Validated using a knockout cell line
Recombinant
RabMAb

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

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
  • Tested applications
    Suitable for: Flow Cyt, IHC-P, WB, ICC/IF, IPmore details
  • Species reactivity
    Reacts with: Rat, Human
  • Immunogen

    Synthetic peptide (the amino acid sequence is considered to be commercially sensitive) within Human PI 3 Kinase p85 beta aa 100-200. The exact sequence is proprietary.
    Database link: O00459

  • Positive control
    • WB: 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. Flow cytometry: HeLa cells. IP: Jurkat cell lysate.
  • General notes

    This product is a recombinant rabbit monoclonal antibody.

    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
Flow Cyt 1/250.
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.

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



  • Predicted band size : 82 kDa

    Lane 1: Wild type HAP1 whole cell lysate (20 µg)
    Lane 2: PIK3R2 (KO) knockout HAP1 whole cell lysate (20 µg)
    Lane 3: HeLa whole cell lysate (20
    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.

  • 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
    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.

  • 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
    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.

  • 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
    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.

  • 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.

  • 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.

  • 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.

  • 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.

  • 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.

  • Flow cytometric analysis of 4% paraformaldehyde-fixed HeLa (Human epithelial cells from cervix adenocarcinoma) cells labeling PI 3 Kinase p85 beta with ab180967 at 1/250 dilution (red) compared with a rabbit monoclonal IgG isotype control (ab172730, black) and a unlabelled control (cells without incubation with primary antibody and secondary antibody; blue). Goat anti rabbit IgG (FITC) at 1/500 dilution was used as the secondary antibody.

  • 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

ab180967 has not yet been referenced specifically in any publications.

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