Overview

  • Product name

    Anti-PKR antibody [Y117]
    See all PKR primary antibodies
  • Description

    Rabbit monoclonal [Y117] to PKR
  • Host species

    Rabbit
  • Specificity

    The antibody does not cross-react with other GCN2 family members.
  • Tested applications

    Suitable for: Flow Cyt, WB, IHC-P, ICC/IF, IPmore details
  • Species reactivity

    Reacts with: Human
  • Immunogen

    Synthetic peptide within Human PKR aa 500-600 (C terminal). The exact sequence is proprietary.

  • Positive control

    • WB: MCF-7, HEK293 and HeLa whole cell lysate (ab150035) and human liver carcinoma tissue lysate. ICC/IF: Wild type HAP1, MCF-7 and HeLa cells. IHC-P: Human liver carcinoma and colon carcinoma tissue.
  • General notes

    Mouse, Rat: We have preliminary internal testing data to indicate this antibody may not react with these species. Please contact us for more information.

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

    We are constantly working hard to ensure we provide our customers with best in class antibodies. As a result of this work we are pleased to now offer this antibody in purified format. We are in the process of updating our datasheets. The purified format is designated 'PUR' on our product labels. If you have any questions regarding this update, please contact our Scientific Support team.

    This product is a recombinant rabbit monoclonal antibody.

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

    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

    Y117
  • Isotype

    IgG
  • Research areas

Applications

Our Abpromise guarantee covers the use of ab32506 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
Flow Cyt Use at an assay dependent concentration.
WB 1/5000 - 1/20000. Detects a band of approximately 68 kDa (predicted molecular weight: 62 kDa).
IHC-P 1/100.
ICC/IF 1/100.

For unpurified use at 1/250 - 1/500

IP 1/80 - 1/100.

Target

  • Function

    Following activation by double-stranded RNA in the presence of ATP, the kinase becomes autophosphorylated and can catalyze the phosphorylation of the translation initiation factor EIF2S1, which leads to an inhibition of the initiation of protein synthesis. Double-stranded RNA is generated during the course of a viral infection.
  • Sequence similarities

    Belongs to the protein kinase superfamily. Ser/Thr protein kinase family. GCN2 subfamily.
    Contains 2 DRBM (double-stranded RNA-binding) domains.
    Contains 1 protein kinase domain.
  • Post-translational
    modifications

    Autophosphorylated on several Ser and Thr residues. Autophosphorylation of Thr-451 is dependent on Thr-446 and is stimulated by dsRNA binding and dimerization. Autophosphorylation apparently leads to the activation of the kinase.
  • Information by UniProt
  • Database links

  • Alternative names

    • Double stranded RNA activated protein kinase; antibody
    • E2AK2_HUMAN antibody
    • eIF-2A protein kinase 2 antibody
    • EIF2AK1 antibody
    • EIF2AK2 antibody
    • Eukaryotic translation initiation factor 2 alpha kinase 2 antibody
    • Eukaryotic translation initiation factor 2-alpha kinase 2 antibody
    • HGNC:9437 antibody
    • Interferon induced double stranded RNA activated protein kinase antibody
    • Interferon inducible elF2 alpha kinase antibody
    • Interferon inducible RNA dependent protein kinase antibody
    • Interferon-induced, double-stranded RNA-activated protein kinase antibody
    • Interferon-inducible RNA-dependent protein kinase antibody
    • MGC126524 antibody
    • P1/eIF-2A protein kinase antibody
    • P1/eIF2A protein kinase antibody
    • p68 kinase antibody
    • PKR antibody
    • PPP1R83 antibody
    • PRKR antibody
    • Protein kinase interferon inducible double stranded RNA dependent antibody
    • Protein kinase RNA activated antibody
    • Protein kinase RNA-activated antibody
    • Protein phosphatase 1 regulatory subunit 83 antibody
    • Serine/threonine protein kinase TIK antibody
    • Tyrosine protein kinase EIF2AK2 antibody
    see all

Images

  • Lanes 1, 3 and 5: PKR knockout HAP1 cell lysate (20 µg)
    Lanes 2, 4 and 6: Wild-type HAP1 cell lysate (20 µg)
    Lanes 1 and 2: Green signal from target - ab32506 observed at 62 kDa
    Lanes 3 and 4: Red signal from loading control - ab8245 observed at 37 kDa
    Lanes 5 and 6: Merged (red and green) signal
    ab32506 was shown to specifically react with PKR when PKR knockout samples were used. Wild-type and PKR knockout samples were subjected to SDS-PAGE. ab32506 and ab8245 (loading control to GAPDH) were diluted 1/10 000 and 1/2000 respectively and incubated overnight at 4°C. Blots were developed with Goat anti-Rabbit IgG H&L (IRDye® 800CW) preadsorbed (ab216773) and Goat anti-Mouse IgG H&L (IRDye® 680RD) preadsorbed (ab216776) secondary antibodies at 1/10 000 dilution for 1 h at room temperature before imaging.

  • ab32506 staining PKR in wild-type HAP1 cells (top panel) and PKR knockout HAP1 cells (bottom panel). The cells were fixed with 100% methanol (5min), permeabilized with 0.1% Triton X-100 for 5 minutes and then blocked with 1% BSA/10% normal goat serum/0.3M glycine in 0.1% PBS-Tween for 1h. The cells were then incubated with ab32506 at 1/400 dilution and ab7291 at 1ug/ml concentration overnight at +4°C, followed by a further incubation at room temperature for 1h with a goat secondary antibody to Rabbit IgG (Alexa Fluor® 488) (ab150081) at 2 μg/ml (shown in green) and a goat secondary antibody to Mouse IgG (Alexa Fluor® 594) (ab150117) at 2ug/ml (shown in pseudo-color red). Nuclear DNA was labelled in blue with DAPI.

    This product also gave a positive signal under the same testing conditions in HAP1 cells fixed with 4% formaldehyde (10 min).


    Image was taken with a confocal microscope (Leica-Microsystems, TCS SP8).

  • All lanes : Anti-PKR antibody [Y117] (ab32506) at 1/20000 dilution

    Lane 1 : MCF-7 (human breast carcinoma) whole cell lysates
    Lane 2 : HEK293 (human embryonic kidney) whole cell lysates

    Lysates/proteins at 20 µg per lane.

    Secondary
    All lanes : Goat Anti-Rabbit IgG H&L (HRP) (ab97051) at 1/100000 dilution

    Predicted band size: 62 kDa
    Additional bands at: 68 kDa. We are unsure as to the identity of these extra bands.



    Purified format.

  • ab32506 staining PKR in human liver carcinoma tissue sections by Immunohistochemistry (IHC-P - paraformaldehyde-fixed, paraffin-embedded sections). Tissue was fixed with paraformaldehyde and antigen retrieval was by heat mediation in a EDTA buffer. Samples were incubated with primary antibody at a dilution of 1/100. A goat anti-rabbit IgG H&L (HRP) ab97051 was used as the secondary antibody at a dilution of 1/500.

    Negative control 1: PBS in place of primary antibody.

  • Flow Cytometry analysis of MCF-7 (human breast carcinoma) cells labeling PKR with purified ab32506 at 1/20 dilution (10ug/ml) (red). Cells were fixed with 4% paraformaldehyde and permeabilised with 90% methanol. A Goat anti rabbit IgG (Alexa Fluor® 488) (1/2000 dilution) was used as the secondary antibody. Rabbit monoclonal IgG (Black) was used as the isotype control, cells without incubation with primary antibody and secondary antibody (Blue) were used as the unlabeled control.

  • Anti-PKR antibody [Y117] (ab32506) at 1/10000 dilution (unpurified) + MCF-7 cell lysate

    Predicted band size: 62 kDa
    Observed band size: 68 kDa
    why is the actual band size different from the predicted?

  • ab32506 immunoprecipitating PKR. 10µg of cell lysate was incubated with primary antibody at a dilution of 1/40 and VeriBlot for IP Detection Reagent (HRP) (ab131366) at a dilution of 1/10000.

    Lane 1: HEK293 (human embryonic kidney) whole cell lysate (10ug)
    Lane 2: HEK293 (human embryonic kidney) whole cell lysate
    Lane 3: Rabbit monoclonal IgG (ab172730) instead of ab32506 in HEK293 (human embryonic kidney) whole cell lysate

  • Immunohistochemical analysis of paraffin-embedded human colon carcinoma using unpurified ab32506 at 1/100 dilution.

  • ab32506 staining PKR in MCF-7 (human breast carcinoma) cells by ICC/IF (Immunocytochemistry/immunofluorescence). Cells were fixed with 4% Paraformaldehyde and permeabilized with 0.1% Triton X-100. Samples were incubated with primary antibody at a dilution of 1/100. A goat anti rabbit IgG (Alexa Fluor® 488) (ab150077) was used as the secondary antibody. ab7291 and ab150120 were used as counterstains for primary antibody ab32506 and secondary antibody ab150077 respectively and DAPI was used as a nuclear counterstain.

    Negative control 1: Rabbit primary antibody and anti-mouse secondary antibody (ab150120)
    Negative control 2: Mouse primary antibody (ab7291) and anti-rabbit secondary antibody (ab150077)

  • Immunofluorescent staining of HeLa cells using unpurified ab32506 at 1/250 dilution.

References

This product has been referenced in:

  • Suzuki S  et al. Dental pulp cell-derived powerful inducer of TNF-a comprises PKR containing stress granule rich microvesicles. Sci Rep 9:3825 (2019). Read more (PubMed: 30846715) »
  • Liu G & Zhang W Long non-coding RNA HOTAIR promotes UVB-induced apoptosis and inflammatory injury by up-regulation of PKR in keratinocytes. Braz J Med Biol Res 51:e6896 (2018). Read more (PubMed: 29898032) »
See all 11 Publications for this product

Customer reviews and Q&As

1-4 of 4 Abreviews or Q&A

Application
Western blot
Sample
Human Cell lysate - whole cell (HEK293)
Loading amount
20 µg
Specification
HEK293
Gel Running Conditions
Non-reduced Denaturing (10% gel)
Blocking step
Milk as blocking agent for 1 hour(s) and 0 minute(s) · Concentration: 5% · Temperature: 22°C

Ms. Samantha Johnston

Verified customer

Submitted Sep 27 2011

Application
Western blot
Sample
Human Cell lysate - whole cell (HeLa Cells)
Loading amount
20 µg
Specification
HeLa Cells
Treatment
DMSO, SiRNA Luciferase
Gel Running Conditions
Non-reduced Denaturing (10)
Blocking step
Milk as blocking agent for 45 minute(s) · Concentration: 5% · Temperature: 21°C

Abcam user community

Verified customer

Submitted Aug 23 2011

Application
Western blot
Sample
Human Cell lysate - whole cell (primary human myoblast)
Loading amount
35 µg
Specification
primary human myoblast
Gel Running Conditions
Reduced Denaturing (4-12%)
Blocking step
odyssey blocking buffer as blocking agent for 1 hour(s) and 0 minute(s) · Concentration: 100% · Temperature: RT°C

Abcam user community

Verified customer

Submitted Nov 23 2010

Question
Answer

Thank you for your enquiry. I can confirm that the concentration of this product is 0.157 mg/ml. Should you require any other information, please do not hesitate to contact me.

Read More

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