Overview

  • Product name

  • Description

    Rabbit polyclonal to PHD3
  • Host species

    Rabbit
  • Tested applications

    Suitable for: IP, ICC/IF, WBmore details
  • Species reactivity

    Reacts with: Mouse, Rat, Human
  • Immunogen

    Synthetic peptide corresponding to Human PHD3 aa 50-150 (C terminal) conjugated to keyhole limpet haemocyanin.
    (Peptide available as ab30781)

  • Positive control

    • In WB this antibody gave a positive signal in whole cell, nuclear and cytoplasmic extracts of DFO-treated HeLa cells. In IF this antibody gave a positive signal in the following formaldehyde fixed cell line: HepG2.

Properties

Applications

Our Abpromise guarantee covers the use of ab30782 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
IP Use at an assay dependent concentration.
ICC/IF Use a concentration of 5 µg/ml.
WB Use a concentration of 1 µg/ml. Detects a band of approximately 27 kDa (predicted molecular weight: 27 kDa).

Target

  • Function

    Catalyzes the post-translational formation of 4-hydroxyproline in hypoxia-inducible factor (HIF) alpha proteins. Hydroxylates HIF-1 alpha at 'Pro-564', and HIF-2 alpha. Functions as a cellular oxygen sensor and, under normoxic conditions, targets HIF through the hydroxylation for proteasomal degradation via the von Hippel-Lindau ubiquitination complex. May play a role in cell growth regulation in muscle cells and in apoptosis in neuronal tissue. Promotes cell death through a caspase-dependent mechanism.
  • Tissue specificity

    Widely expressed at low levels. Expressed at higher levels in heart (cardiac myocytes, aortic endothelial cells and coronary artery smooth muscle) and placenta.
  • Sequence similarities

    Contains 1 Fe2OG dioxygenase domain.
  • Cellular localization

    Cytoplasm. Nucleus.
  • Information by UniProt
  • Database links

  • Alternative names

    • Egl 9 family hypoxia inducible factor 3 antibody
    • Egl nine homolog 3 (C. elegans) antibody
    • Egl nine homolog 3 antibody
    • Egl nine like protein 3 isoform antibody
    • EGL9 homolog of C. elegans 3 antibody
    • EGLN3 antibody
    • EGLN3_HUMAN antibody
    • Factor responsive smooth muscle protein antibody
    • HIF Prolyl Hydroxylase 3 antibody
    • HIF-PH3 antibody
    • HIF-prolyl hydroxylase 3 antibody
    • HIFP4H3 antibody
    • HIFPH3 antibody
    • HPH-1 antibody
    • HPH-3 antibody
    • Hypoxia-inducible factor prolyl hydroxylase 3 antibody
    • P4H3 antibody
    • PHD3 antibody
    • Prolyl Hydroxylase Domain Containing Protein 3 antibody
    • Prolyl hydroxylase domain-containing protein 3 antibody
    • SM20 antibody
    see all

Images

  • All lanes : Anti-PHD3 antibody (ab30782) at 1 µg/ml

    Lane 1 : Hela-Vehicle treated (Negative Control) Whole Cell Lysate (ab116321)
    Lane 2 : Hela-DFO treated (0.5mM, 24h) Whole Cell Lysate (ab116322)
    Lane 3 : Human HeLa Nuclear DFO treated
    Lane 4 : Human HeLa Cytoplasmic DFO treated

    Lysates/proteins at 20 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: 27 kDa
    Observed band size: 27 kDa
    Additional bands at: 18 kDa (possible non-specific binding)


    Exposure time: 20 minutes


    This blot was produced using a 4-12% Bis-tris gel under the MES buffer system. The gel was run at 200V for 35 minutes before being transferred onto a Nitrocellulose membrane at 30V for 70 minutes. The membrane was then blocked for an hour using 5% Bovine Serum Albumin before being incubated with ab30782 overnight at 4°C. Antibody binding was detected using an anti-rabbit antibody conjugated to HRP, and visualised using ECL development solution.

  • All lanes : Anti-PHD3 antibody (ab30782) at 1/1000 dilution

    Lane 1 : Mouse embroyonic fibroblast cell lysate. Treated 21% O2 for 24 hours
    Lane 2 : Mouse embroyonic fibroblast cell lysate. Treated 1% O2 for 24 hours

    Lysates/proteins at 5 µg per lane.

    Secondary
    All lanes : Goat polyclonal anti-rabbit HRP-conjugate at 1/10000 dilution

    Developed using the ECL technique.

    Performed under reducing conditions.

    Predicted band size: 27 kDa


    Exposure time: 30 seconds

    See Abreview

  • ab30782 immunoprecipitating PHD3 from rat PC12 (whole cell lysate - the cells were either normoxia (21% oxygen) or hypoxia (1% hypoxia). A protein A matrix was used with the antibody at a dilution of 1/1000. A non-Abcam antibody was used in the subsequent western blot. The doublet observed is specific to PHD3 - most probably a modified form or an alternately spliced form. Note: This antibody IP's a hypoxia inducible species of ~27 kDa and ~30 kDa. This is the same molecular weight as the human and mouse isoform, not the SM-20 variant with the N-terminal mitochondrial leader sequence.

    Lane 1: normoxia - input

    Lane 2: hypoxia - input

    Lane 3: normoxia - control antibody IP

    Lane 4: hypoxia - control antibody IP

    Lane 5: normoxia - ab30782 IP

    Lane 6: hypoxia - ab30782 IP

    See Abreview

  • ICC/IF image of ab30782 stained HepG2 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 ab30782 at 5µg/ml overnight at +4°C. The secondary antibody (green) was DyLight® 488 goat anti- rabbit (ab96899) 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.

References

This product has been referenced in:

  • Li D  et al. Armadillo repeat containing 12 promotes neuroblastoma progression through interaction with retinoblastoma binding protein 4. Nat Commun 9:2829 (2018). Read more (PubMed: 30026490) »
  • Tang LR  et al. Prolyl hydroxylase domain 3 influences the radiotherapy efficacy of pancreatic cancer cells by targeting hypoxia-inducible factor-1a. Onco Targets Ther 11:8507-8515 (2018). Read more (PubMed: 30555241) »
See all 10 Publications for this product

Customer reviews and Q&As

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1-3 of 3 Abreviews

Application
Western blot
Sample
Mouse Cell lysate - whole cell (mouse embryonic fibroblast)
Loading amount
5 µg
Specification
mouse embryonic fibroblast
Treatment
21% O2 and 1% O2 for 24hrs
Gel Running Conditions
Reduced Denaturing (10%)
Blocking step
Milk as blocking agent for 30 minute(s) · Concentration: 3% · Temperature: 25°C

Abcam user community

Verified customer

Submitted May 17 2013

Application
Immunoprecipitation
Sample
Rat Cell lysate - whole cell (PC12)
Total protein in input
500 µg
Specification
PC12
Treatment
Normoxia and Hypoxia (1% Oxygen, 4 hrs)
Immuno-precipitation step
Protein A

Abcam user community

Verified customer

Submitted Oct 30 2007

Application
Western blot
Sample
Rat Cell lysate - whole cell (PC12)
Loading amount
40 µg
Specification
PC12
Treatment
Normoxia and Hypoxia (1% Oxygen, 4 hrs)
Gel Running Conditions
Reduced Denaturing
Blocking step
Milk as blocking agent for 45 minute(s) · Concentration: 5% · Temperature: 21°C

Abcam user community

Verified customer

Submitted Jun 20 2007

Please note: All products are "FOR RESEARCH USE ONLY. NOT FOR USE IN DIAGNOSTIC PROCEDURES"
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