Validated using a knockout cell line
Recombinant
RabMAb

Recombinant Anti-PI 3 Kinase p85 alpha antibody [EPR18702] - BSA and Azide free (ab223792)

Overview

  • Product name

    Anti-PI 3 Kinase p85 alpha antibody [EPR18702] - BSA and Azide free
    See all PI 3 Kinase p85 alpha primary antibodies
  • Description

    Rabbit monoclonal [EPR18702] to PI 3 Kinase p85 alpha - BSA and Azide free
  • Host species

    Rabbit
  • Tested applications

    Suitable for: Flow Cyt, ICC/IF, IP, WBmore details
  • Species reactivity

    Reacts with: Mouse, Rat, Human
  • Immunogen

    Synthetic peptide within Rat PI 3 Kinase p85 alpha aa 600-700. The exact sequence is proprietary.
    Database link: Q63787

  • Positive control

    • WB: Human PI3K p85 alpha full length recombinant protein; Human fetal liver, fetal heart and fetal kidney lysates; HepG2, MCF7, Raji, Jurkat, C6, RAW 264.7, PC-12 and NIH/3T3 whole cell lysates; Mouse brain, heart, kidney and spleen lysates; Rat brain, heart, kidney and spleen lysates. ICC/IF: HepG2 and NIH/3T3 cells. Flow Cyt: NIH/3T3 cells; IP: MCF7 whole cell lysate.
  • General notes

    Ab223792 is the carrier-free version of ab191606. This format is designed for use in antibody labeling, including fluorochromes, metal isotopes, oligonucleotides, enzymes.

     

    Our carrier-free formats are supplied in a buffer free of BSA, sodium azide and glycerol for higher conjugation efficiency.

    Use our conjugation kits  for antibody conjugates that are ready-to-use in as little as 20 minutes with <1 minute hands-on-time and 100% antibody recovery: available for fluorescent dyes, HRP, biotin and gold.

    ab223792 is compatible with the Maxpar® Antibody Labeling Kit from Fluidigm.

    Maxpar® is a trademark of Fluidigm Canada Inc.

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

    This product is a recombinant rabbit monoclonal antibody.

Properties

Applications

Our Abpromise guarantee covers the use of ab223792 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 Use at an assay dependent concentration.

ab199376 - Rabbit monoclonal IgG, is suitable for use as an isotype control with this antibody.

 

ICC/IF Use at an assay dependent concentration.
IP Use at an assay dependent concentration.
WB Use at an assay dependent concentration. Detects a band of approximately 85,46 kDa (predicted molecular weight: 84 kDa).

Target

  • Function

    Binds to activated (phosphorylated) protein-Tyr kinases, through its SH2 domain, and acts as an adapter, mediating the association of the p110 catalytic unit to the plasma membrane. Necessary for the insulin-stimulated increase in glucose uptake and glycogen synthesis in insulin-sensitive tissues.
  • Tissue specificity

    Isoform 2 is expressed in skeletal muscle and brain, and at lower levels in kidney and cardiac muscle. Isoform 2 and isoform 4 are present in skeletal muscle (at protein level).
  • Sequence similarities

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

    The SH3 domain mediates the binding to CBLB, and to HIV-1 Nef.
  • Post-translational
    modifications

    Polyubiquitinated in T-cells by CBLB; which does not promote proteasomal degradation but impairs association with CD28 and CD3Z upon T-cell activation.
    Phosphorylated. Dephosphorylated by PTPRJ.
  • Information by UniProt
  • Database links

  • Alternative names

    • GRB1 antibody
    • p85 alpha antibody
    • p85 antibody
    • P85A_HUMAN antibody
    • Phosphatidylinositol 3 kinase associated p 85 alpha antibody
    • Phosphatidylinositol 3 kinase regulatory 1 antibody
    • Phosphatidylinositol 3 kinase, regulatory subunit, polypeptide 1 (p85 alpha) antibody
    • Phosphatidylinositol 3-kinase 85 kDa regulatory subunit alpha antibody
    • Phosphatidylinositol 3-kinase regulatory subunit alpha antibody
    • Phosphoinositide 3 kinase, regulatory subunit 1 (alpha) antibody
    • PI3 kinase p85 antibody
    • PI3 kinase p85 subunit alpha antibody
    • PI3-kinase regulatory subunit alpha antibody
    • PI3-kinase subunit p85-alpha antibody
    • PI3K antibody
    • PI3K p85 antibody
    • PI3K regulatory subunit alpha antibody
    • Pik3r1 antibody
    • PtdIns 3 kinase p85 alpha antibody
    • PtdIns-3-kinase regulatory subunit alpha antibody
    • PtdIns-3-kinase regulatory subunit p85-alpha antibody
    see all

Images

  • This WB data was generated using the same anti-PI 3 Kinase p85 alpha antibody clone [EPR18702] in a different buffer formulation containing PBS, BSA, glycerol, and sodium azide (cat# ab191606).

    Lane 1: Wild type HAP1 whole cell lysate (20 µg)
    Lane 2: PIK3R1 knockout HAP1 whole cell lysate (20 µg)
    Lane 3: Jurkat whole cell lysate (20 µg)

    Lanes 1 - 4: Merged signal (red and green). Green - ab191606 observed at 90 kDa. Red - loading control, ab9484, observed at 37 kDa.

    ab191606 was shown to specifically react with PIK3R1 when PIK3R1 knockout samples were used. Wild-type and PIK3R1 knockout samples were subjected to SDS-PAGE. Ab191606 and ab9484 (Mouse anti GAPDH loading control) were incubated overnight at 4°C at 1000 dilution and 1/20000 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 1/20000 dilution for 1 hour at room temperature before imaging.

  • Overlay histogram showing HepG2 cells fixed in 4% PFA and stained with ab191606 at a dilution of 1/80 (red line). The secondary antibody used was Alexa Fluor® 488 goat anti-rabbit at a dilution of 1/500. Rabbit monoclonal IgG (ab172730) was used as an isotype control (black line) and cells incubated in the absence of both primary and secondary antibody were used as a negative control (blue line).

    This data was developed using the same antibody clone in a different buffer formulation containing PBS, BSA, glycerol, and sodium azide (ab191606).

  • Immunofluorescent analysis of 4% paraformaldehyde-fixed, 0.1% Triton X-100 permeabilized HepG2 (Human liver hepatocellular carcinoma cell line) cells labeling PI3K p85 with ab191606 at 1/250 dilution, followed by Goat Anti-Rabbit IgG (Alexa Fluor® 488) (ab150077) secondary antibody at 1/1000 dilution (green). Confocal image showing cytoplasmic staining on HepG2 cell line. The nuclear counter stain is DAPI (blue). Tubulin is detected with Anti-alpha Tubulin antibody - Loading Control (ab7291)  at 1/1000 dilution and Goat Anti-Mouse IgG (AlexaFluor®594) preadsorbed (ab150120)  at 1/1000 dilution (red).
    The negative controls are as follows:
    -ve control 1: ab191606 at 1/500 dilution followed by ab150120  at 1/1000 dilution.
    -ve control 2: ab7291  at 1/1000 dilution followed by ab150077 at 1/1000 dilution.

    This data was developed using the same antibody clone in a different buffer formulation containing PBS, BSA, glycerol, and sodium azide (ab191606).

  • Immunofluorescent analysis of 4% paraformaldehyde-fixed, 100% Methanol permeabilized NIH/3T3 (Mouse embryonic fibroblast cell line) cells labeling PI3K p85 with ab191606 at 1/250 dilution, followed by Goat anti-rabbit IgG (Alexa Fluor® 488) (ab150077) secondary antibody at 1/1000 dilution (green). Confocal image showing cytoplasmic staining on NIH/3T3 cell line. The nuclear counter stain is DAPI (blue). Tubulin is detected with Anti-alpha Tubulin antibody - Loading Control (ab7291) at  1/1000 dilution and Goat Anti-Mouse IgG  (AlexaFluor®594) preadsorbed (ab150120) at 1/1000 dilution (red).
    The negative controls are as follows:
    -ve control 1: ab191606 at 1/500 dilution followed by ab150120  at 1/1000 dilution.
    -ve control 2: ab7291 at 1/1000 dilution followed by ab150077  at 1/1000 dilution.

    This data was developed using the same antibody clone in a different buffer formulation containing PBS, BSA, glycerol, and sodium azide (ab191606).

  • Flow cytometric analysis of 4% paraformaldehyde-fixed NIH/3T3 (Mouse embryonic fibroblast cell line) cells labeling PI3K p85 with ab191606 at 1/150 dilution (red) compared with a Rabbit IgG,monoclonal -  Isotype control (ab172730) (black) and an unlabelled control (cells without incubation with primary antibody and secondary antibody) (blue). Goat anti Rabbit IgG (Alexa Fluor® 488) at 1/500 dilution was used as the secondary antibody.

    This data was developed using the same antibody clone in a different buffer formulation containing PBS, BSA, glycerol, and sodium azide (ab191606).

  • PI3K p85 was immunoprecipitated from 1mg of MCF7 (Human breast adenocarcinoma cell line) whole cell lysate with ab191606 at 1/50 dilution. Western blot was performed from the immunoprecipitate using ab191606 at 1/1000 dilution. VeriBlot for IP Detection Reagent (HRP) (ab131366), was used for detection at 1/10000 dilution.

    Lane 1: MCF7 whole cell lysate, 10µg (Input).

    Lane 2: ab191606 IP in MCF7 whole cell lysate.

    Lane 3: Rabbit IgG, monoclonal - Isotype Control (ab172730) instead of ab191606 in MCF7 whole cell lysate.

    Blocking and dilution buffer and concentration: 5% NFDM/TBST.

    Exposure time: 5 seconds.

    This data was developed using the same antibody clone in a different buffer formulation containing PBS, BSA, glycerol, and sodium azide (ab191606).

References

ab223792 has not yet been referenced specifically in any publications.

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