Recombinant
RabMAb

Recombinant Anti-LRP6 (phospho S1490) antibody [EP2360Y] (ab76417)

Overview

  • Product name

    Anti-LRP6 (phospho S1490) antibody [EP2360Y]
    See all LRP6 primary antibodies
  • Description

    Rabbit monoclonal [EP2360Y] to LRP6 (phospho S1490)
  • Host species

    Rabbit
  • Specificity

    This antibody detects LRP6 phosphorylated on serine 1490.
  • Tested applications

    Suitable for: WBmore details
    Unsuitable for: ICC/IF
  • Species reactivity

    Reacts with: Human
  • Immunogen

    A phospho specific peptide corresponding to residues surrounding serine 1490 of human LRP6.

  • Positive control

  • General notes

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

     

    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

  • Form

    Liquid
  • Storage instructions

    Shipped at 4°C. Upon delivery aliquot and store at -20°C. Avoid freeze / thaw cycles.
  • Storage buffer

    pH: 7.20
    Preservative: 0.01% Sodium azide
    Constituents: 59% PBS, 40% Glycerol, 0.05% BSA
  • Concentration information loading...
  • Purity

    Protein A purified
  • Clonality

    Monoclonal
  • Clone number

    EP2360Y
  • Isotype

    IgG
  • Research areas

Applications

Our Abpromise guarantee covers the use of ab76417 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 1/500 - 1/2000. Predicted molecular weight: 179 kDa.Can be blocked with LRP6 (phospho S1490) peptide (ab192756).
  • Application notes
    Is unsuitable for ICC/IF.
  • Target

    • Function

      Component of the Wnt-Fzd-LRP5-LRP6 complex that triggers beta-catenin signaling through inducing aggregation of receptor-ligand complexes into ribosome-sized signalsomes. Cell-surface coreceptor of Wnt/beta-catenin signaling, which plays a pivotal role in bone formation. The Wnt-induced Fzd/LRP6 coreceptor complex recruits DVL1 polymers to the plasma membrane which, in turn, recruits the AXIN1/GSK3B-complex to the cell surface promoting the formation of signalsomes and inhibiting AXIN1/GSK3-mediated phosphorylation and destruction of beta-catenin. Required for posterior patterning of the epiblast during gastrulation.
    • Tissue specificity

      Widely co-expressed with LRP5 during embryogenesis and in adult tissues.
    • Involvement in disease

      Defects in LRP6 are the cause of autosomal dominant coronary artery disease type 2 (ADCAD2) [MIM:610947].
    • Sequence similarities

      Belongs to the LDLR family.
      Contains 4 EGF-like domains.
      Contains 3 LDL-receptor class A domains.
      Contains 20 LDL-receptor class B repeats.
    • Domain

      The YWTD-EGF-like domains 1 and 2 are required for the interaction with Wnt-frizzled complex. The YWTD-EGF-like domains 3 and 4 are required for the interaction with DKK1.
      The PPPSP motifs play a central role in signal transduction by being phosphorylated, leading to activate the Wnt signaling pathway.
    • Post-translational
      modifications

      Dual phosphorylation of cytoplasmic PPPSP motifs sequentially by GSK3 and CK1 is required for AXIN1-binding, and subsequent stabilization and activation of beta-catenin via preventing GSK3-mediated phosphorylation of beta-catenin. Phosphorylated, in vitro, by GRK5/6 within and outside the PPPSP motifs. Phosphorylation at Ser-1490 by CDK14 during G2/M phase leads to regulation of the Wnt signaling pathway during the cell cycle. Phosphorylation by GSK3B is induced by RPSO1 binding and inhibited by DKK1. Phosphorylated, in vitro, by casein kinase I on Thr-1479.
      Undergoes gamma-secretase-dependent regulated intramembrane proteolysis (RIP). The extracellular domain is first released by shedding, and then, through the action of gamma-secretase, the intracellular domain (ICD) is released into the cytoplasm where it is free to bind to GSK3B and to activate canonical Wnt signaling.
      Palmitoylation on the two sites near the transmembrane domain leads to release of LRP6 from the endoplasmic reticulum.
      Mono-ubiquitinated which retains LRP6 in the endoplasmic reticulum.
      N-glycosylation is required for cell surface location.
    • Cellular localization

      Membrane. Endoplasmic reticulum. On Wnt signaling, undergoes a cycle of caveolin- or clathrin-mediated endocytosis and plasma membrane location. Released from the endoplasmic reticulum on palmitoylation. Mono-ubiquitination retains it in the endoplasmic reticulum in the absence of palmitoylation. On Wnt signaling, phosphorylated, aggregates and colocalizes with AXIN1 and GSK3B at the plasma membrane in LRP6-signalsomes. Chaperoned to the plasma membrane by MESD.
    • Information by UniProt
    • Database links

    • Alternative names

      • ADCAD2 antibody
      • C030016K15Rik antibody
      • Cd antibody
      • FLJ90062 antibody
      • FLJ90421 antibody
      • LDL receptor related protein 6 antibody
      • Low density lipoprotein receptor related protein 6 antibody
      • Low-density lipoprotein receptor-related protein 6 antibody
      • LRP-6 antibody
      • LRP6 antibody
      • LRP6_HUMAN antibody
      • OTTHUMP00000238979 antibody
      • OTTHUMP00000238980 antibody
      • OTTHUMP00000238982 antibody
      • STHAG7 antibody
      see all

    Images

    • All lanes : Anti-LRP6 (phospho S1490) antibody [EP2360Y] (ab76417) at 1/2000 dilution

      Lane 1 : Untreated HeLa (Human cervix adenocarcinoma epithelial cell) whole cell lysate
      Lane 2 : Untreated HeLa (Human cervix adenocarcinoma epithelial cell) whole cell lysate. The membrane was then treated with Alkaline 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: 179 kDa
      Observed band size: 180 kDa
      why is the actual band size different from the predicted?



      Blocking and diluting buffer: 5% NFDM/TBST

    • All lanes : Anti-LRP6 (phospho S1490) antibody [EP2360Y] (ab76417) at 1/1000 dilution

      Lane 1 : HeLa cell
      lysates, untreated
      Lane 2 : HeLa cell
      lysates treated with Calyculin

      Lysates/proteins at 10 µg per lane.

      Secondary
      All lanes : HRP labelled goat anti-rabbit at 1/2000 dilution

      Predicted band size: 179 kDa
      Observed band size: 210 kDa why is the actual band size different from the predicted?

    References

    This product has been referenced in:

    • Sun Y  et al. LncRNA H19/miR-194/PFTK1 axis modulates the cell proliferation and migration of pancreatic cancer. J Cell Biochem 120:3874-3886 (2019). Read more (PubMed: 30474270) »
    See 1 Publication for this product

    Customer reviews and Q&As

    1-3 of 3 Abreviews or Q&A

    Answer

    Thanks for your reply.

    I believe the data suggests that LRP6 is phosphorylated on S1490 in HEK293T cells but that there is no further induction of phosphorylation at this residue upon stimulation. That is why the band is the same intensity in both stimulated and untreated cells.

    If you have access to Hela cells, I would suggest doing the experiment with this cell line as, at least on the data we have,S1490 phosphorylation is induced upon stimulation in Hela cells.

    I hope this helps. Please contact me with any further comments or questions.

    Read More

    Question

    It will take approximately 5 minutes to complete the questionnaire. Please fill-in all fields so that we can better assist you.



    1) Abcam product code ab

    ab76417



    3) Description of the problem: no band / high background (many non-specific bands) / wrong band size / other?

    Many non-specific bands, no difference in positive control (WNT3a treated)



    4) Sample preparation:

    Type of sample (whole cell lysates, fraction, recombinant protein…): cell lysates

    Species : HEK293T

    Lysis buffer : RIPA

    Protease inhibitors: 100mM PMSF, Aprotinin, 0.5M NaF, 100mM Na3VO4

    Phosphatase inhibitors : see above

    Reducing agent : Invitrogen NuPAGE Reducing Agent (10x)

    Boiling for ≥5 min? yes

    Protein loaded ug/lane or cells/lane : 10ug/lane

    Positive control : WNT3a conditioned media

    Negative control : L-cell conditioned media





    5) Percentage of gel : NuPAGE 4-12% Bis-Tris

    Type of membrane: nitrocellulose

    Protein transfer verified , yes

    Blocking agent and concentration: 5% BSA in TBST

    Blocking time 1h

    Blocking temperature 4°C





    6) Primary antibody (If more than one was used, describe in “additional notes”) :

    Concentration or dilution 1/1000

    Diluent buffer 5%BSA/TBST

    Incubation time: o.n.

    Incubation temperature: 4°C





    7) Secondary antibody:

    Species: goat

    Reacts against: rabbit

    Concentration or dilution : 1/10 000

    Diluent buffer TBST

    Incubation time 1h

    Incubation temperature: at room temperature

    Fluorochrome or enzyme conjugate: HRP-conjugated





    8) Washing after primary and secondary antibodies:

    Buffer TBST

    Number of washes 4x 8min





    9)Detection method: ECL detection kit





    10) How many times have you run this staining? 2x

    Do you obtain the same results every time? yes

    What steps have you altered to try and optimize the use of this antibody? Different gel, incubation at 4°C and not for 2h at room temperature.

    Read More
    Answer

    HEK293T is not a suitable positive control for the LRP6 Phospho (pS1490) antibody (see attachment).
    We did not see a significant difference after treatment of 293T cells with Calyculin A. The best positive control is HeLa,
    as shown on the datasheet.
    Please let me know if you have any questions.

    Read More

    Answer

    Thank you for contacting us.

    I am sorry to hear you are experiencing difficulties with one of our products. We take product complaints very seriously, and investigate every product that we feel may not be performing correctly.

    Thank you for providing the details you have. In order to collect some further details regarding the samples tested and the protocol used I have attached a questionnaire to this email. This should not take longer than 5-10 minutes to complete. Once we receive the completed questionnaire, we will look at the protocol and see if there are any suggestions we can make that may improve the results. This information will also allow us to investigate this case internally and initiate any additional testing where necessary.

    If the product was purchased in the last six months and is being used according to our Abpromise, we would be happy to replace or refund the antibody.

    I look forward to receiving your reply.

    Read More

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