Validated using a knockout cell line
Recombinant
RabMAb

Recombinant Anti-PI 3 Kinase p85 beta antibody [EPR18416] (ab180967)

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Key features and details

  • Produced recombinantly (animal-free) for high batch-to-batch consistency and long term security of supply
  • Rabbit monoclonal [EPR18416] to PI 3 Kinase p85 beta
  • Suitable for: IHC-P, WB, ICC/IF, IP
  • Knockout validated
  • Reacts with: Rat, Human

Overview

  • Product name

    Anti-PI 3 Kinase p85 beta antibody [EPR18416]
    See all PI 3 Kinase p85 beta primary antibodies

  • Description

    Rabbit monoclonal [EPR18416] to PI 3 Kinase p85 beta

  • Host species

    Rabbit

  • Tested applications

    Suitable for: IHC-P, WB, ICC/IF, IPmore details
    Unsuitable for: Flow Cyt

  • Species reactivity

    Reacts with: Rat, Human

  • Immunogen

    Synthetic peptide within Human PI 3 Kinase p85 beta aa 100-200. The exact sequence is proprietary.
    Database link: O00459

  • Positive control

    • WB: HAP1, HeLa, Jurkat, HEK293 cell lysates; Human fetal brain and fetal kidney lysates; Rat brain and spleen lysates; PC12 cell lysate. IHC-P: Human colonic adenocarcinoma, Human colon, rat testis tissues. ICC/IF: HeLa and Jurkat cells. IP: Jurkat cell lysate.

  • General notes

     

     

    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.

Properties

Applications

Our Abpromise guarantee covers the use of ab180967 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
IHC-P 1/100. Perform heat mediated antigen retrieval with Tris/EDTA buffer pH 9.0 before commencing with IHC staining protocol.
WB 1/2000. Detects a band of approximately 82 kDa (predicted molecular weight: 82 kDa).
ICC/IF 1/500.
IP 1/80.
  • Application notes
    Is unsuitable for Flow Cyt.

    • Target

    • Function

      Regulatory subunit of phosphoinositide-3-kinase (PI3K), a kinase that phosphorylates PtdIns(4,5)P2 (Phosphatidylinositol 4,5-bisphosphate) to generate phosphatidylinositol 3,4,5-trisphosphate (PIP3). PIP3 plays a key role by recruiting PH domain-containing proteins to the membrane, including AKT1 and PDPK1, activating signaling cascades involved in cell growth, survival, proliferation, motility and morphology. Binds to activated (phosphorylated) protein-tyrosine kinases, through its SH2 domain, and acts as an adapter, mediating the association of the p110 catalytic unit to the plasma membrane. Indirectly regulates autophagy (PubMed:23604317). Promotes nuclear translocation of XBP1 isoform 2 in a ER stress- and/or insulin-dependent manner during metabolic overloading in the liver and hence plays a role in glucose tolerance improvement.

    • Involvement in disease

      Megalencephaly-polymicrogyria-polydactyly-hydrocephalus syndrome 1

    • Sequence similarities

      Belongs to the PI3K p85 subunit family.
      Contains 1 Rho-GAP domain.
      Contains 2 SH2 domains.
      Contains 1 SH3 domain.

    • Domain

      The SH2 2 domain is required for interaction with FBXL2 and PTPN13.

    • Post-translational
      modifications

      Phosphorylated in response to signaling from activated receptor-type protein kinases (PubMed:19690332, PubMed:20068231). Dephosphorylated by PTPRJ (PubMed:18348712). Dephosphorylated at Tyr-655 by PTPN13. Phosphorylation of Tyr-655 impairs while its dephosphorylation promotes interaction with FBXL2 and SCF(FBXL2)-mediated polyubiquitination (PubMed:23604317).
      Ubiquitinated. Polyubiquitination by the SCF(FBXL2) complex probably promotes proteasomal degradation of PIK3R2.
    • Information by UniProt

    • Database links

      • Alternative names

        • p85 antibody
        • p85 beta antibody
        • P85B antibody
        • P85B_HUMAN antibody
        • Phosphatidylinositol 3 kinase antibody
        • Phosphatidylinositol 3 kinase regulatory beta subunit antibody
        • Phosphatidylinositol 3 kinase regulatory subunit beta antibody
        • Phosphatidylinositol 3 kinase regulatory subunit polypeptide 2 antibody
        • Phosphatidylinositol 3 kinase, regulatory subunit, polypeptide 2 (p85 beta) antibody
        • Phosphatidylinositol 3-kinase 85 kDa regulatory subunit beta antibody
        • Phosphatidylinositol 3-kinase regulatory subunit beta antibody
        • Phosphoinositide 3 kinase regulatory subunit 2 (beta) antibody
        • Phosphoinositide 3 kinase regulatory subunit 2 antibody
        • Phosphoinositide 3 kinase regulatory subunit polypeptide 2 (p85 beta) antibody
        • Phosphoinositide 3 kinase regulatory subunit polypeptide 2 antibody
        • Phosphoinositide 3 kinase, regulatory subunit 2 (beta) antibody
        • Phosphoinositide 3 kinase, regulatory subunit 2 (p85 beta) antibody
        • PI3 kinase p85 beta subunit antibody
        • PI3 kinase p85 subunit beta antibody
        • PI3-kinase regulatory subunit beta antibody
        • PI3-kinase subunit p85-beta antibody
        • PI3K antibody
        • PI3K regulatory subunit beta antibody
        • PIK3R 2 antibody
        • PIK3R2 antibody
        • PtdIns 3 kinase p85 beta antibody
        • PtdIns-3-kinase regulatory subunit beta antibody
        • PtdIns-3-kinase regulatory subunit p85-beta antibody
        see all

      Images

      • Western blot - Anti-PI 3 Kinase p85 beta antibody [EPR18416] (ab180967)
        Western blot - Anti-PI 3 Kinase p85 beta antibody [EPR18416] (ab180967)
        All lanes : Anti-PI 3 Kinase p85 beta antibody [EPR18416] (ab180967) at 1/2000 dilution

        Lane 1 : Wild-type HEK-293T cell lysate
        Lane 2 : PIK3R2 knockout HEK-293T cell lysate

        Lysates/proteins at 20 µg per lane.

        Performed under reducing conditions.

        Predicted band size: 82 kDa
        Observed band size: 85 kDa
        why is the actual band size different from the predicted?



        Lanes 1- 2: Merged signal (red and green). Green - ab180967 observed at 85 kDa. Red - Anti-GAPDH antibody [6C5] - Loading Control (ab8245) observed at 37 kDa.

         ab180967 was shown to react with PI 3 Kinase p85 beta in wild-type HEK-293T cells in western blot. Loss of signal was observed when knockout cell line ab266799 (knockout cell lysate ab257586) was used. Wild-type HEK-293T and PIK3R2 knockout HEK-293T cell lysates were subjected to SDS-PAGE. ab180967 and Anti-GAPDH antibody [6C5] - Loading Control (ab8245) overnight at 4°C at a 1 in 2000 dilution and a 1 in 20000 dilution respectively. Blots were developed with Goat anti-Rabbit IgG H&L (IRDye®800CW) preadsorbed (ab216773) and Goat anti-Mouse IgG H&L (IRDye®680RD) preadsorbed (ab216776) secondary antibodies at 1 in 20000 dilution for 1 hour at room temperature before imaging.

      • Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-PI 3 Kinase p85 beta antibody [EPR18416] (ab180967)
        Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-PI 3 Kinase p85 beta antibody [EPR18416] (ab180967)

        Immunohistochemical analysis of paraffin-embedded Human colon tissue labeling PI 3 Kinase p85 beta using ab180967 at 1/100 dilution. A Goat Anti-Rabbit IgG H&L (HRP) (ab97051) was used as secondary at 1/500 dilution. Cytoplasm staining on epithelial cells of Human colon was observed. Counter stained with Hematoxylin.

        Negative control: Used PBS instead of primary antibody, secondary antibody is Goat Anti-Rabbit IgG H&L (HRP) (ab97051) at 1/500 dilution.

        Perform heat mediated antigen retrieval with Tris/EDTA buffer pH 9.0 before commencing with IHC staining protocol.

      • Immunocytochemistry/ Immunofluorescence - Anti-PI 3 Kinase p85 beta antibody [EPR18416] (ab180967)
        Immunocytochemistry/ Immunofluorescence - Anti-PI 3 Kinase p85 beta antibody [EPR18416] (ab180967)

        Immunofluorescent analysis of 4% paraformaldehyde-fixed, 0.1% Triton X-100 permeabilized HeLa (Human epithelial cells from cervix adenocarcinoma) cells labeling PI 3 Kinase p85 beta with ab180967 at 1/500 dilution, followed by Goat anti-rabbit IgG (Alexa Fluor® 488) (ab150077) secondary antibody at 1/1000 dilution (green).

        Confocal image showing nuclear and cytoplasmic staining on HeLa cell line.

        The nuclear counter stain is DAPI (blue).

        Tubulin is detected with ab7291 (anti-Tubulin mouse mAb) at 1/1000 dilution and ab150120 (AlexaFluor®594 Goat anti-Mouse secondary) at 1/1000 dilution (red).
        The negative controls are as follows:
        1. ab180967 at 1/500 dilution followed by ab150120 (AlexaFluor®594 Goat anti-Mouse secondary) at 1/1000 dilution.
        2. ab7291 (anti-Tubulin mouse mAb) at 1/1000 dilution followed by ab150077 (Alexa Fluor®488 Goat Anti-Rabbit IgG H&L) at 1/1000 dilution.

      • Western blot - Anti-PI 3 Kinase p85 beta antibody [EPR18416] (ab180967)
        Western blot - Anti-PI 3 Kinase p85 beta antibody [EPR18416] (ab180967)

        Lane 1: Wild type HAP1 whole cell lysate (20 µg)
        Lane 2: PIK3R2 knockout HAP1 whole cell lysate (20 µg)
        Lane 3: HeLa whole cell lysate (20 µg)
        Lanes 1 - 3: Merged signal (red and green). Green - ab180967 observed at 85 kDa. Red - loading control, ab8245, observed at 37 kDa.

        ab180967 was shown to specifically react with PIK3R2 (KO) when PIK3R2 (KO) knockout samples were used. Wild-type and PIK3R2 (KO) knockout samples were subjected to SDS-PAGE. Ab180967 and ab8245 (Mouse anti GAPDH loading control) were incubated overnight at 4°C at 2000 dilution and 1/10000 dilution respectively. Blots were developed with Goat anti-Rabbit IgG H&L (IRDye® 800CW) preabsorbed ab216773 and Goat anti-Mouse IgG H&L (IRDye® 680RD) preabsorbed ab216776 secondary antibodies at 1/10000 dilution for 1 hour at room temperature before imaging.

      • Western blot - Anti-PI 3 Kinase p85 beta antibody [EPR18416] (ab180967)
        Western blot - Anti-PI 3 Kinase p85 beta antibody [EPR18416] (ab180967)
        All lanes : Anti-PI 3 Kinase p85 beta antibody [EPR18416] (ab180967) at 1/2000 dilution

        Lane 1 : HeLa (Human epithelial cells from cervix adenocarcinoma) whole cell lysate
        Lane 2 : Jurkat (Human T cell leukemia cells from peripheral blood) whole cell lysate
        Lane 3 : HEK293 (Human epithelial cells from embryonic kidney) whole cell lysate

        Lysates/proteins at 20 µg per lane.

        Secondary
        All lanes : Goat Anti-Rabbit IgG H&L (HRP) (ab97051) at 1/1000 dilution

        Predicted band size: 82 kDa
        Observed band size: 82 kDa


        Exposure time: 30 seconds


        5% NFDM/TBST: Blocking and diluting buffer.

      • Western blot - Anti-PI 3 Kinase p85 beta antibody [EPR18416] (ab180967)
        Western blot - Anti-PI 3 Kinase p85 beta antibody [EPR18416] (ab180967)
        All lanes : Anti-PI 3 Kinase p85 beta antibody [EPR18416] (ab180967) at 1/2000 dilution

        Lane 1 : Human fetal brain lysate
        Lane 2 : Human fetal kidney lysate

        Lysates/proteins at 10 µg per lane.

        Secondary
        All lanes : Anti-Rabbit IgG (HRP), specific to the non-reduced form of IgG at 1/1000 dilution

        Predicted band size: 82 kDa
        Observed band size: 82 kDa


        Exposure time: 3 minutes


        5% NFDM/TBST: Blocking and diluting buffer.

      • Western blot - Anti-PI 3 Kinase p85 beta antibody [EPR18416] (ab180967)
        Western blot - Anti-PI 3 Kinase p85 beta antibody [EPR18416] (ab180967)
        All lanes : Anti-PI 3 Kinase p85 beta antibody [EPR18416] (ab180967) at 1/2000 dilution

        Lane 1 : Rat brain lysate
        Lane 2 : Rat spleen lysate
        Lane 3 : PC-12 (Rat adrenal gland pheochromocytoma) whole cell lysate

        Lysates/proteins at 10 µg per lane.

        Secondary
        All lanes : Goat Anti-Rabbit IgG H&L (HRP) (ab97051) at 1/1000 dilution

        Predicted band size: 82 kDa
        Observed band size: 82 kDa


        Exposure time: 30 seconds


        5% NFDM/TBST: Blocking and diluting buffer.

      • Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-PI 3 Kinase p85 beta antibody [EPR18416] (ab180967)
        Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-PI 3 Kinase p85 beta antibody [EPR18416] (ab180967)

        Immunohistochemical analysis of paraffin-embedded Human colonic adenocarcinoma tissue labeling PI 3 Kinase p85 beta using ab180967 at 1/100 dilution. A Goat Anti-Rabbit IgG H&L (HRP) (ab97051) was used as secondary at 1/500 dilution. Nucleus and cytoplasm staining on tumor cells of colonic adenocarcinoma was observed. Counter stained with Hematoxylin.

        Negative control: Used PBS instead of primary antibody, secondary antibody is Goat Anti-Rabbit IgG H&L (HRP) (ab97051) at 1/500 dilution.

        Perform heat mediated antigen retrieval with Tris/EDTA buffer pH 9.0 before commencing with IHC staining protocol.

      • Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-PI 3 Kinase p85 beta antibody [EPR18416] (ab180967)
        Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-PI 3 Kinase p85 beta antibody [EPR18416] (ab180967)

        Immunohistochemical analysis of paraffin-embedded Rat testis tissue labeling PI 3 Kinase p85 beta using ab180967 at 1/100 dilution. A Goat Anti-Rabbit IgG H&L (HRP) (ab97051) was used as secondary at 1/500 dilution. Nucleus and cytoplasm staining on spermatogenic cell of rat testis was observed. Counter stained with Hematoxylin.

        Negative control: Used PBS instead of primary antibody, secondary antibody is Goat Anti-Rabbit IgG H&L (HRP) (ab97051) at 1/500 dilution.

        Perform heat mediated antigen retrieval with Tris/EDTA buffer pH 9.0 before commencing with IHC staining protocol.

      • Immunocytochemistry/ Immunofluorescence - Anti-PI 3 Kinase p85 beta antibody [EPR18416] (ab180967)
        Immunocytochemistry/ Immunofluorescence - Anti-PI 3 Kinase p85 beta antibody [EPR18416] (ab180967)

        Immunofluorescent analysis of 4% paraformaldehyde-fixed, 0.1% Triton X-100 permeabilized Jurkat (Human T cell leukemia cells from peripheral blood) cells labeling PI 3 Kinase p85 beta with ab180967 at 1/500 dilution, followed by Goat anti-rabbit IgG (Alexa Fluor® 488) (ab150077) secondary antibody at 1/1000 dilution (green).

        Confocal image showing nuclear and weakly cytoplasmic staining on Jurkat cell line.

        The nuclear counter stain is DAPI (blue).

        Tubulin is detected with ab7291 (anti-Tubulin mouse mAb) at 1/1000 dilution and ab150120 (AlexaFluor®594 Goat anti-Mouse secondary) at 1/1000 dilution (red).
        The negative controls are as follows:
        1. ab180967 at 1/500 dilution followed by ab150120 (AlexaFluor®594 Goat anti-Mouse secondary) at 1/1000 dilution.
        2. ab7291 (anti-Tubulin mouse mAb) at 1/1000 dilution followed by ab150077 (Alexa Fluor®488 Goat Anti-Rabbit IgG H&L) at 1/1000 dilution.

      • Immunoprecipitation - Anti-PI 3 Kinase p85 beta antibody [EPR18416] (ab180967)
        Immunoprecipitation - Anti-PI 3 Kinase p85 beta antibody [EPR18416] (ab180967)

        PI 3 Kinase p85 beta was immunoprecipitated from 1mg of HeLa (Human epithelial cells from cervix adenocarcinoma) whole cell lysate with ab180967 at 1/80 dilution. Western blot was performed from the immunoprecipitate using ab180967 at 1/10000 dilution. Anti-Rabbit IgG (HRP), specific to the non-reduced form of IgG, was used as secondary antibody at 1/1500 dilution.

        Lane 1: HeLa whole cell lysate10 µg (Input).

        Lane 2: ab180967 IP in HeLa whole cell lysate.

        Lane 3: Rabbit monoclonal IgG (ab172730) instead of ab180967 in HeLa whole cell lysate.
        Blocking and dilution buffer and concentration: 5% NFDM/TBST.

        Exposure time: 10 seconds.

      References (7)

      Publishing research using ab180967? Please let us know so that we can cite the reference in this datasheet.

      ab180967 has been referenced in 7 publications.

      • Yue Z  et al. Diallyl Disulfide Induces Apoptosis and Autophagy in Human Osteosarcoma MG-63 Cells through the PI3K/Akt/mTOR Pathway. Molecules 24:N/A (2019). PubMed: 31340526
      • Hao P  et al. UBE2T promotes proliferation and regulates PI3K/Akt signaling in renal cell carcinoma. Mol Med Rep 20:1212-1220 (2019). PubMed: 31173226
      • Ding X  et al. Long non-coding RNA FOXO1 inhibits lung cancer cell growth through down-regulating PI3K/AKT signaling pathway. Iran J Basic Med Sci 22:491-498 (2019). PubMed: 31217928
      • Du Q  et al. Propofol inhibits proliferation and epithelial-mesenchymal transition of MCF-7 cells by suppressing miR-21 expression. Artif Cells Nanomed Biotechnol 47:1265-1271 (2019). PubMed: 30942630
      • Zhang Y  et al. Ailanthone up-regulates miR-449a to restrain acute myeloid leukemia cells growth, migration and invasion. Exp Mol Pathol 108:114-120 (2019). PubMed: 31002772
      • Leng X  et al. Low-intensity pulsed ultrasound promotes proliferation and migration of HaCaT keratinocytes through the PI3K/AKT and JNK pathways. Braz J Med Biol Res 51:e7862 (2018). PubMed: 30365726
      • Amaral C  et al. Hormone-dependent breast cancer: Targeting autophagy and PI3K overcomes Exemestane-acquired resistance. J Steroid Biochem Mol Biol N/A:N/A (2018). WB . PubMed: 29791862

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