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  1. Link

    hif-1-alpha-antibody-chip-grade-ab2185.pdf

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Cardiovascular Hypoxia HIF
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Anti-HIF-1 alpha antibody - ChIP Grade (ab2185)

  • Datasheet
  • SDS
Reviews (10)Q&A (34)References (123)

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  • ChIP - Anti-HIF-1 alpha antibody - ChIP Grade (ab2185)
  • Immunocytochemistry/ Immunofluorescence - Anti-HIF-1 alpha antibody - ChIP Grade (ab2185)
  • Western blot - Anti-HIF-1 alpha antibody - ChIP Grade (ab2185)
  • Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-HIF-1 alpha antibody - ChIP Grade (ab2185)
  • Immunoprecipitation - Anti-HIF-1 alpha antibody - ChIP Grade (ab2185)

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  • Datasheet
  • References (123)
  • Protocols

Overview

  • Product name

    Anti-HIF-1 alpha antibody - ChIP Grade
    See all HIF-1 alpha primary antibodies
  • Description

    Rabbit polyclonal to HIF-1 alpha - ChIP Grade
  • Host species

    Rabbit
  • Tested applications

    Suitable for: IHC-P, WB, IP, GSA, IHC-Fr, ICC, ICC/IF, ChIPmore details
  • Species reactivity

    Reacts with: Mouse, Rat, Guinea pig, Human, Xenopus laevis, Monkey
    Predicted to work with: Cow
  • Immunogen

    Fusion protein corresponding to Human HIF-1 alpha aa 432-528.

  • Positive control

    • ICC/IF: HeLa cells. IHC-P: Human kidney tissue. ChIP: HeLa cells. WB: Rat hypoxic nuclear lysate. IP: PC-3 cell lysate treated with cobalt chloride.
  • General notes

    HIF-1 alpha can be a difficult target to work with so we have compiled a summary of all the important information you need to know including useful tips. This can be found in the protocols tab or alternatively click here to download the English version and here to download the Mandarin.

    Under normoxic conditions HIF-1 alpha has a short half-life. It is largely undetectable in cells or tissues grown under normoxic conditions. It is stabilized only at O2 concentrations below 5% and upon stabilization under hypoxic conditions HIF-1 translocates to the nucleus. Therefore we recommend western blots using nuclear extracts and running Hypoxia treated samples as positive control (ab180880). Hypoxia can be induced with treatment using certain agents e.g. CoCl2 or DFO, etc. so proper sample preparation is critical.

Properties

  • Form

    Liquid
  • Storage instructions

    Shipped at 4°C. Store at +4°C short term (1-2 weeks). Upon delivery aliquot. Store at -20°C. Avoid freeze / thaw cycle.
  • Storage buffer

    Preservative: 0.05% Sodium azide
    Constituent: PBS
  • Concentration information loading...
  • Purity

    Immunogen affinity purified
  • Clonality

    Polyclonal
  • 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
    • Epigenetics and Nuclear Signaling
    • ChIP assays
    • ChIP antibodies
    • 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

Associated products

  • Assay kits

    • HIF-1 alpha Transcription Factor Assay Kit (ab133104)
  • ChIP Related Products

    • Rabbit Anti-Mouse IgG H&L (ab46540)
    • ChIP Kit (ab500)
  • Compatible Secondaries

    • Goat Anti-Rabbit IgG H&L (Alexa Fluor® 488) (ab150077)
    • Goat Anti-Rabbit IgG H&L (HRP) (ab205718)
  • Isotype control

    • Rabbit IgG, polyclonal - Isotype Control (ChIP Grade) (ab171870)
  • 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)
    • Hela-Vehicle treated (Negative Control) Whole Cell Lysate (ab116321)
    • Hela-DFO treated (0.5mM, 24h) Whole Cell Lysate (ab116322)
    • Human HIF-1 alpha ELISA Kit (ab171577)

Applications

Our Abpromise guarantee covers the use of ab2185 in the following tested applications.

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

Application Abreviews Notes
IHC-P 1/100.
WB 1/500 - 1/1000.
IP 1/1000.
GSA Use at an assay dependent dilution.
IHC-Fr 1/10 - 1/2000.
ICC Use at an assay dependent concentration.
ICC/IF 1/10 - 1/2000.
EMSA 1/1 - 1/100.
ChIP Use at an assay dependent concentration.

25 µl / 15 millions cells

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: 281814 Cow
    • Entrez Gene: 3091 Human
    • Entrez Gene: 15251 Mouse
    • Entrez Gene: 29560 Rat
    • Entrez Gene: 100337573 Xenopus laevis
    • Entrez Gene: 380141 Xenopus laevis
    • Omim: 603348 Human
    • SwissProt: Q16665 Human
    • SwissProt: Q61221 Mouse
    • SwissProt: O35800 Rat
    • SwissProt: Q9I8A9 Xenopus laevis
    • Unigene: 597216 Human
    • Unigene: 3879 Mouse
    • Unigene: 446610 Mouse
    • Unigene: 10852 Rat
    • Unigene: 4589 Xenopus laevis
    see all
  • 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

  • ChIP - Anti-HIF-1 alpha antibody - ChIP Grade (ab2185)
    ChIP - Anti-HIF-1 alpha antibody - ChIP Grade (ab2185)This image is courtesy of a review submitted by Yolanda Cuevas, Hospital La Princesa.

    ChIP analysis of HIF-1-alpha genomic sequences from HeLa cells cultivated in normoxic (N) or hypoxic (Hx) conditions, using a HIF1-alpha polyclonal antibody (ab2185). For a negative control, IgG was used and the input as a positive control in the subsequent PCR. Primers for known target genes were used HIF1 alpha, A. EPO and B. VEGF.

  • Immunocytochemistry/ Immunofluorescence - Anti-HIF-1 alpha antibody - ChIP Grade (ab2185)
    Immunocytochemistry/ Immunofluorescence - Anti-HIF-1 alpha antibody - ChIP Grade (ab2185)

    ICC/IF image of ab2185 stained HeLa cells. The cells were 4% formaldehyde fixed (10 min) and then incubated in 1%BSA / 10% normal goat serum / 0.3M glycine in 0.1% PBS-Tween for 1h to permeabilise the cells and block non-specific protein-protein interactions. The cells were then incubated with the antibody (ab2185, 1/1000 dilution) overnight at +4°C. The secondary antibody (green) was Alexa Fluor® 488 goat anti-rabbit IgG (H+L) used at a 1/1000 dilution for 1h. Alexa Fluor® 594 WGA was used to label plasma membranes (red) at a 1/200 dilution for 1h. DAPI was used to stain the cell nuclei (blue) at a concentration of 1.43µM.

  • Western blot - Anti-HIF-1 alpha antibody - ChIP Grade (ab2185)
    Western blot - Anti-HIF-1 alpha antibody - ChIP Grade (ab2185)
    All lanes : Anti-HIF-1 alpha antibody - ChIP Grade (ab2185)

    Lane 1 : Rat nuclear extract lysate - normoxic
    Lane 2 : Rat nuclear extract lysate - hypoxic
  • Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-HIF-1 alpha antibody - ChIP Grade (ab2185)
    Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-HIF-1 alpha antibody - ChIP Grade (ab2185)

    Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) analysis of Human kidney tissue sections labeling HIF-1-alpha with ab2185.

  • Immunoprecipitation - Anti-HIF-1 alpha antibody - ChIP Grade (ab2185)
    Immunoprecipitation - Anti-HIF-1 alpha antibody - ChIP Grade (ab2185)
    All lanes : Anti-HIF-1 alpha antibody - ChIP Grade (ab2185)

    Lane 1 : PC-3 cell lysates - untreated
    Lane 2 : PC-3 cell lysates - treated with 1µl cobalt chloride
    Lane 3 : PC-3 cell lysates - treated with 3µl cobalt chloride
    Lane 4 : PC-3 cell lysates - treated with 5µl cobalt chloride

Protocols

  • Nuclear Extract Preparation for HIF 1 alpha
  • Protocol Booklet - HIF 1 alpha - useful tips and information

Datasheets and documents

    • Datasheet
    • SDS
  • References

    This product has been referenced in:

    • Chen X  et al. Copper promotes the migration of bone marrow mesenchymal stem cells via Rnd3-dependent cytoskeleton remodeling. J Cell Physiol 235:221-231 (2020). Read more (PubMed: 31187497) »
    • Liu FY  et al. TLR9 is essential for HMGB1-mediated post-myocardial infarction tissue repair through affecting apoptosis, cardiac healing, and angiogenesis. Cell Death Dis 10:480 (2019). Read more (PubMed: 31209243) »
    See all 123 Publications for this product

    Publishing research using ab2185? Please let us know so that we can cite the reference in this datasheet.

    Customer reviews and Q&As

    Show All Reviews Q&A
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    1-10 of 10 Abreviews

    Immunocytochemistry/ Immunofluorescence abreview for Anti-HIF-1 alpha antibody - ChIP Grade

    Excellent
    Abreviews
    Abreviews
    abreview image
    Application
    Immunocytochemistry/ Immunofluorescence
    Sample
    Human Cell (HEK293)
    Permeabilization
    Yes - 0.1% Triton X100
    Specification
    HEK293
    Blocking step
    Serum as blocking agent for 2 hour(s) and 0 minute(s) · Concentration: 5% · Temperature: 25°C
    Fixative
    Formaldehyde
    Read More

    Abcam user community

    Verified customer

    Submitted Jul 18 2017

    Western blot abreview for Anti-HIF-1 alpha antibody - ChIP Grade

    Inconclusive
    Abreviews
    Abreviews
    abreview image
    Application
    Western blot
    Sample
    Notothenia coriiceps Cell lysate - other (heart)
    Gel Running Conditions
    Reduced Denaturing
    Loading amount
    50 µg
    Treatment
    CTmax
    Specification
    heart
    Blocking step
    Milk as blocking agent for 10 hour(s) and 0 minute(s) · Concentration: 0.1% · Temperature: 4°C
    Read More

    Abcam user community

    Verified customer

    Submitted Jan 13 2017

    Western blot abreview for Anti-HIF-1 alpha antibody - ChIP Grade

    Inconclusive
    Abreviews
    Abreviews
    abreview image
    Application
    Western blot
    Sample
    Chaenocephalus aceratus Cell lysate - other (heart)
    Gel Running Conditions
    Reduced Denaturing
    Loading amount
    50 µg
    Treatment
    CTmax
    Specification
    heart
    Blocking step
    BSA as blocking agent for 10 hour(s) and 0 minute(s) · Concentration: 0.1% · Temperature: 4°C
    Read More

    Abcam user community

    Verified customer

    Submitted Jan 13 2017

    Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) abreview for Anti-HIF-1-alpha antibody - ChIP Grade

    Excellent
    Abreviews
    Abreviews
    abreview image
    Application
    Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections)
    Sample
    Human Tissue sections (necrotic melanoma xenograft)
    Antigen retrieval step
    Heat mediated - Buffer/Enzyme Used: Tris/EDTA pH 9
    Permeabilization
    No
    Specification
    necrotic melanoma xenograft
    Blocking step
    Serum as blocking agent for 1 hour(s) and 0 minute(s) · Concentration: 15% · Temperature: 20°C
    Fixative
    Formaldehyde
    Read More

    Abcam user community

    Verified customer

    Submitted Dec 18 2015

    Immunocytochemistry

    Good
    Abreviews
    Abreviews
    abreview image
    Application
    Immunocytochemistry
    Blocking step
    BSA as blocking agent for 1 hour(s) and 0 minute(s) · Concentration: 1% · Temperature: 25°C
    Sample
    Mouse Cultured Cells (Neuro2A- neuroblastoma)
    Specification
    Neuro2A- neuroblastoma
    Permeabilization
    Yes - 0.3% Triton X-100
    Fixative
    Paraformaldehyde
    Read More

    Abcam user community

    Verified customer

    Submitted Jun 04 2013

    Western blot abreview for Anti-HIF1 alpha antibody - ChIP Grade

    Good
    Abreviews
    Abreviews
    abreview image
    Application
    Western blot
    Sample
    Mouse Tissue lysate - whole (Striatum (Brain))
    Loading amount
    50 µg
    Specification
    Striatum (Brain)
    Gel Running Conditions
    Reduced Denaturing (8%)
    Blocking step
    BSA as blocking agent for 1 hour(s) and 0 minute(s) · Concentration: 2% · Temperature: 25°C
    Read More

    Abcam user community

    Verified customer

    Submitted Feb 14 2013

    Immunocytochemistry/ Immunofluorescence abreview for Anti-HIF1 alpha antibody - ChIP Grade

    Good
    Abreviews
    Abreviews
    abreview image
    Application
    Immunocytochemistry/ Immunofluorescence
    Sample
    Mouse Cell (sarcoma (MFH) subcutaneous xenografts)
    Specification
    sarcoma (MFH) subcutaneous xenografts
    Fixative
    Paraformaldehyde
    Permeabilization
    Yes - 0.1% Triton-X100 in PBS
    Blocking step
    Serum as blocking agent for 2 hour(s) and 0 minute(s) · Concentration: 5% · Temperature: 20°C
    Read More

    Tatiana Karakasheva

    Verified customer

    Submitted Jan 11 2012

    Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) abreview for Anti-HIF1 alpha antibody - ChIP Grade

    Good
    Abreviews
    Abreviews
    abreview image
    Application
    Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections)
    Sample
    Mouse Tissue sections (sarcoma (MFH) subcutaneous xenografts)
    Specification
    sarcoma (MFH) subcutaneous xenografts
    Fixative
    Paraformaldehyde
    Antigen retrieval step
    Heat mediated - Buffer/Enzyme Used: citric acid
    Permeabilization
    Yes - 0.1% Triton-X100 in PBS
    Blocking step
    Serum as blocking agent for 2 hour(s) and 0 minute(s) · Concentration: 5% · Temperature: 20°C
    Read More

    Tatiana Karakasheva

    Verified customer

    Submitted Jan 11 2012

    ChIP abreview for Anti-HIF1 alpha antibody - ChIP Grade

    Excellent
    Abreviews
    Abreviews
    abreview image
    Application
    ChIP
    Sample
    Human Cell lysate - nuclear (Glioma cell line)
    Specification
    Glioma cell line
    Type
    Cross-linking (X-ChIP)
    Duration of cross-linking step: 15 minute(s) and 0 second(s)
    Specification of the cross-linking agent: Formaldehyde
    Detection step
    Real-time PCR
    Positive control
    Region in the EGLN3 gene
    Negative control
    Region on chromosome 12 (Untr12) that is far from any known gene annotation and not expected to be bound by HIF1 alpha
    Read More

    Mr. Genpathway Inc

    Verified customer

    Submitted Apr 21 2009

    Immunoprecipitation abreview for Anti-HIF1 alpha antibody - ChIP Grade

    Average
    Abreviews
    Abreviews
    Application
    Immunoprecipitation
    Sample
    Human Cell lysate - whole cell (A549)
    Total protein in input
    8e+006 cells
    Specification
    A549
    Immuno-precipitation step
    Protein A
    Read More

    Abcam user community

    Verified customer

    Submitted May 14 2007

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