Overview

  • Product name
    Anti-eIF4E (phospho S209) antibody
    See all eIF4E primary antibodies
  • Description
    Rabbit polyclonal to eIF4E (phospho S209)
  • Host species
    Rabbit
  • Specificity
    This phosphorylation site specific antibody is selective for eIF-4E containing a phosphate on serine 209.
  • Tested applications
    Suitable for: WBmore details
  • Species reactivity
    Reacts with: Human
    Predicted to work with: Mouse, Rat, Rabbit, a wide range of other species
  • Immunogen

    Synthetic peptide (Human) derived from the region of eIF-4E that contains serine 209, based on the Homo sapiens sequence. This region is identical among many species including rat, mouse and rabbit.

  • Positive control
    • HeLa cell lysate

Properties

Applications

Our Abpromise guarantee covers the use of ab4774 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. Detects a band of approximately 25 kDa (predicted molecular weight: 25 kDa).

Target

  • Function
    Its 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 similarities
    Belongs 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

Images

  • HeLa cell lysates (A) and HeLa cell lysates treated with alkaline phosphatase (B) were resolved by SDS-PAGE on a 4-20% Tris-glycine gel. The proteins were then transferred to PVDF membrane. Membranes were incubated with 1 µg/mL ab4774. After washing, membranes were incubated with goat  (ab’)2 anti-rabbit IgG alkaline phosphatase and bands were detected using the Tropix WesternStar detection method. HeLa cell lysates (A) and HeLa cell lysates treated with alkaline phosphatase (B) were resolved by SDS-PAGE on a 4-20% Tris-glycine gel. The proteins were then transferred to PVDF membrane. Membranes were incubated with 1 µg/mL ab4774. After washing, membranes were incubated with goat (ab’)2 anti-rabbit IgG alkaline phosphatase and bands were detected using the Tropix WesternStar detection method.
  • Extracts of HeLa cells were resolved by SDS-PAGE on a 4-20% Tris-glycine gel and transferred to PVDF. The membrane was blocked with a 5% BSA-TBST buffer for one hour at room temperature, either left untreated (1-4) or treated with lambda (?) phosphatase (5), and then incubated with the eIF4E (phospho S209) antibody for two hours at room temperature in a 3% BSA-TBST buffer, following prior incubation with: no peptide (1,5), the nonphosphopeptide corresponding to the phosphopeptide immunogen (2), a generic phosphoserine-containing peptide (3), or the phosphopeptide immunogen (4). After washing, the membrane was incubated with goat F(ab’)2 anti-rabbit IgG alkaline phosphatase and signals were detected using the Pierce SuperSignal™ method.

    The data show that only the peptide corresponding to eIF4E (phospho S209) blocks the antibody signal, demonstrating the specificity of the antibody. The data also show that phosphatase stripping eliminates the signal, verifying that the antibody i

References

This product has been referenced in:
  • Kapasi P  et al. L13a blocks 48S assembly: role of a general initiation factor in mRNA-specific translational control. Mol Cell 25:113-26 (2007). Read more (PubMed: 17218275) »
See 1 Publication for this product

Customer reviews and Q&As

1-2 of 2 Abreviews or Q&A

Question
Answer

Thank you for your enquiry. The concentration is 0.5 mg/mL. I hope this information helps, please do not hesitate to contact us if you need any more advice or information.

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Question

Here are two blots, one from abCAM and one from cell signaling! And all the details you requested! Lot Number 63956, Abcam order number: 49277, P.O. number: 3P668468. 1. Please describe the problem (high background, wrong band size, more bands, no band etc). Wrong size approximately 36kD and 65kD! 2. On what material are you testing the antibody in WB? Species? Brain tissue lyaste from Guinea Pig along with Pos and Neg cell line lyastae! Cell extract/ Nuclear extract? Whole cell lysate. Purified protein? No Recombinant protein? No 3. How much protein did you load? From 10 to 50 ug, we always run a standard curve with increasing amount of protein to evaluate an antibody of interest before doing the various experimental conditions. How did you prepare the lysate for the analysis (protease inhibito 50 mM Tris-HCl, Standard lysis buffer with protease inhibitors and phosphatase inhibitors. Did you heat the samples? Yes 4. Primary Antibody Specification (in which species was it raised against)? In Rabbit. ab4744, you have all the details! At what dilution(s) have you tested this antibody? 1ug/ml Incubation time, wash step? 2 hours in 1% non-fat dry milk solution in 20 mM TBST, ph 7.6 as suggested in the protocol supplied with the antibody. Three wash steps at room temp for 5 min each. 5. Secondary Antibody Specification (in which species was it raised against)? Goat. Anti-rabbit IgG whole molecule, HRP conjugate At what dilution(s) have you tested this antibody? 1:2500 Incubation time, wash step? 1 hr at room temp, wash step as for primary antibody. Do you know whether the problems you are experiencing come from th No 6. What detection method are you using? Chemiluminescent detection method. 7. Background bands Have you eliminated the possibility that any background bands coul Is the blocking step sufficient? (We recommend blocking the membra YES Are your washing steps sufficiently stringent? (Multiple short was YES At what size are the bands migrating? Could they be degradation pr Approx 36kD and 65kD Please provide an image of your blot (as an e-mail attachment, a f Attached. 8. Optimization attempts How many times have you tried the Western? Two times! And we do western almost everyday in our lab, check the band with your competitor on the image labeled Cell Signaling Technol on the right hand side.. Do you obtain the same results every time e.g. are background band YES What steps have you altered? YES, overnight incubation with primary antibody! 9. Did you apply positive and negative controls along with the samples? Please specify. YES, see band at 36 kD and not at approx. 25kD with abCAM. On the contrary, see band at lower than 29kD with cell signaling tehnol. product. Using abCAM antibody, we did not get band on western blot at the right molecular weight. Please do the needful.

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Answer

Thank you very much for the details that you have provided, and I'm sorry to hear that you are experiencing difficulty with this antibody. As stated on the online datasheet, this particular antibody has been characterized using HeLa cell lysates and has not yet been tested on other species although it is expected to cross-react with a wide range of other species, due to sequence homology. We have not had any other complaints regarding this antibody; the problem may very well be due to the batch you received. I can offer you a replacement batch to try or a refund, please let me know which one you would prefer.

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