucleic Acids Res. 38:D142-D148 (2010) .

    Information by UniProt
  • Database links

    • Entrez Gene: 3091 Human
    • Omim: 603348 Human
    • SwissProt: Q16665 Human
    • Unigene: 597216 Human
    • Alternative names

      • ARNT interacting protein antibody
      • ARNT-interacting protein antibody
      • Basic helix loop helix PAS protein MOP1 antibody
      • Basic-helix-loop-helix-PAS protein MOP1 antibody
      • bHLHe78 antibody
      • Class E basic helix-loop-helix protein 78 antibody
      • HIF 1A antibody
      • HIF 1alpha antibody
      • HIF-1-alpha antibody
      • HIF-1alpha antibody
      • HIF-alpha antibody
      • HIF1 A antibody
      • HIF1 Alpha antibody
      • HIF1 antibody
      • HIF1-alpha antibody
      • HIF1A antibody
      • HIF1A_HUMAN antibody
      • hifla antibody
      • Hypoxia inducible factor 1 alpha antibody
      • Hypoxia inducible factor 1 alpha isoform I.3 antibody
      • Hypoxia inducible factor 1 alpha subunit antibody
      • Hypoxia inducible factor 1 alpha subunit basic helix loop helix transcription factor antibody
      • Hypoxia inducible factor 1, alpha subunit (basic helix loop helix transcription factor) antibody
      • Hypoxia inducible factor1alpha antibody
      • Hypoxia-inducible factor 1-alpha antibody
      • Member of PAS protein 1 antibody
      • Member of PAS superfamily 1 antibody
      • Member of the PAS Superfamily 1 antibody
      • MOP 1 antibody
      • MOP1 antibody
      • PAS domain-containing protein 8 antibody
      • PASD 8 antibody
      • PASD8 antibody
      see all

    Images

    • Western blot - Anti-HIF-1 alpha antibody [EP1215Y] (ab51608)
      Western blot - Anti-HIF-1 alpha antibody [EP1215Y] (ab51608)This image is courtesy of an anonymous Abreview
      All lanes : Anti-HIF-1 alpha antibody [EP1215Y] (ab51608) at 1/2000 dilution

      Lane 1 : MCF-7 (normoxia)
      Lane 2 : MCF-7 treated with 0.5% oxygen for 24 hours

      Lysates/proteins at 30000 cells per lane.

      Secondary
      All lanes : Polyclonal Swine anti-rabbit IgG HRP at 1/1000 dilution

      Predicted band size: 93 kDa



      Blocking buffer: 5% milk for 16 hours at 4°C. 

      See Abreview

    • Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-HIF-1 alpha antibody [EP1215Y] (ab51608)
      Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-HIF-1 alpha antibody [EP1215Y] (ab51608)Zeindl-Eberhart E et al. Epithelial-mesenchymal transition induces endoplasmic-reticulum-stress response in human colorectal tumor cells. PLoS One 9:e87386 (2014).

      Immunohistochemical analysis of Formalin-fixed paraffin-embedded human CRC tumour tissue using ab51608 for HIF-1 alpha staining. Endogenous peroxidase of sections was inhibited by 7.5% H2O2 at room temperature

      In central tumor areas of human CRCs β-catenin was typically localized at the cell membrane (A) whereas only a weak staining was observed for cytoplasmic GRP78 (B) and HIF-1 alpha staining was found to be negative (C). At the invasion front strong nuclear β-catenin was detectable indicating EMT (D, G). In corresponding regions strong cytoplasmic GRP78 expression was found (E, H). In some of the cases an intense nuclear HIF-1 alpha staining was observed (F, with hypoxia), but not in others (I, without hypoxia) (magnification 200×; scale bar: 100 µm).

    • Immunocytochemistry/ Immunofluorescence - Anti-HIF-1 alpha antibody [EP1215Y] (ab51608)
      Immunocytochemistry/ Immunofluorescence - Anti-HIF-1 alpha antibody [EP1215Y] (ab51608)

      ab51608 staining HIF-1-alpha in HeLa cell line treated with Cocl2 by ICC/IF (Immunocytochemistry/immunofluorescence). Cells were fixed with 4% Paraformaldehyde permeabilized with 0.1% Triton X-100. Samples were incubated with primary antibody (1/500). An Alexa Fluor® 488-conjugated Goat anti-rabbit IgG(1/200) was used as the secondary antibody. Nuclei were counterstained with DAPI(right hand image).

    • ChIC/CUT&RUN sequencing - Anti-HIF-1 alpha antibody [EP1215Y] (ab51608)
      ChIC/CUT&RUN sequencing - Anti-HIF-1 alpha antibody [EP1215Y] (ab51608)

      CUT&RUN was performed using the ChIC/CUT&RUN pAG-MNAse ab285373, 2.5 x 10^5 HeLa (Human cervix adenocarcinoma epithelial cell line) cells treated with Cocl2 (500 μM 20h+4h) and MG-132(10µM 4h) and 5µg of ab51608 [EP1215Y]. The resulting DNA was sequenced on the NovaSeq 6000 (PE150).

      Additional screenshots of mapped reads can be downloaded here.

    • Flow Cytometry (Intracellular) - Anti-HIF-1 alpha antibody [EP1215Y] (ab51608)
      Flow Cytometry (Intracellular) - Anti-HIF-1 alpha antibody [EP1215Y] (ab51608)

      Overlay histogram showing HeLa untreated (Blue line) and HeLa treated (Red line - Deferoxamine, 1mM, 24 hours) cells stained with ab51608. The cells were fixed with 80% methanol (5 min) and then permeabilized with 0.1% PBS-Tween for 20 min. The cells were then incubated in 1x PBS / 10% normal goat serum / 0.3M glycine to block non-specific protein-protein interactions followed by the antibody (ab51608, 1/11709 dilution) for 30 min at 22°C. The secondary antibody used was Alexa Fluorr®488 goat anti-rabbit IgG (H&L) (ab150081) at 1/2000 dilution for 30 min at 22°C. 

      Acquisition of >5,000 events were collected using a 20mW Argon ion laser (488nm) and 525/30 bandpass filter. 

       

       

    • Immunoprecipitation - Anti-HIF-1 alpha antibody [EP1215Y] (ab51608)
      Immunoprecipitation - Anti-HIF-1 alpha antibody [EP1215Y] (ab51608)

      HIF-1-alpha was immunoprecipitated using 0.5mg HeLa Nuclear DFO treated whole cell extract (ab180880), 5µg of Rabbit polyclonal to HIF1 alpha and 50µl of protein G magnetic beads (+). No antibody was added to the control (-).

      The antibody was incubated under agitation with Protein G beads for 10min, HeLa DFO treated whole cell extract lysate diluted in RIPA buffer was added to each sample and incubated for a further 10min under agitation.

      Proteins were eluted by addition of 40µl SDS loading buffer and incubated for 10min at 70°C; 10µl of each sample was separated on a SDS PAGE gel, transferred to a nitrocellulose membrane, blocked with 5% BSA and probed with unpurified ab51608.

      Secondary: Mouse monoclonal [SB62a] Secondary Antibody to Rabbit IgG light chain (HRP) (ab99697).

      Band: 110kDa; HIF1 alpha

    • Immunocytochemistry/ Immunofluorescence - Anti-HIF-1 alpha antibody [EP1215Y] (ab51608)
      Immunocytochemistry/ Immunofluorescence - Anti-HIF-1 alpha antibody [EP1215Y] (ab51608)

      HeLa cells were untreated or treated with 1mM Deferoxamine (DFO) for 24h and fixed with paraformaldehyde for imaging by fluorescent microscopy. Cells were blocked and stained with 1X blocking buffer (ab126587). Unpurified ab51608 was used at 1:500. DAPI was used to label the nucleus. HIF1 alpha staining is absent in untreated cells and induced by DFO treatment. HIF1 alpha localizes to the nucleus.

    • ChIC/CUT&RUN sequencing - Anti-HIF-1 alpha antibody [EP1215Y] (ab51608)
      ChIC/CUT&RUN sequencing - Anti-HIF-1 alpha antibody [EP1215Y] (ab51608)

      CUT&RUN was performed using the ChIC/CUT&RUN pAG-MNAse ab285373, 105 HeLa cells, and 5µg of ab51608 [EP1215Y]. The resulting DNA was sequenced on the Illumina NovaSeq 6000 to a depth of 10 million reads.

      Additional screenshots of mapped reads can be downloaded here.

    • Western blot - Anti-HIF-1 alpha antibody [EP1215Y] (ab51608)
      Western blot - Anti-HIF-1 alpha antibody [EP1215Y] (ab51608)
      Anti-HIF-1 alpha antibody [EP1215Y] (ab51608) at 1/100 dilution + Ramos Cells treated with Cocl2 at 10 µg

      Secondary
      Goat Anti-Rabbit IgG, (H+L), HRP- conjugated at 1/1000 dilution

      Predicted band size: 93 kDa

    • Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-HIF-1 alpha antibody [EP1215Y] (ab51608)
      Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-HIF-1 alpha antibody [EP1215Y] (ab51608)

      ab51608 staining HIF-1-alpha in Human ovarian carcinoma tissue sections by Immunohistochemistry (IHC-P - paraformaldehyde-fixed, paraffin-embedded sections). Tissue was fixed and paraffin-embedded, antigen retrieval was by heat mediation in Tris/EDTA buffer pH9. Samples were incubated with primary antibody (1/100). An undiluted HRP-conjugated anti-rabbit IgG was used as the secondary antibody. Tissue counterstained with Hematoxylin.

    • Immunocytochemistry/ Immunofluorescence - Anti-HIF-1 alpha antibody [EP1215Y] (ab51608)
      Immunocytochemistry/ Immunofluorescence - Anti-HIF-1 alpha antibody [EP1215Y] (ab51608)

      Unpurified ab51608 staining HIF-1-alpha in HepG2 cells treated with baicalein (ab120723), by ICC/IF. Increase in HIF-1-alpha expression correlates with increased concentration of baicalein as described in literature.
      The cells were incubated at 37°C for 6h in media containing different concentrations of ab120723 (baicalein) in DMSO, fixed with 4% formaldehyde for 10 minutes at room temperature and blocked with PBS containing 10% goat serum, 0.3 M glycine, 1% BSA and 0.1% tween for 2h at room temperature. Staining of the treated cells with ab51608 (5 µg/ml) was performed overnight at 4°C in PBS containing 1% BSA and 0.1% tween. A DyLight 488 goat anti-rabbit polyclonal antibody (ab96899) at 1/250 dilution was used as the secondary antibody.

    • Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-HIF-1 alpha antibody [EP1215Y] (ab51608)
      Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-HIF-1 alpha antibody [EP1215Y] (ab51608)Chen L et al. HIF-1 alpha overexpression correlates with poor overall survival and disease-free survival in gastric cancer patients post-gastrectomy. PLoS One 9:e90678 (2014).

      Immunohistochemical analysis of paraffin-embedded formalin-fixed human gastric cancer tissue stained for HIF-1 alpha using ab15608 at 1/600 dilution.  Tissue sections were counterstained with Mayer's hematoxylin. Citrate buffer (pH 6.0) antigen retrieval using standard methodology

      C. HIF-1 alpha was located mainly in the nucleus of tumor cells (positive expression ×400).
      D. HIF-1 alpha original magnification ×100.

    • Immunocytochemistry/ Immunofluorescence - Anti-HIF-1 alpha antibody [EP1215Y] (ab51608)
      Immunocytochemistry/ Immunofluorescence - Anti-HIF-1 alpha antibody [EP1215Y] (ab51608)

      ab51608 staining of HIF-1-alpha in untreated HeLa cell line by ICC/IF (Immunocytochemistry/immunofluorescence). Cells were fixed with 4% Paraformaldehyde permeabilized with 0.1% Triton X-100. Samples were incubated with primary antibody (1/500). An Alexa Fluor® 488-conjugated Goat anti-rabbit IgG(1/200) was used as the secondary antibody. Nuclei were counterstained with DAPI(right hand Image).

    • Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-HIF-1 alpha antibody [EP1215Y] (ab51608)
      Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-HIF-1 alpha antibody [EP1215Y] (ab51608)

      Immunohistochemical analysis using unpurified ab51608 showing positive staining in Breast carcinoma tissue.

    • Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-HIF-1 alpha antibody [EP1215Y] (ab51608)
      Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-HIF-1 alpha antibody [EP1215Y] (ab51608)

      Immunohistochemical analysis using unpurified ab51608 showing positive staining in Colonic adenocarcinoma tissue. Heat mediated antigen retrieval was performed via the microwave method before commencing with IHC staining protocol.

    • Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-HIF-1 alpha antibody [EP1215Y] (ab51608)
      Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-HIF-1 alpha antibody [EP1215Y] (ab51608)

      Immunohistochemical analysis using unpurified ab51608 showing positive staining in Squamous cell cervical carcinoma tissue. Heat mediated antigen retrieval was performed via the microwave method before commencing with IHC staining protocol.

    • Western blot - Anti-HIF-1 alpha antibody [EP1215Y] (ab51608)
      Western blot - Anti-HIF-1 alpha antibody [EP1215Y] (ab51608)
      All lanes : Anti-HIF-1 alpha antibody [EP1215Y] (ab51608) at 1/2000 dilution (Unpurified)

      Lane 1 : HeLa nuclear extract lysate (ab150036)
      Lane 2 : Hela-DFO treated (0.5mM, 24h) Nuclear Lysate (ab180880)

      Lysates/proteins at 40 µg per lane.

      Secondary
      All lanes : Goat Anti-Rabbit IgG H&L (HRP) (ab97051) at 1/10000 dilution

      Developed using the ECL technique.

      Performed under reducing conditions.

      Predicted band size: 93 kDa
      Observed band size: 110 kDa why is the actual band size different from the predicted?


      Exposure time: 8 minutes


      Abcam recommends using 5% milk as the blocking agent, decreasing to 2% milk during primary and secondary incubation. Abcam welcomes customer feedback and would appreciate any comments regarding this product and the data presented above.

    • Western blot - Anti-HIF-1 alpha antibody [EP1215Y] (ab51608)
      Western blot - Anti-HIF-1 alpha antibody [EP1215Y] (ab51608)Image courtesy of Teresa Otto (University of Duisburg-Essen, Germany)

      Anti-HIF-1-alpha unpurified antibody (ab51608) reactivity with reduced Hep3B cell lysate after transient transfection of scrambled siRNA (lanes1-3 and 7-9) or HIF-1-alpha siRNA (lanes 4-6 and 10-12). Cells were incubated at with 21% O2 (lanes 1-6) or 1% O2 (lanes 7-12) for 4h before lysis. After SDS-PAGE, membranes were blocked in 5% milk for 1h at 25°C before incubation with unpurified ab51608 (1/1,000 dilution 5% milk) for 16h at 4ºC. The blot was then incubated with an anti-Rabbit HRP-conjugated secondary antibody before developing with ECL.

      See Abreview

    • Western blot - Anti-HIF-1 alpha antibody [EP1215Y] (ab51608)
      Western blot - Anti-HIF-1 alpha antibody [EP1215Y] (ab51608)
      All lanes : Anti-HIF-1 alpha antibody [EP1215Y] (ab51608) at 1/2000 dilution (unpurified)

      Lane 1 : HeLa Whole Cell Lysate (untreated, negative control)
      Lane 2 : HeLa DFO treated (0.5mM, 24h) Whole Cell Lysate
      Lane 3 : HeLa Nuclear Cell Lysate (untreated, negative control)
      Lane 4 : HeLa Nuclear DFO treated (0.5mM, 24h) Cell Lysate

      Lysates/proteins at 40 µg per lane.

      Secondary
      All lanes : Goat Anti-Rabbit IgG H&L (HRP) (ab97051) at 1/10000 dilution

      Performed under reducing conditions.

      Predicted band size: 93 kDa
      Observed band size: 110 kDa why is the actual band size different from the predicted?


      Exposure time: 2 minutes


      Abcam recommends using 5% milk as the blocking agent, decreasing to 2% milk during primary and secondary incubation. Abcam welcomes customer feedback and would appreciate any comments regarding this product and the data presented above.

    • OI-RD Scanning - Anti-HIF-1 alpha antibody [EP1215Y] (ab51608)
      OI-RD Scanning - Anti-HIF-1 alpha antibody [EP1215Y] (ab51608)
      Equilibrium disassociation constant (KD)
      Learn more about KD

      Click here to learn more about KD
    • Anti-HIF-1 alpha antibody [EP1215Y] (ab51608)
      Anti-HIF-1 alpha antibody [EP1215Y] (ab51608)

    Protocols

    To our knowledge, customised protocols are not required for this product. Please try the standard protocols listed below and let us know how you get on.

    Click here to view the general protocols

    Datasheets and documents

    • SDS download

    • Datasheet download

      Download

    References (244)

    Publishing research using ab51608? Please let us know so that we can cite the reference in this datasheet.

    ab51608 has been referenced in 244 publications.

    • Shi M  et al. APCCDC20-mediated degradation of PHD3 stabilizes HIF-1a and promotes tumorigenesis in hepatocellular carcinoma. Cancer Lett 496:144-155 (2021). PubMed: 33039559
    • Liu X  et al. Huaier shows anti-cancer activities by inhibition of cell growth, migration and energy metabolism in lung cancer through PI3K/AKT/HIF-1a pathway. J Cell Mol Med 25:2228-2237 (2021). PubMed: 33377619
    • Zhang W  et al. PlGF knockdown attenuates hypoxia-induced stimulation of cell proliferation and glycolysis of lung adenocarcinoma through inhibiting Wnt/ß-catenin pathway. Cancer Cell Int 21:18 (2021). PubMed: 33407494
    • Zhu F  et al. Tubular Numb promotes renal interstitial fibrosis via modulating HIF-1a protein stability. Biochim Biophys Acta Mol Basis Dis 1867:166081 (2021). PubMed: 33486098
    • Wang Z  et al. Resveratrol alleviates bleomycin-induced pulmonary fibrosis via suppressing HIF-1a and NF-?B expression. Aging (Albany NY) 13:4605-4616 (2021). PubMed: 33495418
    View all Publications for this product

    Customer reviews and Q&As

    Show All Reviews Q&A
    Submit a review Submit a question

    1-10 of 17 Abreviews or Q&A

    Western blot abreview for Anti-HIF1 alpha [EP1215Y] antibody

    Excellent
    Abreviews
    Abreviews
    abreview image
    Application
    Western blot
    Loading amount
    10 µg
    Gel Running Conditions
    Reduced Denaturing (7.5%, 1.5 mm)
    Sample
    Human Cell lysate - whole cell (Hep3B)
    Specification
    Hep3B
    Treatment
    1-6: 4h 21% oxygen, 7-12: 4h 1% oxygen 1-3 and 7-9: 50nM scrambled siRNA, 4-6 and 10-12: 50nM HIF-1alpha siRNA
    Blocking step
    Milk as blocking agent for 1 hour(s) and 0 minute(s) · Concentration: 5% · Temperature: 20°C
    Read More
    The reviewer received a reward from Abcam’s Loyalty Program in thanks for submitting this Abreview and for helping the scientific community make better-informed decisions.

    DR. Teresa Otto

    Verified customer

    Submitted Sep 09 2013

    Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) abreview for Anti-HIF-1 alpha antibody [EP1215Y]

    Excellent
    Abreviews
    Abreviews
    abreview image
    Application
    Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections)
    Sample
    Human Tissue sections (Lung Cancer (Xenograft))
    Antigen retrieval step
    Heat mediated - Buffer/Enzyme Used: Tris Buffer pH9
    Permeabilization
    No
    Specification
    Lung Cancer (Xenograft)
    Blocking step
    Protein Block Dako as blocking agent for 5 minute(s) · Concentration: 100% · Temperature: 20°C
    Fixative
    Formaldehyde
    Read More
    The reviewer received a reward from Abcam’s Loyalty Program in thanks for submitting this Abreview and for helping the scientific community make better-informed decisions.

    MS. Loreen Weichert

    Verified customer

    Submitted Aug 06 2021

    Western blot abreview for Anti-HIF-1 alpha antibody [EP1215Y]

    Excellent
    Abreviews
    Abreviews
    abreview image
    Application
    Western blot
    Sample
    Human Cell lysate - whole cell (RPE1 cell line)
    Gel Running Conditions
    Reduced Denaturing
    Loading amount
    30 µg
    Specification
    RPE1 cell line
    Blocking step
    Milk as blocking agent for 1 hour(s) and 0 minute(s) · Concentration: 5% · Temperature: 24°C
    Read More
    The reviewer received a reward from Abcam’s Loyalty Program in thanks for submitting this Abreview and for helping the scientific community make better-informed decisions.

    Abcam user community

    Verified customer

    Submitted Mar 05 2021

    Western blot abreview for Anti-HIF-1 alpha antibody [EP1215Y]

    Good
    Abreviews
    Abreviews
    abreview image
    Application
    Western blot
    Sample
    Human Cell lysate - whole cell (T-cell Lymphoma)
    Gel Running Conditions
    Non-reduced Non-Denaturing (Native) (4-15)
    Loading amount
    20 µg
    Treatment
    Hypoxic environment 1% O2
    Specification
    T-cell Lymphoma
    Blocking step
    Odessey blocking buffer as blocking agent for 1 hour(s) and 0 minute(s) · Concentration: 50% · Temperature: 25°C
    Read More
    The reviewer received a reward from Abcam’s Loyalty Program in thanks for submitting this Abreview and for helping the scientific community make better-informed decisions.

    DR. Tanya Hundal

    Verified customer

    Submitted Dec 03 2020

    Immunohistochemistry (Frozen sections) abreview for Anti-HIF-1 alpha antibody [EP1215Y]

    Good
    Abreviews
    Abreviews
    abreview image
    Application
    Immunohistochemistry (Frozen sections)
    Sample
    Human Tissue sections (human frozen skin sections)
    Permeabilization
    No
    Specification
    human frozen skin sections
    Fixative
    50:50 acetone and methonal 10mins
    Read More
    The reviewer received a reward from Abcam’s Loyalty Program in thanks for submitting this Abreview and for helping the scientific community make better-informed decisions.

    Abcam user community

    Verified customer

    Submitted Jan 30 2019

    Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) abreview for Anti-HIF-1 alpha antibody [EP1215Y]

    Inconclusive
    Abreviews
    Abreviews
    abreview image
    Application
    Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections)
    Sample
    Mouse Tissue sections (Mouse breast cancer)
    Antigen retrieval step
    Heat mediated - Buffer/Enzyme Used: Tris ph9 / citrate pH6
    Permeabilization
    Yes - Tween 20 (0.1%)
    Specification
    Mouse breast cancer
    Blocking step
    BSA as blocking agent for 2 hour(s) and 0 minute(s) · Concentration: 1% · Temperature: rt°C
    Fixative
    Formaldehyde
    Read More
    The reviewer received a reward from Abcam’s Loyalty Program in thanks for submitting this Abreview and for helping the scientific community make better-informed decisions.

    Abcam user community

    Verified customer

    Submitted Aug 10 2018

    Immunocytochemistry/ Immunofluorescence abreview for Anti-HIF-1-alpha antibody [EP1215Y]

    Inconclusive
    Abreviews
    Abreviews
    abreview image
    Application
    Immunocytochemistry/ Immunofluorescence
    Sample
    Human Cell (Human pluripotent SC derived epicardial like cells)
    Permeabilization
    Yes - Saponin
    Specification
    Human pluripotent SC derived epicardial like cells
    Blocking step
    Serum as blocking agent for 15 minute(s) · Concentration: 4% · Temperature: 4°C
    Fixative
    Paraformaldehyde
    Read More
    The reviewer received a reward from Abcam’s Loyalty Program in thanks for submitting this Abreview and for helping the scientific community make better-informed decisions.

    Abcam user community

    Verified customer

    Submitted Oct 31 2016

    Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) abreview for Anti-HIF-1-alpha antibody [EP1215Y]

    Inconclusive
    Abreviews
    Abreviews
    abreview image
    Application
    Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections)
    Sample
    Mouse Tissue sections (femur head)
    Antigen retrieval step
    Heat mediated - Buffer/Enzyme Used: 0.01 M Citrate Buffer pH6.0
    Permeabilization
    Yes - 0.1%Triton X 100
    Specification
    femur head
    Blocking step
    Serum as blocking agent for 1 hour(s) and 0 minute(s) · Concentration: 10% · Temperature: RT°C
    Fixative
    Formaldehyde
    Read More
    The reviewer received a reward from Abcam’s Loyalty Program in thanks for submitting this Abreview and for helping the scientific community make better-informed decisions.

    Abcam user community

    Verified customer

    Submitted Jun 13 2016

    Western blot abreview for Anti-HIF-1-alpha antibody [EP1215Y]

    Good
    Abreviews
    Abreviews
    abreview image
    Application
    Western blot
    Sample
    Mouse Cell lysate - whole cell (TSC cell)
    Gel Running Conditions
    Non-reduced Denaturing
    Loading amount
    60 µg
    Specification
    TSC cell
    Blocking step
    BSA as blocking agent for 1 hour(s) and 0 minute(s) · Concentration: 5% · Temperature: RT°C
    Read More
    The reviewer received a reward from Abcam’s Loyalty Program in thanks for submitting this Abreview and for helping the scientific community make better-informed decisions.

    Abcam user community

    Verified customer

    Submitted Dec 18 2015

    Western blot abreview for Anti-HIF-1-alpha antibody [EP1215Y]

    Good
    Abreviews
    Abreviews
    abreview image
    Application
    Western blot
    Sample
    Human Cell lysate - whole cell (human hepatocytes)
    Gel Running Conditions
    Reduced Denaturing
    Loading amount
    20 µg
    Treatment
    hypoxia (2% O2)
    Specification
    human hepatocytes
    Blocking step
    Milk as blocking agent for 1 hour(s) and 0 minute(s) · Concentration: 5% · Temperature: 23°C
    Read More
    The reviewer received a reward from Abcam’s Loyalty Program in thanks for submitting this Abreview and for helping the scientific community make better-informed decisions.

    Abcam user community

    Verified customer

    Submitted Sep 01 2015

    1-10 of 17 Abreviews or Q&A

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