Anti-eIF4E (phospho S209) antibody [EP2151Y] (ab76256)

Overview

  • Product nameAnti-eIF4E (phospho S209) antibody [EP2151Y]
    See all eIF4E primary antibodies
  • Description
    Rabbit monoclonal [EP2151Y] to eIF4E (phospho S209)
  • Tested applicationsSuitable for: ICC/IF, WB, IP, IHC-Pmore details
  • Species reactivity
    Reacts with: Mouse, Rat, Human, Pig
  • Immunogen

    Synthetic peptide (the amino acid sequence is considered to be commercially sensitive) corresponding to Human eIF4E.
    Database link: P06730

  • Positive control
    • WB: 293 cell lysate treated with alkaline phosphatase and HEK293 cell lysate treated with Dexamethasone. IHC-P: human breast carcinoma tissue. ICC/IF: HEK293 cells.
  • General notes

    This product is a recombinant rabbit monoclonal antibody. Produced using Abcam’s RabMAb® technology. RabMAb® technology is covered by the following U.S. Patents, No. 5,675,063 and/or 7,429,487.

    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. 

    Alternative versions available:
    Anti-eIF4E (phospho S209) antibody (Biotin) [EP2151Y] (ab201471)
    Anti-eIF4E (phospho S209) antibody (HRP) [EP2151Y] (ab200858)

Properties

Applications

Our Abpromise guarantee covers the use of ab76256 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
ICC/IF 1/500.
WB 1/1000 - 1/100000. Detects a band of approximately 25 kDa (predicted molecular weight: 25 kDa).
IP 1/40 - 1/60.
IHC-P 1/50 - 1/250. Perform heat mediated antigen retrieval before commencing with IHC staining protocol.

See protocols (link: http://www.abcam.com/protocols/ihc-antigen-retrieval-protocol).

Target

  • FunctionIts translation stimulation activity is repressed by binding to the complex CYFIP1-FMR1 (By similarity). Recognizes and binds the 7-methylguanosine-containing mRNA cap during an early step in the initiation of protein synthesis and facilitates ribosome binding by inducing the unwinding of the mRNAs secondary structures. Component of the CYFIP1-EIF4E-FMR1 complex which binds to the mRNA cap and mediates translational repression. In the CYFIP1-EIF4E-FMR1 complex this subunit mediates the binding to the mRNA cap.
  • Sequence similaritiesBelongs to the eukaryotic initiation factor 4E family.
  • Post-translational
    modifications
    Phosphorylation increases the ability of the protein to bind to mRNA caps and to form the eIF4F complex.
  • Information by UniProt
  • Database links
  • Alternative names
    • AUTS19 antibody
    • CBP antibody
    • eIF 4E antibody
    • eIF 4F 25 kDa subunit antibody
    • EIF 4F antibody
    • eIF-4E antibody
    • eIF-4F 25 kDa subunit antibody
    • eIF4E antibody
    • EIF4E1 antibody
    • EIF4EL1 antibody
    • EIF4F antibody
    • Eukaryotic translation initiation factor 4 E antibody
    • Eukaryotic translation initiation factor 4E antibody
    • Eukaryotic translation initiation factor 4E like 1 antibody
    • IF4E_HUMAN antibody
    • Messanger RNA Cap Binding Protein eIF 4E antibody
    • MGC111573 antibody
    • mRNA cap binding protein antibody
    • mRNA cap-binding protein antibody
    see all

Anti-eIF4E (phospho S209) antibody [EP2151Y] images

  • All lanes : Anti-eIF4E (phospho S209) antibody [EP2151Y] (ab76256) at 1/100000 dilution (purified)

    Lane 1 : Untreated HEK293 whole cell lysate
    Lane 2 : HEK293 cells treated with 10uM dexamethasone for 1 hour whole cell lysate
    Lane 3 : HEK293 cells treated with 10uM dexamethasone for 1 hour whole cell lysate.The membrane was then incubated with alkaline phosphatase.

    Lysates/proteins at 10 µg per lane.

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

    Predicted band size : 25 kDa
    Observed band size : 25 kDa


    Exposure time : 30 seconds

    Blocking buffer and concentration 2% BSA/TBST.
    Diluting buffer and concentration 2% BSA/TBST.

  • Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) analysis of human cervix carcinoma tissue labelling eIF4E with purified ab76256 at 1/50. Heat mediated antigen retrieval was performed using Tris/EDTA buffer pH 9. ab97051, a goat anti-rabbit IgG H&L (HRP) was used as the secondary antibody (1/500). Negative control using PBS instead of primary antibody. Counterstained with hematoxylin.
  • Immunocytochemistry/Immunofluorescence analysis of untreated, 20% serum treated and 20% serum + LP treated NIH/3T3 cells labelling eIF4E (phospho S209) with ab76256 (left) and eIF4E with ab33766 (right) both at a dilution of 1/500.

    Cells were fixed with 100% methanol. ab150077, an Alexa Fluor® 488-conjugated goat anti-rabbit IgG (1/1000) was used as the secondary antibody. DAPI (blue) was used as the nuclear counterstain. ab7291, a mouse anti-tubulin (1/1000) and ab150120, an Alexa Fluor® 594-conjugated goat anti-mouse IgG (1/1000) were also used.

    The image shows increased cytoplasmic staining after 20% serum treatment on NIH3T3 cells when compared with no serum treated cells. The LP treatment decreased the increased cytoplasmic staining caused by 20% serum.

    ab33766 was used as a Pan control for ab76256. The results showed cytoplasmic staining on no serum, 20% serum and 20% serum +LP treated NIH3T3 cells.

  • ab76256 (purified) at 1/40 immunoprecipitating eIF4E (phospho S209) in HEK293 whole cell lysate. 10 ug of cell lysate was present in the input. For western blotting, a HRP-conjugated Veriblot for IP secondary antibody (ab131366) (1/1,500) was used as the secondary antibody. A rabbit monoclonal IgG (ab172730) was used intead of ab128913 as a negative control (Lane 3).

    Blocking buffer and concentration: 5% NFDM/TBST.

    Diluting buffer and concentration: 5% NFDM /TBST.

  • All lanes : Anti-eIF4E (phospho S209) antibody [EP2151Y] (ab76256) at 1/1000 dilution (purified)

    Lane 1 : Mouse spleen lysate
    Lane 2 : Rat brain lysate
    Lane 3 : Pig heart lysate

    Lysates/proteins at 20 µg per lane.

    Secondary
    Goat Anti-Rabbit IgG H&L (HRP) (ab97051) at 1/1000 dilution (Goat Anti-Rabbit IgG, (H+L), Peroxidase conjugated)

    Predicted band size : 25 kDa

    Blocking buffer and concentration: 5% NFDM/TBST.
    Diluting buffer and concentration: 5% NFDM /TBST.

  • Dot blot analysis of eIF4E (pS209) peptide (Lane 1) and eIF4E non-phospho peptide (Lane 2) labelling eIF4E (pS209) with purified ab76256 at a dilution of 1/1000. ab97051 (Peroxidase conjugated goat anti-rabbit IgG (H+L)) was used as the secondary antibody at a dilution of 1/100000.

    Blocking and dilution buffer: 5% NFDM/TBST.

    Exposure time: 3 minutes.

  • Immunocytochemistry/Immunofluorescence analysis of serum starved HEK293 cells treated with CGP 57380 ab120365) labelling eIF4E (phospho S209) with unpurified ab32124 at 1/100. Decrease in eIF4E (phospho S209) expression correlates with increased concentration of CGP 57380, as described in literature.
    The cells were incubated at 37°C for 1h in media containing different concentrations of ab120365 (CGP 57380) in DMSO, fixed with 100% methanol for 5 minutes at -20°C and blocked with PBS containing 10% goat serum, 0.3 M glycine, 1% BSA and 0.1% tween for 2h at room temperature. Staining of the treated cells with unpurified ab76256 was performed overnight at 4°C in PBS containing 1% BSA and 0.1% tween. A DyLight 488 goat anti-rabbit polyclonal antibody (ab96899) at 1/250 dilution was used as the secondary antibody. Nuclei were counterstained with DAPI and are shown in blue.

  • All lanes : Anti-eIF4E (phospho S209) antibody [EP2151Y] (ab76256) at 1/50000 dilution (purified)

    Lane 1 : Untreated HEK293 cell lysate
    Lane 2 : HEK293 treated with 10mM Dexamethasone 1 hour lysate

    Lysates/proteins at 10 µg per lane.

    Secondary
    Goat Anti-Rabbit IgG H&L (HRP) (ab97051) at 1/10000 dilution (Goat Anti-Rabbit IgG, (H+L), Peroxidase conjugated)

    Predicted band size : 25 kDa

    Blocking buffer and concentration: 5% NFDM/TBST.
    Diluting buffer and concentration: 5% NFDM /TBST.

  • All lanes : Anti-eIF4E (phospho S209) antibody [EP2151Y] (ab76256) at 1/50000 dilution (purified)

    Lane 1 : Untreated 293 cell lysate
    Lane 2 : 293 cell lysate treated with alkaline phosphatase

    Lysates/proteins at 10 µg per lane.

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

    Predicted band size : 25 kDa


    Exposure time : 1 minute

    Blocking and dilution buffer: 5% NFDM/TBST.

  • All lanes : Anti-eIF4E (phospho S209) antibody [EP2151Y] (ab76256) at 1/100000 dilution (purified)

    Lane 1 : Untreated HEK293 cell lysate
    Lane 2 : HEK293 cell lysate - treated with Dexamethasone

    Lysates/proteins at 10 µg per lane.

    Secondary
    Goat Anti-Rabbit IgG H&L (HRP) (ab97051) at 1/10000 dilution

    Predicted band size : 25 kDa

    Exposure time:

    eIF4E pS209: 15 seconds.

    eIF4E: 3 minutes.

     

    Blocking and dilution buffer: 5% NFDM/TBST.

References for Anti-eIF4E (phospho S209) antibody [EP2151Y] (ab76256)

This product has been referenced in:
  • Grzmil M  et al. MNK1 pathway activity maintains protein synthesis in rapalog-treated gliomas. J Clin Invest 124:742-54 (2014). IHC-P ; Human . Read more (PubMed: 24401275) »
  • Zheng J  et al. Phosphorylated Mnk1 and eIF4E are associated with lymph node metastasis and poor prognosis of nasopharyngeal carcinoma. PLoS One 9:e89220 (2014). IHC-P ; Human . Read more (PubMed: 24551240) »

See all 10 Publications for this product

Product Wall

Abcam guarantees this product to work in the species/application used in this Abreview.
Application Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections)
Sample Human Tissue sections (Kidney)
Antigen retrieval step Heat mediated - Buffer/Enzyme Used: Ventana Ultra CC1
Permeabilization No
Specification Kidney
Blocking step Optiview H2O2 as blocking agent for 8 minute(s) · Concentration: 0.04% · Temperature: 36°C
Fixative Formaldehyde
Username

Mr. Alex Bowman

Verified customer

Submitted Mar 08 2016

Ab76256 reconnait uniquement la protéine phosphorilée au résidu Serine 209. Comme vous pouvez le voir sur notre western blot, quand la protéine est déphosphorylée avec de l’alkaline phosphatase (AP) l&rsq...

Read More

Please note: All products are "FOR RESEARCH USE ONLY AND ARE NOT INTENDED FOR DIAGNOSTIC OR THERAPEUTIC USE"