Recombinant Anti-PKR antibody [EPR19374] - BSA and Azide free (ab224887)
Key features and details
- Produced recombinantly (animal-free) for high batch-to-batch consistency and long term security of supply
- Rabbit monoclonal [EPR19374] to PKR - BSA and Azide free
- Suitable for: Flow Cyt (Intra), WB, ICC/IF, IP
- Knockout validated
- Reacts with: Mouse, Rat, Human
Related conjugates and formulations
Overview
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Product name
Anti-PKR antibody [EPR19374] - BSA and Azide free
See all PKR primary antibodies -
Description
Rabbit monoclonal [EPR19374] to PKR - BSA and Azide free -
Host species
Rabbit -
Tested applications
Suitable for: Flow Cyt (Intra), WB, ICC/IF, IPmore details -
Species reactivity
Reacts with: Mouse, Rat, Human -
Immunogen
Recombinant fragment. This information is proprietary to Abcam and/or its suppliers.
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Positive control
- WB: A459, K562, HeLa, Jurkat, 4T1, MCF7, HepG2 and bEnd.3 whole cell lysates. Mouse brain, cerebral cortex, hippocampus, lung, thymus and heart lysates; Rat brain, cerebral cortex, heart and spleen lysates; ICC/IF: bEnd.3 and A459 cells. Flow Cyt (Intra): bEnd.3 cells. IP: Mouse hippocampus lysate.
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General notes
ab224887 is the carrier-free version of ab184257.
Our carrier-free antibodies are typically supplied in a PBS-only formulation, purified and free of BSA, sodium azide and glycerol. The carrier-free buffer and high concentration allow for increased conjugation efficiency.
This conjugation-ready format is designed for use with fluorochromes, metal isotopes, oligonucleotides, and enzymes, which makes them ideal for antibody labelling, functional and cell-based assays, flow-based assays (e.g. mass cytometry) and Multiplex Imaging applications.
Use our conjugation kits for antibody conjugates that are ready-to-use in as little as 20 minutes with <1 minute hands-on-time and 100% antibody recovery: available for fluorescent dyes, HRP, biotin and gold.
This product is compatible with the Maxpar® Antibody Labeling Kit from Fluidigm, without the need for antibody preparation. Maxpar® is a trademark of Fluidigm Canada Inc.
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
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
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Form
Liquid -
Storage instructions
Shipped at 4°C. Store at +4°C. Do Not Freeze. -
Storage buffer
pH: 7.2
Constituent: PBS -
Carrier free
Yes -
Concentration information loading...
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Purity
Protein A purified -
Clonality
Monoclonal -
Clone number
EPR19374 -
Isotype
IgG -
Research areas
Associated products
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Alternative Versions
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Compatible Secondaries
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Conjugation kits
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Isotype control
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KO cell lines
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KO cell lysates
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Recombinant Protein
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Related Products
Applications
The Abpromise guarantee
Our Abpromise guarantee covers the use of ab224887 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 |
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Flow Cyt (Intra) |
Use at an assay dependent concentration.
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WB |
Use at an assay dependent concentration. Detects a band of approximately 58 kDa (predicted molecular weight: 58 kDa).
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ICC/IF |
Use at an assay dependent concentration.
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IP |
Use at an assay dependent concentration.
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Notes |
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Flow Cyt (Intra)
Use at an assay dependent concentration. |
WB
Use at an assay dependent concentration. Detects a band of approximately 58 kDa (predicted molecular weight: 58 kDa). |
ICC/IF
Use at an assay dependent concentration. |
IP
Use at an assay dependent concentration. |
Target
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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
modificationsAutophosphorylated 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
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Database links
- Entrez Gene: 5610 Human
- Entrez Gene: 19106 Mouse
- Entrez Gene: 54287 Rat
- Omim: 176871 Human
- SwissProt: P19525 Human
- SwissProt: Q03963 Mouse
- SwissProt: Q63184 Rat
- Unigene: 131431 Human
see all -
Alternative names
- Double stranded RNA activated protein kinase; antibody
- E2AK2_HUMAN antibody
- eIF-2A protein kinase 2 antibody
see all
Images
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All lanes : Anti-PKR antibody [EPR19374] (ab184257) at 1/1000 dilution
Lane 1 : Wild-type HeLa (Human epithelial cell line from cervix adenocarcinoma) whole cell lysate
Lane 2 : EIF2AK2 knockout HeLa (Human epithelial cell line from cervix adenocarcinoma) whole cell lysate
Lane 3 : Wild-type A549 (Human lung carcinoma cell line) whole cell lysate
Lane 4 : EIF2AK2 knockout A549 (Human lung carcinoma cell line) whole cell lysate
Lane 5 : K562 (Human chronic myelogenous leukemia lymphoblast cell line) whole cell lysate
Lysates/proteins at 20 µg per lane.
Secondary
All lanes : Goat anti-Rabbit IgG H&L (IRDye® 800CW) preadsorbed (ab216773) at 1/10000 dilution
Predicted band size: 58 kDa
Observed band size: 70 kDa why is the actual band size different from the predicted?This data was developed using ab184257, the same antibody clone in a different buffer formulation.
Lanes 1-5: Merged signal (red and green). Green - ab184257 observed at 70 kDa. Red - loading control ab8245 observed at 36 kDa.
ab184257 Anti-PKR antibody [EPR19374] was shown to specifically react with PKR in wild-type HeLa cells. Loss of signal was observed when knockout cell line ab261824 (knockout cell lysate ab256899) was used. Wild-type and PKR knockout samples were subjected to SDS-PAGE. ab184257 and Anti-GAPDH antibody [6C5] - Loading Control (ab8245) were incubated overnight at 4°C at 1 in 1000 dilution and 1 in 20000 dilution respectively. 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 in 20000 dilution for 1 hour at room temperature before imaging.
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All lanes : Anti-PKR antibody [EPR19374] (ab184257) at 1/1000 dilution
Lane 1 : Wild-type A549 cell lysate
Lane 2 : EIF2AK2 knockout A549 cell lysate
Lane 3 : K-562 cell lysate
Lysates/proteins at 20 µg per lane.
Secondary
All lanes : Goat anti-Rabbit IgG H&L (IRDye® 800CW) preadsorbed (ab216773) at 1/10000 dilution
Predicted band size: 58 kDa
Observed band size: 70 kDa why is the actual band size different from the predicted?This data was developed using the same antibody clone in a different buffer formulation (ab184257).
Lanes 1-3: Merged signal (red and green). Green - ab184257 observed at 70 kDa. Red - loading control ab8245 observed at 36 kDa.
ab184257 Anti-PKR antibody [EPR19374] was shown to specifically react with PKR in wild-type A549 cells. Loss of signal was observed when knockout cell line ab266999 (knockout cell lysate ab256900) was used. Wild-type and PKR knockout samples were subjected to SDS-PAGE. ab184257 and Anti-GAPDH antibody [6C5] - Loading Control (ab8245) were incubated at room temperature for 2. 5 hours at 1 in 1000 dilution and 1 in 20000 dilution respectively. 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 in 20000 dilution for 1 hour at room temperature before imaging.
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Anti-PKR antibody [EPR19374] (ab184257) at 1/1000 dilution + bEnd.3 (mouse brain endothelial cell) whole cell lysate at 20 µg
Secondary
Goat Anti-Rabbit IgG H&L (HRP) (ab97051) at 1/100000 dilution
Predicted band size: 58 kDa
Exposure time: 114 secondsThis data was developed using ab184257, the same antibody clone in a different buffer formulation.
Blocking and diluting buffer: 5% NFDM/TBST
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This data was developed using ab184257, the same antibody clone in a different buffer formulation.
Immunofluorescent analysis of 4% paraformaldehyde-fixed, 0.1% Triton X-100 permeabilized bEnd.3 (mouse brain endothelial cell) labeling PKR with ab184257 at 1/50 dilution, followed by Goat Anti-Rabbit IgG H&L (Alexa Fluor® 488) preabsorbed (ab150081) secondary antibody at 1/1000 dilution (green).
Anti-alpha Tubulin mouse monoclonal antibody - Microtubule Marker (Alexa Fluor® 594) (ab195889) at 1/200 was used as a counterstain
The nuclear counterstain is DAPI (blue)
Confocal image showing cytoplasmic and weak nuclear staining in bEnd.3 cell line.
Image was taken with a confocal microscope(Leica-Microsystems, TCS SP8).
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This data was developed using ab184257, the same antibody clone in a different buffer formulation.
Immunofluorescent analysis of 4% paraformaldehyde-fixed, 0.1% Triton X-100 permeabilized A549 (human lung carcinoma epithelial cell) labeling PKR with ab184257 at 1/50 dilution, followed by Goat Anti-Rabbit IgG H&L (Alexa Fluor® 488) preabsorbed (ab150081) secondary antibody at 1/1000 dilution (green).
Anti-alpha Tubulin mouse monoclonal antibody - Microtubule Marker (Alexa Fluor® 594) (ab195889) at 1/200 was used as a counterstain
The nuclear counterstain is DAPI (blue)
Confocal image showing cytoplasmic and weak nuclear staining in A549 cell line.
Image was taken with a confocal microscope(Leica-Microsystems, TCS SP8).
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This data was developed using ab184257, the same antibody clone in a different buffer formulation.
Intracellular flow cytometric analysis of 4% paraformaldehyde-fixed, 90% methanol permeabilised bEnd.3 (mouse brain endothelial cell) cells labeling PKR with ab184257 at 1/500 dilution (red) compared with a Rabbit monoclonal IgG isotype control (ab172730; black) and an unlabelled control (cells without incubation with primary antibody and secondary antibody; blue). Goat Anti-Rabbit IgG (Alexa Fluor® 488, ab150081) at 1/5000 dilution was used as the secondary antibody.
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This data was developed using ab184257, the same antibody clone in a different buffer formulation.
PKR was immunoprecipitated from mouse hippocampus lysate with ab184257 at 1/30 dilution (2μg in 0.35mg lysates).
Western blot was performed from the immunoprecipitate using ab184257 at 1/1000 dilution.
VeriBlot for IP Detection Reagent (HRP) (ab131366), was used for detection at 1/5000 dilution.
Lane 1: Mouse hippocampus tissue lysate 10 μg (Input).
Lane 2: ab184257 IP in Mouse hippocampus tissue lysate.
Lane 3: Rabbit IgG,monoclonal [EPR19374] - Isotype Control (ab172730) instead of ab184257 in mouse hippocampus tissue lysate.
Blocking and dilution buffer and concentration: 5% NFDM/TBST.
Exposure time: 31 seconds.
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.
Datasheets and documents
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Datasheet download
Certificate of Compliance
References (0)
ab224887 has not yet been referenced specifically in any publications.