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重组RabMAb

重组Anti-PTEN (phospho T366)抗体[EP229] (ab109454)

Submit a review Q&A (3)References (13)
Western blot - Anti-PTEN (phospho T366) antibody [EP229] (ab109454)
  • Western blot - Anti-PTEN (phospho T366) antibody [EP229] (ab109454)
  • Western blot - Anti-PTEN (phospho T366) antibody [EP229] (ab109454)
  • Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-PTEN (phospho T366) antibody [EP229] (ab109454)
  • Dot Blot - Anti-PTEN (phospho T366) antibody [EP229] (ab109454)
  • Anti-PTEN (phospho T366) antibody [EP229] (ab109454)

Key features and details

  • Produced recombinantly (animal-free) for high batch-to-batch consistency and long term security of supply
  • Rabbit monoclonal [EP229] to PTEN (phospho T366)
  • Suitable for: WB, IHC-P, Dot blot
  • Reacts with: Mouse, Rat, Human

Conjugates logo Related conjugates and formulations

Carrier Free

概述

  • 产品名称

    Anti-PTEN (phospho T366)抗体[EP229]
    参阅全部 PTEN 一抗

  • 描述

    兔单克隆抗体[EP229] to PTEN (phospho T366)

  • 宿主

    Rabbit

  • 特异性

    The mouse and rat recommendation is based on the WB results. We do not guarantee IHC-P for mouse and rat.

  • 经测试应用

    适用于: WB, IHC-P, Dot blotmore details
    不适用于: Flow Cyt

  • 种属反应性

    与反应: Mouse, Rat, Human

  • 免疫原

    Synthetic peptide. This information is proprietary to Abcam and/or its suppliers.

  • 阳性对照

    • WB: HeLa (Human cervix adenocarcinoma epithelial cell), NIH/3T3 (Mouse embryonic fibroblast), and C6 (Rat glial tumor glial cell) whole cell lysate untreated, treated with 100ng/ml Calyculin A for 30 minutes, and ted with 100ng/ml Calyculin A for 30 minutes whole cell lysate then the membrane was incubated with Alkaline phosphatase. IHC-P: Human breast ductal carcinoma tissue and prostate cancer sections.

  • 常规说明

    This product has switched from a hybridoma to recombinant production method on 9th June 2023.

    PTEN is a protein implicated in several disease, including certain cancers and neurological diseases. PTEN is expressed ubiquitously throughout the body and acts as a phosphatase to dephosphorylate phosphatidylinositol (3,4,5)-trisphosphate. This is important in the inhibition of the Akt signalling pathway, which plays an important role in regulating cellular behaviours such as cell growth, survival, and migration.

     

    This product is a recombinant monoclonal antibody, which offers several advantages including:

    • - High batch-to-batch consistency and reproducibility
    • - Improved sensitivity and specificity
    • - Long-term security of supply
    • - Animal-free production
    For more information see here.

    Our RabMAb® technology is a patented hybridoma-based technology for making rabbit monoclonal antibodies. For details on our patents, please refer to RabMAb® patents.

性能

应用

The Abpromise guarantee

Abpromise™承诺保证使用ab109454于以下的经测试应用

“应用说明”部分 下显示的仅为推荐的起始稀释度；实际最佳的稀释度/浓度应由使用者检定。

应用 Ab评论 说明
WB
1/10000. Detects a band of approximately 54 kDa (predicted molecular weight: 47 kDa).
IHC-P
1/2000. Perform heat mediated antigen retrieval before commencing with IHC staining protocol.

The mouse and rat recommendation is based on the WB results. We do not guarantee IHC-P for mouse and rat.
Dot blot
1/10000.
说明
WB
1/10000. Detects a band of approximately 54 kDa (predicted molecular weight: 47 kDa).
IHC-P
1/2000. Perform heat mediated antigen retrieval before commencing with IHC staining protocol.

The mouse and rat recommendation is based on the WB results. We do not guarantee IHC-P for mouse and rat.
Dot blot
1/10000.
应用说明
Is unsuitable for Flow Cyt.

靶标

  • 功能

    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.

  • 组织特异性

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

  • 疾病相关

    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.

  • 序列相似性

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

  • 结构域

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

  • 翻译后修饰

    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.

  • 细胞定位

    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

  • 数据库链接

    see all

  • 别名

    • 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

图片

  • Western blot - Anti-PTEN (phospho T366) antibody [EP229] (ab109454)
    Western blot - Anti-PTEN (phospho T366) antibody [EP229] (ab109454)
    All lanes : Anti-PTEN (phospho T366) antibody [EP229] - BSA and Azide free (ab208104) at 1/100000 dilution

    Lane 1 : HeLa (Human cervix adenocarcinoma epithelial cell) whole cell lysate
    Lane 2 : HeLa (Human cervix adenocarcinoma epithelial cell) treated with 100ng/ml Calyculin A for 30 minutes whole cell lysate
    Lane 3 : HeLa (Human cervix adenocarcinoma epithelial cell) treated with 100ng/ml Calyculin A for 30 minutes whole cell lysate, then the membrane was incubated 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

    Developed using the ECL technique.

    Performed under reducing conditions.

    Predicted band size: 47 kDa


    Exposure time: 80 seconds


    This data was developed using the same antibody clone in a different buffer formulation - BSA and Azide free (ab208104).

    Recommended concentration for ab109454: 1/10000 dilution (0.1ug/ml).

    Blocking and diluting buffer and concentration: 5% NFDM/TBST,

    Primary and secondary antibodies were incubated for 1h at room temperature.

  • Western blot - Anti-PTEN (phospho T366) antibody [EP229] (ab109454)
    Western blot - Anti-PTEN (phospho T366) antibody [EP229] (ab109454)
    All lanes : Anti-PTEN (phospho T366) antibody [EP229] - BSA and Azide free (ab208104) at 1/100000 dilution

    Lane 1 : NIH/3T3 (Mouse embryonic fibroblast) whole cell lysate
    Lane 2 : NIH/3T3 (Mouse embryonic fibroblast) treated with 100 ng/ml Calyculin A for 30 minutes whole cell lysate
    Lane 3 : NIH/3T3 (Mouse embryonic fibroblast) treated with 100 ng/ml Calyculin A for 30 minutes whole cell lysate, then the membrane was incubated 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

    Developed using the ECL technique.

    Performed under reducing conditions.

    Predicted band size: 47 kDa


    Exposure time: 40 seconds


    This data was developed using the same antibody clone in a different buffer formulation - BSA and Azide free (ab208104).

    Recommended concentration for ab109454: 1/10000 dilution (0.1ug/ml).

    Blocking and diluting buffer and concentration: 5% NFDM/TBST,

    Primary and secondary antibodies were incubated for 1h at room temperature.

  • Western blot - Anti-PTEN (phospho T366) antibody [EP229] (ab109454)
    Western blot - Anti-PTEN (phospho T366) antibody [EP229] (ab109454)
    All lanes : Anti-PTEN (phospho T366) antibody [EP229] - BSA and Azide free (ab208104) at 1/100000 dilution

    Lane 1 : C6 (Rat glial tumor glial cell) whole cell lysate
    Lane 2 : C6 (Rat glial tumor glial cell) treated with 100 ng/ml Calyculin A for 30 minutes whole cell lysate
    Lane 3 : C6 (Rat glial tumor glial cell) treated with 100 ng/ml Calyculin A for 30 minutes whole cell lysate, then the membrane was incubated 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

    Developed using the ECL technique.

    Performed under reducing conditions.

    Predicted band size: 47 kDa


    Exposure time: 20 seconds


    This data was developed using the same antibody clone in a different buffer formulation - BSA and Azide free (ab208104).

    Recommended concentration for ab109454: 1/10000 dilution (0.1ug/ml).

    Blocking and diluting buffer and concentration: 5% NFDM/TBST,

    Primary and secondary antibodies were incubated for 1h at room temperature.

  • Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-PTEN (phospho T366) antibody [EP229] (ab109454)
    Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-PTEN (phospho T366) antibody [EP229] (ab109454)

    This data was developed using the same antibody clone in a different buffer formulation - BSA and Azide free (ab208104).

    Recommended concentration for ab109454: 1/200 dilution.

    The human prostate cancer sections were performed by Heat mediated antigen retrieval with Tris-EDTA buffer (pH 9.0, epitope retrieval solution2) for 20 mins, then treated with or without alkaline phosphatase. After alkaline phosphatase treatment, the sections were labelled with ab208104 at 1/2000 dilution for 30 mins at room temperature. The Rabbit specific IHC polymer detection kit HRP/DAB (ab209101) were used as secondary antibody. The sections Counterstained with hematoxylin. The immunostaining was performed on a Leica Biosystems BOND® RX instrument.

    Human prostate cancer without alkaline phosphatase treatment (image A), no signal was detected when treated with alkaline phosphatase (image B).

  • Dot Blot - Anti-PTEN (phospho T366) antibody [EP229] (ab109454)
    Dot Blot - Anti-PTEN (phospho T366) antibody [EP229] (ab109454)

    This data was developed using the same antibody clone in a different buffer formulation - BSA and Azide free (ab208104).

    Recommended concentration for ab109454: 1/1000 dilution.

    Dot blot analysis of PTEN (pT366) peptide (Lane 1), and PTEN non-phospho peptide (Lane 2) incubated with ab208104 at a dilution of 1/10000.
    ab97051 (Peroxidase conjugated goat anti-rabbit IgG (H+L)) was used as the secondary antibody at a dilution of 1/100,000.

    Exposure time: 180s

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

文献 (13)

发表研究结果有使用 ab109454？请让我们知道，以便我们可以引用本数据表中的参考文章。

ab109454 被引用在 13 文献中.

  • Zhang T  et al. Gut microbiota relieves inflammation in the substantia nigra of chronic Parkinson's disease by protecting the function of dopamine neurons. Exp Ther Med 23:52 (2022). PubMed: 34934429
  • Xing Y  et al. Methylated Vnn1 at promoter regions induces asthma occurrence via the PI3K/Akt/NF?B-mediated inflammation in IUGR mice. Biol Open 9:N/A (2020). PubMed: 32139393
  • Li B  et al. The MSC-Derived Exosomal lncRNA H19 Promotes Wound Healing in Diabetic Foot Ulcers by Upregulating PTEN via MicroRNA-152-3p. Mol Ther Nucleic Acids 19:814-826 (2020). PubMed: 31958697
  • Engqvist H  et al. Immunohistochemical validation of COL3A1, GPR158 and PITHD1 as prognostic biomarkers in early-stage ovarian carcinomas. BMC Cancer 19:928 (2019). PubMed: 31533654
  • 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). PubMed: 30964887
View all Publications for this product

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1-3 of 3 Abreviews or Q&A

Question

As per our conversation, I have attached a powerpoint containing two images, one for p-PTEN and one for PTEN. Notice that the PTEN image contains a band that appears to be appropriately at 54kda, but the p-PTEN image contains two bands that are above and below band for PTEN.

I used 50 micrograms of protein, and the cells used were MCF-7.

Thanks,

Read More
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

Question

I edited the powerpoint to indicate which lysates were not treated (-) and which were treated with Trastuzumab (+). Each of the six pairs of lanes contains a different transfected variant of the MCF-7 cell line (e.g. MCF-7, MCF-7 with EGFR knockout, etc). The same samples correspond to the lanes for the ab109454 image.

Please let me know what other information you require.

Thanks,

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

Strong bands around 50 and 58 kDa using treated and untreated MCF7 cell lysates.

Read More
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!

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