Name: Anti-PKR antibody [Y117]
Brand: RabMAb
SKU: ab32506
Rating: 4 (3 reviews)

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Western blot - Anti-PKR antibody [Y117] (ab32506)

Key features and details

Produced recombinantly (animal-free) for high batch-to-batch consistency and long term security of supply
Rabbit monoclonal [Y117] to PKR
Suitable for: Flow Cyt (Intra), WB, IHC-P, ICC/IF, IP
Knockout validated
Reacts with: Human

Alexa Fluor® 488 Alexa Fluor® 647 APC Carrier Free HRP PE

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

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 (glycerin, glycerine), 0.05% BSA
Concentration information loading...
Purity

Protein A purified
Clonality

Monoclonal
Clone number

Y117
Isotype

IgG
Research areas

Alternative Versions
Compatible Secondaries
- Goat anti-Rabbit IgG H&L (IRDye® 800CW) preadsorbed (ab216773)
Isotype control
- Rabbit IgG, monoclonal [EPR25A] - Isotype Control (ab172730)
Positive Controls
- HEK-293 whole cell lysate (ab7902)
Recombinant Protein
- Recombinant human PKR protein (ab71666)

The Abpromise guarantee

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 (Intra)		Use at an assay dependent concentration.
WB	(3)	1/5000 - 1/20000. Detects a band of approximately 68 kDa (predicted molecular weight: 62 kDa).
IHC-P		1/100. Perform heat mediated antigen retrieval with Tris/EDTA buffer pH 9.0 before commencing with IHC staining protocol.
ICC/IF		1/100. For unpurified use at 1/250 - 1/500
IP		1/80 - 1/100.

Notes
Flow Cyt (Intra) 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. Perform heat mediated antigen retrieval with Tris/EDTA buffer pH 9.0 before commencing with IHC staining protocol.
ICC/IF 1/100. For unpurified use at 1/250 - 1/500
IP 1/80 - 1/100.

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.
Target information above from: UniProt accession P19525 The UniProt Consortium
The Universal Protein Resource (UniProt) in 2010
Nucleic Acids Res. 38:D142-D148 (2010) .

Information by UniProt
Database links
- Entrez Gene: 5610 Human
- Omim: 176871 Human
- SwissProt: P19525 Human
- Unigene: 131431 Human
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

Western blot - Anti-PKR antibody [Y117] (ab32506)

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.
Immunocytochemistry/ Immunofluorescence - Anti-PKR antibody [Y117] (ab32506)

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).
Western blot - Anti-PKR antibody [Y117] (ab32506)

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.
Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-PKR antibody [Y117] (ab32506)

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 (Intracellular) - Anti-PKR antibody [Y117] (ab32506)

Intracellular 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.
Western blot - Anti-PKR antibody [Y117] (ab32506)

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?
Immunoprecipitation - Anti-PKR antibody [Y117] (ab32506)

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
Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-PKR antibody [Y117] (ab32506)

Immunohistochemical analysis of paraffin-embedded human colon carcinoma using unpurified ab32506 at 1/100 dilution.
Immunocytochemistry/ Immunofluorescence - Anti-PKR antibody [Y117] (ab32506)

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)
Immunocytochemistry/ Immunofluorescence - Anti-PKR antibody [Y117] (ab32506)

Immunofluorescent staining of HeLa cells using unpurified ab32506 at 1/250 dilution.
Anti-PKR antibody [Y117] (ab32506)

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

SDS download

Country/Region

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Datasheet download

Download

Publishing research using ab32506? Please let us know so that we can cite the reference in this datasheet.

ab32506 has been referenced in 23 publications.

McFadden MJ et al. Post-transcriptional regulation of antiviral gene expression by N6-methyladenosine. Cell Rep 34:108798 (2021). PubMed: 33657363
Tusi SK et al. The alternative initiation factor eIF2A plays key role in RAN translation of myotonic dystrophy type 2 CCUG•CAGG repeats. Hum Mol Genet 30:1020-1029 (2021). PubMed: 33856033
Montavon TC et al. Human DICER helicase domain recruits PKR and modulates its antiviral activity. PLoS Pathog 17:e1009549 (2021). PubMed: 33984068
Piazzi M et al. Expression of the double-stranded RNA-dependent kinase PKR influences osteosarcoma attachment independent growth, migration, and invasion. J Cell Physiol 235:1103-1119 (2020). PubMed: 31240713
Cheng J et al. Typical Stress Granule Proteins Interact with the 3' Untranslated Region of Enterovirus D68 To Inhibit Viral Replication. J Virol 94:N/A (2020). PubMed: 31941779

View all Publications for this product

Customer reviews and Q&As

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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

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. 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

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 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

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 Nov 23 2010

Question

Please advise what is the concentration of this ab ( ug/ul)?

Abcam community

Verified customer

Asked on Aug 29 2007

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.

Abcam Scientific Support

Answered on Aug 29 2007

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

Related conjugates and formulations

Overview

Product name

Description

Host species

Specificity

Tested applications

Species reactivity

Immunogen

Positive control

General notes

Properties

Form

Storage instructions

Storage buffer

Purity

Clonality

Clone number

Isotype

Research areas

Associated products

Alternative Versions

Compatible Secondaries

Isotype control

Positive Controls

Recombinant Protein

Applications

The Abpromise guarantee

Target

Function

Sequence similarities

Post-translationalmodifications

Database links

Alternative names

Images

Protocols

Datasheets and documents

SDS download

Datasheet download

References (23)

Customer reviews and Q&As

Western blot abreview for Anti-PKR antibody [Y117]

Western blot abreview for Anti-PKR antibody [Y117]

Western blot abreview for Anti-PKR antibody [Y117]

Post-translational
modifications