Recombinant
RabMAb

Anti-PKC (phospho T514) antibody [EP2730Y] (ab109539)

Overview

  • Product name
    Anti-PKC (phospho T514) antibody [EP2730Y]
    See all PKC primary antibodies
  • Description
    Rabbit monoclonal [EP2730Y] to PKC (phospho T514)
  • Host species
    Rabbit
  • Tested applications
    Suitable for: WB, IP, IHC-Pmore details
  • Species reactivity
    Reacts with: Mouse, Rat, Human
  • Immunogen

    Synthetic peptide corresponding to Human PKC (phospho T514).
    Database link: P05129

  • Positive control
    • HeLa cell lysate; Human brain tissue; Mouse cerebellum; Rat brain; Human cerebral cortex.
  • General notes

    A trial size is available to purchase for this antibody.

    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

Applications

Our Abpromise guarantee covers the use of ab109539 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
WB 1/1000 - 1/10000. Predicted molecular weight: 78 kDa.
IP 1/20.

For unpurified use at 1/50.

IHC-P 1/300. Perform heat mediated antigen retrieval with citrate buffer pH 6 before commencing with IHC staining protocol.

For unpurified use at 1/100 - 1/250. 

Target

  • Function
    Calcium-activated, phospholipid- and diacylglycerol (DAG)-dependent serine/threonine-protein kinase that is involved in positive and negative regulation of cell proliferation, apoptosis, differentiation, migration and adhesion, tumorigenesis, cardiac hypertrophy, angiogenesis, platelet function and inflammation, by directly phosphorylating targets such as RAF1, BCL2, CSPG4, TNNT2/CTNT, or activating signaling cascade involving MAPK1/3 (ERK1/2) and RAP1GAP. Involved in cell proliferation and cell growth arrest by positive and negative regulation of the cell cycle. Can promote cell growth by phosphorylating and activating RAF1, which mediates the activation of the MAPK/ERK signaling cascade, and/or by up-regulating CDKN1A, which facilitates active cyclin-dependent kinase (CDK) complex formation in glioma cells. In intestinal cells stimulated by the phorbol ester PMA, can trigger a cell cycle arrest program which is associated with the accumulation of the hyper-phosphorylated growth-suppressive form of RB1 and induction of the CDK inhibitors CDKN1A and CDKN1B. Exhibits anti-apoptotic function in glioma cells and protects them from apoptosis by suppressing the p53/TP53-mediated activation of IGFBP3, and in leukemia cells mediates anti-apoptotic action by phosphorylating BCL2. During macrophage differentiation induced by macrophage colony-stimulating factor (CSF1), is translocated to the nucleus and is associated with macrophage development. After wounding, translocates from focal contacts to lamellipodia and participates in the modulation of desmosomal adhesion. Plays a role in cell motility by phosphorylating CSPG4, which induces association of CSPG4 with extensive lamellipodia at the cell periphery and polarization of the cell accompanied by increases in cell motility. Is highly expressed in a number of cancer cells where it can act as a tumor promoter and is implicated in malignant phenotypes of several tumors such as gliomas and breast cancers. Negatively regulates myocardial contractility and positively regulates angiogenesis, platelet aggregation and thrombus formation in arteries. Mediates hypertrophic growth of neonatal cardiomyocytes, in part through a MAPK1/3 (ERK1/2)-dependent signaling pathway, and upon PMA treatment, is required to induce cardiomyocyte hypertrophy up to heart failure and death, by increasing protein synthesis, protein-DNA ratio and cell surface area. Regulates cardiomyocyte function by phosphorylating cardiac troponin T (TNNT2/CTNT), which induces significant reduction in actomyosin ATPase activity, myofilament calcium sensitivity and myocardial contractility. In angiogenesis, is required for full endothelial cell migration, adhesion to vitronectin (VTN), and vascular endothelial growth factor A (VEGFA)-dependent regulation of kinase activation and vascular tube formation. Involved in the stabilization of VEGFA mRNA at post-transcriptional level and mediates VEGFA-induced cell proliferation. In the regulation of calcium-induced platelet aggregation, mediates signals from the CD36/GP4 receptor for granule release, and activates the integrin heterodimer ITGA2B-ITGB3 through the RAP1GAP pathway for adhesion. During response to lipopolysaccharides (LPS), may regulate selective LPS-induced macrophage functions involved in host defense and inflammation. But in some inflammatory responses, may negatively regulate NF-kappa-B-induced genes, through IL1A-dependent induction of NF-kappa-B inhibitor alpha (NFKBIA/IKBA). Upon stimulation with 12-O-tetradecanoylphorbol-13-acetate (TPA), phosphorylates EIF4G1, which modulates EIF4G1 binding to MKNK1 and may be involved in the regulation of EIF4E phosphorylation. Phosphorylates KIT, leading to inhibition of KIT activity. Phosphorylates ATF2 which promotes cooperation between ATF2 and JUN, activating transcription.
  • Sequence similarities
    Belongs to the protein kinase superfamily. AGC Ser/Thr protein kinase family. PKC subfamily.
    Contains 1 AGC-kinase C-terminal domain.
    Contains 1 C2 domain.
    Contains 2 phorbol-ester/DAG-type zinc fingers.
    Contains 1 protein kinase domain.
  • Cellular localization
    Cytoplasm. Cell membrane. Mitochondrion membrane. Nucleus.
  • Information by UniProt
  • Database links
  • Alternative names
    • KPCA_HUMAN antibody
    • PKC alpha antibody
    • PKC beta antibody
    • PKC delta antibody
    • PKC epsilon antibody
    • PKC gamma antibody
    • PKC zeta antibody
    • PKC-A antibody
    • PKC-alpha antibody
    • PKC2 antibody
    • PKCA antibody
    • PKCB antibody
    • PKCD antibody
    • PKCE antibody
    • PKCG antibody
    • PRKCA antibody
    • PRKCB antibody
    • PRKCD antibody
    • PRKCE antibody
    • PRKCG antibody
    • PRKCZ antibody
    • Protein kinase C alpha antibody
    • Protein kinase C alpha type antibody
    • Protein kinase C beta antibody
    • Protein kinase C delta antibody
    • Protein kinase C epsilon antibody
    • Protein kinase C gamma antibody
    • Protein kinase C zeta antibody
    see all

Images

  • ab109539 (purified) at 1:20 dilution (2µg) immunoprecipitating PKC in Rat brain lysate.
    Lane 1 (input): Rat brain lysate 10µg
    Lane 2 (+): ab109539 & Rat brain lysate
    Lane 3 (-): Rabbit monoclonal IgG (ab172730) instead of ab109539 in Rat brain lysate
    For western blotting, VeriBlot for IP secondary antibody (HRP) (ab131366) was used as the secondary antibody at 1:1000 dilution.
    Blocking and diluting buffer: 5% NFDM/TBST.
  • Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) analysis of Rat cerebellum tissue sections labeling PKC with Purified ab109539 at 1:300 dilution (1.14 µg/ml). Heat mediated antigen retrieval was performed using Perform heat mediated antigen retrieval using citrate buffer, pH 6.0. Tissue was counterstained with Hematoxylin. ImmunoHistoProbe one step HRP Polymer (ready to use) secondary antibody was used at 1:0 dilution. PBS instead of the primary antibody was used as the negative control.
  • Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) analysis of Mouse cerebellum tissue sections labeling PKC with Purified ab109539 at 1:300 dilution (1.14 µg/ml). Heat mediated antigen retrieval was performed using Perform heat mediated antigen retrieval using citrate buffer, pH 6.0. Tissue was counterstained with Hematoxylin. ImmunoHistoProbe one step HRP Polymer (ready to use) secondary antibody was used at 1:0 dilution. PBS instead of the primary antibody was used as the negative control.
  • Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) analysis of Human cerebral cortex tissue sections labeling PKC with Purified ab109539 at 1:300 dilution (1.14 µg/ml). Heat mediated antigen retrieval was performed using Perform heat mediated antigen retrieval using citrate buffer, pH 6.0. Tissue was counterstained with Hematoxylin. ImmunoHistoProbe one step HRP Polymer (ready to use) secondary antibody was used at 1:0 dilution. PBS instead of the primary antibody was used as the negative control.
  • All lanes : Anti-PKC (phospho T514) antibody [EP2730Y] (ab109539) at 1/2000 dilution (purified)

    Lane 1 : Rat brain lysates
    Lane 2 : Rat brain lysates and then the membrane was incubated with phosphatase.

    Lysates/proteins at 15 µg per lane.

    Secondary
    All lanes : Goat Anti-Rabbit IgG H&L (HRP) (ab97051) at 1/20000 dilution

    Predicted band size: 78 kDa



    Blocking and diluting buffer : 5% NFDM/TBST
  • All lanes : Anti-PKC (phospho T514) antibody [EP2730Y] (ab109539) at 1/2000 dilution (purified)

    Lane 1 : Mouse cerebellum lysates
    Lane 2 : Mouse cerebellum lysates and then the membrane was incubated with phosphatase.

    Lysates/proteins at 15 µg per lane.

    Secondary
    All lanes : Goat Anti-Rabbit IgG H&L (HRP) (ab97051) at 1/20000 dilution

    Predicted band size: 78 kDa



    Blocking and diluting buffer : 5% NFDM/TBST
  • All lanes : Anti-PKC (phospho T514) antibody [EP2730Y] (ab109539) at 1/2000 dilution (unpurified)

    Lane 1 : Active human PKC alpha full length protein.
    Lane 2 : Active human PKC beta 1 full length protein.
    Lane 3 : Active human PKC beta 2 full length protein.
    Lane 4 : Active human PKC delta full length protein.
    Lane 5 : Active human PKC eta full length protein.
    Lane 6 : Active human PKC epsilon full length protein.
    Lane 7 : Active human PKC theta full length protein.
    Lane 8 : Active human PKC mu full length protein.

    Secondary
    All lanes : Goat Anti-Rabbit IgG H&L (HRP) (ab97051) at 1/20000 dilution

    Predicted band size: 78 kDa
    Observed band size: 100-150 kDa (why is the actual band size different from the predicted?)


    Exposure time: 3 minutes


    Blocking and Diluting buffer and concentration

    Active human PKC alpha full length protein (Catalog#ab55672) contains aa1-672 with GST-tag; Active human PKC beta 1 full length protein(Catalog#ab60840) contains aa1-671 with GST-tag; Active human PKC beta 2 full length protein(Catalog#ab60841) contains aa1-673 with GST-tag; Active human PKC delta full length protein(Catalog#ab60844) contains aa1-676 with GST-tag; Active human PKC eta full length protein(Catalog#ab60849) contains aa1-683 with GST-tag; Active human PKC epsilon full length protein(Catalog#ab60847) contains aa1-737 with GST-tag; Active human PKC theta full length protein(Catalog#ab56641) contains aa1-706 with GST-tag; Active human PKC mu full length protein(Catalog#ab60873) contains aa1-912 with GST-tag.

  • All lanes : Anti-PKC (phospho T514) antibody [EP2730Y] (ab109539) at 1/5000 dilution (purified)

    Lane 1 : HeLa (Human cervix adenocarcinoma epithelial cell) whole cell lysates
    Lane 2 : HeLa (Human cervix adenocarcinoma epithelial cell) treated with Phorbol-12-myristate-13-acetate whole cell lysates
    Lane 3 : HeLa (Human cervix adenocarcinoma epithelial cell) treated with Phorbol-12-myristate-13-acetate whole cell lysates. The membrane was then incubated with phosphatase.

    Lysates/proteins at 15 µg per lane.

    Secondary
    All lanes : Goat Anti-Rabbit IgG H&L (HRP) (ab97051) at 1/20000 dilution

    Predicted band size: 78 kDa
    Observed band size: 80 kDa (why is the actual band size different from the predicted?)


    Exposure time: 3 minutes


    Blocking and dilution buffer: 5% NFDM/TBST.

  • All lanes : Anti-PKC (phospho T514) antibody [EP2730Y] (ab109539) at 0.02 µg/ml (unpurified)

    Lane 1 : HeLa cell lysate with PKC gamma (pT514) peptide
    Lane 2 : HeLa cell lysate with PKC gamma non-phospho peptide
    Lane 3 : HeLa cell lysate

    Predicted band size: 78 kDa
    Observed band size: 80 kDa (why is the actual band size different from the predicted?)


    Exposure time: 15 seconds
  • Unpurified ab109539 at 1/100 dilution staining PKC (phospho T514) in paraffin-embedded Human brain tissue by Immunohistochemistry.

References

This product has been referenced in:
  • Huang L  et al. Congenital absence of corticospinal tract does not severely affect plastic changes of the developing postnatal spinal cord. Neuroscience 301:338-50 (2015). IHC-P, IHC-Fr ; Mouse . Read more (PubMed: 26079333) »
  • Huang L  et al. Ketamine potentiates hippocampal neurodegeneration and persistent learning and memory impairment through the PKC?-ERK signaling pathway in the developing brain. Brain Res 1476:164-71 (2012). WB . Read more (PubMed: 22985497) »

See all 3 Publications for this product

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