Recombinant
RabMAb

Recombinant Anti-PTEN (phospho T366) antibody [EP229] (ab109454)

Overview

  • Product name

    Anti-PTEN (phospho T366) antibody [EP229]
    See all PTEN primary antibodies
  • Description

    Rabbit monoclonal [EP229] to PTEN (phospho T366)
  • Host species

    Rabbit
  • Specificity

    ab109454 only detects PTEN phosphorylated on T366.
  • Tested applications

    Suitable for: WB, IP, IHC-P, Dot blotmore details
    Unsuitable for: Flow Cyt
  • Species reactivity

    Reacts with: Mouse, Rat, Human
  • Immunogen

    Synthetic peptide (the amino acid sequence is considered to be commercially sensitive) corresponding to Human PTEN (phospho T366).

  • Positive control

    • WB: Mouse brain tissue lysate and C6 cell lysate treated with Calyculin A. IHC-P: Human breast ductal carcinoma tissue.
  • General notes

     

     

    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 ab109454 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. Detects a band of approximately 54 kDa (predicted molecular weight: 47 kDa).
IP 1/50.
IHC-P 1/100 - 1/250. Perform heat mediated antigen retrieval before commencing with IHC staining protocol.
Dot blot Use at an assay dependent concentration.
  • Application notes
    Is unsuitable for Flow Cyt.
  • Target

    • Function

      Tumor suppressor. Acts as a dual-specificity protein phosphatase, dephosphorylating tyrosine-, serine- and threonine-phosphorylated proteins. Also acts as a lipid phosphatase, removing the phosphate in the D3 position of the inositol ring from phosphatidylinositol 3,4,5-trisphosphate, phosphatidylinositol 3,4-diphosphate, phosphatidylinositol 3-phosphate and inositol 1,3,4,5-tetrakisphosphate with order of substrate preference in vitro PtdIns(3,4,5)P3 > PtdIns(3,4)P2 > PtdIns3P > Ins(1,3,4,5)P4. The lipid phosphatase activity is critical for its tumor suppressor function. Antagonizes the PI3K-AKT/PKB signaling pathway by dephosphorylating phosphoinositides and thereby modulating cell cycle progression and cell survival. The unphosphorylated form cooperates with AIP1 to suppress AKT1 activation. Dephosphorylates tyrosine-phosphorylated focal adhesion kinase and inhibits cell migration and integrin-mediated cell spreading and focal adhesion formation. Plays a role as a key modulator of the AKT-mTOR signaling pathway controlling the tempo of the process of newborn neurons integration during adult neurogenesis, including correct neuron positioning, dendritic development and synapse formation. May be a negative regulator of insulin signaling and glucose metabolism in adipose tissue. The nuclear monoubiquitinated form possesses greater apoptotic potential, whereas the cytoplasmic nonubiquitinated form induces less tumor suppressive ability. In motile cells, suppresses the formation of lateral pseudopods and thereby promotes cell polarization and directed movement.
      Isoform alpha: Functional kinase, like isoform 1 it antagonizes the PI3K-AKT/PKB signaling pathway. Plays a role in mitochondrial energetic metabolism by promoting COX activity and ATP production, via collaboration with isoform 1 in increasing protein levels of PINK1.
    • Tissue specificity

      Expressed at a relatively high level in all adult tissues, including heart, brain, placenta, lung, liver, muscle, kidney and pancreas.
    • Involvement in disease

      Cowden syndrome 1
      Lhermitte-Duclos disease
      Bannayan-Riley-Ruvalcaba syndrome
      Squamous cell carcinoma of the head and neck
      Endometrial cancer
      PTEN mutations are found in a subset of patients with Proteus syndrome, a genetically heterogeneous condition. The molecular diagnosis of PTEN mutation positive cases classifies Proteus syndrome patients as part of the PTEN hamartoma syndrome spectrum. As such, patients surviving the early years of Proteus syndrome are likely at a greater risk of developing malignancies.
      Glioma 2
      VACTERL association with hydrocephalus
      Prostate cancer
      Macrocephaly/autism syndrome
      A microdeletion of chromosome 10q23 involving BMPR1A and PTEN is a cause of chromosome 10q23 deletion syndrome, which shows overlapping features of the following three disorders: Bannayan-Zonana syndrome, Cowden disease and juvenile polyposis syndrome.
    • Sequence similarities

      Contains 1 C2 tensin-type domain.
      Contains 1 phosphatase tensin-type domain.
    • Domain

      The C2 domain binds phospholipid membranes in vitro in a Ca(2+)-independent manner; this binding is important for its tumor suppressor function.
    • Post-translational
      modifications

      Constitutively phosphorylated by CK2 under normal conditions. Phosphorylated in vitro by MAST1, MAST2, MAST3 and STK11. Phosphorylation results in an inhibited activity towards PIP3. Phosphorylation can both inhibit or promote PDZ-binding. Phosphorylation at Tyr-336 by FRK/PTK5 protects this protein from ubiquitin-mediated degradation probably by inhibiting its binding to NEDD4. Phosphorylation by ROCK1 is essential for its stability and activity. Phosphorylation by PLK3 promotes its stability and prevents its degradation by the proteasome.
      Monoubiquitinated; monoubiquitination is increased in presence of retinoic acid. Deubiquitinated by USP7; leading to its nuclear exclusion. Monoubiquitination of one of either Lys-13 and Lys-289 amino acid is sufficient to modulate PTEN compartmentalization. Ubiquitinated by XIAP/BIRC4.
    • Cellular localization

      Secreted. May be secreted via a classical signal peptide and reenter into cells with the help of a poly-Arg motif and Cytoplasm. Nucleus. Nucleus, PML body. Monoubiquitinated form is nuclear. Nonubiquitinated form is cytoplasmic. Colocalized with PML and USP7 in PML nuclear bodies. XIAP/BIRC4 promotes its nuclear localization.
    • Information by UniProt
    • Database links

    • Alternative names

      • 10q23del antibody
      • BZS antibody
      • DEC antibody
      • GLM2 antibody
      • MGC11227 antibody
      • MHAM antibody
      • MMAC1 antibody
      • MMAC1 phosphatase and tensin homolog deleted on chromosome 10 antibody
      • Mutated in multiple advanced cancers 1 antibody
      • Phosphatase and tensin homolog antibody
      • Phosphatase and tensin like protein antibody
      • Phosphatidylinositol 3,4,5-trisphosphate 3-phosphatase and dual-specificity protein phosphatase PTEN antibody
      • Pten antibody
      • PTEN_HUMAN antibody
      • PTEN1 antibody
      • TEP1 antibody
      see all

    Images

    • All lanes : Anti-PTEN (phospho T366) antibody [EP229] (ab109454) at 1/20000 dilution

      Lane 1 : Untreated Hela whole cell lysate
      Lane 2 : HeLa treated with Calyculin A whole cell lysate
      Lane 3 : HeLa whole cell lysate treated with Calyculin A and phosphatase on the membrane

      Lysates/proteins at 10 µg per lane.

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

      Predicted band size: 47 kDa


      Exposure time: 1 minute


      Blocking and dilution buffer: 5% NFDM/TBST.

    • Dot blot analysis of PTEN (pT366+ pS370) peptide (Lane 1), PTEN (pS370) peptide (Lane 2), PTEN (pT366) peptide (Lane 3), and PTEN non-phospho peptide (Lane 4) labelling PTEN phospho T366 with ab109454 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: 10 seconds.

    • ab109454 at 1/100 dilution staining PTEN in Human breast ductal carcinoma by Immunohistochemistry, Paraffin-embedded tissue.

      Perform heat mediated antigen retrieval before commencing with IHC staining protocol.

    References

    This product has been referenced in:

    • Stump B  et al. Glycogen synthase kinase 3-ß inhibition induces lymphangiogenesis through ß-catenin-dependent and mTOR-independent pathways. PLoS One 14:e0213831 (2019). Read more (PubMed: 30964887) »
    • Gong L  et al. miR-222 promotes invasion and migration of ovarian carcinoma by targeting PTEN. Oncol Lett 16:984-990 (2018). Read more (PubMed: 29963173) »
    See all 8 Publications for this product

    Customer reviews and Q&As

    1-3 of 3 Abreviews or Q&A

    Answer

    Thank you so much for sending on this data. Could you also tell me which lysates (treated vs untreated) were loaded into each lane? Are the same samples in corresponding lanes of the ab32199 image?

    Thanks again, and I look forward to hearing from you. Please let me know if you have any further questions at this time.

    Read More

    Answer

    Thank you for your patience while I have been discussing this case with the scientists in the lab. I sincerely apologize for the delay in getting back to you.

    Using both ab32199 and ab109454, we have seen similar bands to yours. For example, the band around 37 kDa with ab32199 is also seen in certain samples but not in others. The specific PTEN bandis normally seenaround 54-55 kDa, however, not around 50-58 kDa whichyou have seen with ab109454. I've looked through the literature, and molecular weight changes due to cleavage and/or phosphorylation could explain the difference in band size. However, I am not sure how to explain why these two bands would not be picked up by ab32199, which is specific for a different region of the protein and should pick up both phosphorylated and non-phosphorylated PTEN. The results also don't correlate with the trastuzumab treatment, which I also don't have a good explanation for.

    Have you run any more blots since we were last in touch? At this point, I would be happy to send a replacement vial of ab102949 to compare with these results. We also have several other antibodies that are specific for different PTEN phosphorylation sites, such as ab30654 and ab76431. Would you like to try one of these? Alternatively I can issue you a credit or refund.

    I apologize again for the delay and that we don't have a better explanation for these results, but please let me know if you have any further questions or if there is anything else that we can do for you, and I'll be happy to help.

    Read More

    Question
    Answer

    Thank you for your call today and for letting us know about the trouble with ab109454.

    As we discussed, you can send images in a reply to this email, and I will get in touch with the lab to see if we have more information about the extra bands onthe blot.

    For our records, could you also tell me how much protein was loaded per lane of the blot?

    We do fully guarantee this antibody to work with human lysates in Western blot, so if you are not satisfied with the results at any time, I would be happy to send a replacement or issue a credit or refund.

    Please let me know if you have any further questions or if there is anything else that we can do for you. Have a great day!

    Read More

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