Overview

  • Product name

    Mouse PRAS40 ELISA Kit
    See all PRAS40 kits
  • Detection method

    Colorimetric
  • Precision

    Intra-assay
    Sample n Mean SD CV%
    Cell extract 5 5.8%
    Inter-assay
    Sample n Mean SD CV%
    Cell extract 3 9.4%
  • Sample type

    Cell culture extracts, Tissue Extracts
  • Assay type

    Sandwich (quantitative)
  • Sensitivity

    23.5 pg/ml
  • Range

    156 pg/ml - 10000 pg/ml
  • Recovery

    Sample specific recovery
    Sample type Average % Range
    Cell culture extracts 100 96% - 104%

  • Assay time

    1h 30m
  • Assay duration

    One step assay
  • Species reactivity

    Reacts with: Mouse, Rat, Human
  • Product overview

    PRAS40 in vitro SimpleStep ELISA® (Enzyme-Linked Immunosorbent Assay) kit is designed for the quantitative measurement of PRAS40 protein in cell and tissue extract.


    The SimpleStep ELISA® employs an affinity tag labeled capture antibody and a reporter conjugated detector antibody which immunocapture the sample analyte in solution. This entire complex (capture antibody/analyte/detector antibody) is in turn immobilized via immunoaffinity of an anti-tag antibody coating the well. To perform the assay, samples or standards are added to the wells, followed by the antibody mix. After incubation, the wells are washed to remove unbound material.  TMB substrate is added and during incubation is catalyzed by HRP, generating blue coloration. This reaction is then stopped by addition of Stop Solution completing any color change from blue to yellow. Signal is generated proportionally to the amount of bound analyte and the intensity is measured at 450 nm. Optionally, instead of the endpoint reading, development of TMB can be recorded kinetically at 600 nm.

  • Notes

    PRAS40 (also known as Proline-rich AKT1 substrate 1) is a subunit of the mTORC1 (mammalian target of rapamycin complex 1) protein complex which regulates cell growth and survival in response to nutrient and hormone signals.  PRAS40 is a direct phosphorylation substrate of AKT and PRAS40 is overexpressed in some cancers.  The standard protein in this kit is full length mouse PRAS40 expressed and purified from E.coli.  Mouse PRAS40 has 97% and 93% sequence identity to rat and human PRAS40, respectively.

  • Tested applications

    Suitable for: Sandwich ELISAmore details
  • Platform

    Pre-coated microplate (12 x 8 well strips)

Properties

  • Storage instructions

    Store at +4°C. Please refer to protocols.
  • Components 1 x 96 tests
    10X Mouse PRAS40 Capture Antibody 1 x 600µl
    10X Mouse PRAS40 Detector Antibody 1 x 600µl
    10X Wash Buffer PT (ab206977) 1 x 20ml
    50X Cell Extraction Enhancer Solution (ab193971) 1 x 1ml
    5X Cell Extraction Buffer PTR (ab193970) 1 x 10ml
    Antibody Diluent 5BR 1 x 6ml
    Mouse PRAS40 Lyophilized Recombinant Protein 2 vials
    Plate Seals 1 unit
    Sample Diluent NS 1 x 12ml
    SimpleStep Pre-Coated 96-Well Microplate (ab206978) 1 unit
    Stop Solution 1 x 12ml
    TMB Development Solution 1 x 12ml
  • Research areas

  • Function

    Subunit of mTORC1, which regulates cell growth and survival in response to nutrient and hormonal signals. mTORC1 is activated in response to growth factors or amino-acids. Growth factor-stimulated mTORC1 activation involves a AKT1-mediated phosphorylation of TSC1-TSC2, which leads to the activation of the RHEB GTPase that potently activates the protein kinase activity of mTORC1. Amino-acid-signaling to mTORC1 requires its relocalization to the lysosomes mediated by the Ragulator complex and the Rag GTPases. Activated mTORC1 up-regulates protein synthesis by phosphorylating key regulators of mRNA translation and ribosome synthesis. mTORC1 phosphorylates EIF4EBP1 and releases it from inhibiting the elongation initiation factor 4E (eiF4E). mTORC1 phosphorylates and activates S6K1 at 'Thr-389', which then promotes protein synthesis by phosphorylating PDCD4 and targeting it for degradation. Within mTORC1, AKT1S1 negatively regulates mTOR activity in a manner that is dependent on its phosphorylation state and binding to 14-3-3 proteins. Inhibits RHEB-GTP-dependent mTORC1 activation. Substrate for AKT1 phosphorylation, but can also be activated by AKT1-independent mechanisms. May also play a role in nerve growth factor-mediated neuroprotection.
  • Tissue specificity

    Widely expressed with highest levels of expression in liver and heart. Expressed at higher levels in cancer cell lines (e.g. A549 and HeLa) than in normal cell lines (e.g. HEK293).
  • Post-translational
    modifications

    Phosphorylated by AKT1. Phosphorylation relieves inhibitory function on mTORC1.
  • Cellular localization

    Cytoplasm > cytosol. Found in the cytosolic fraction of the brain.
  • Information by UniProt
  • Alternative names

    • 40 kDa proline rich AKT substrate
    • 40 kDa proline-rich AKT substrate
    • AKT1 S1
    • AKT1 substrate 1
    • AKT1 substrate 1 (proline rich)
    • AKT1S 1
    • AKT1S1
    • AKTS1_HUMAN
    • Lobe
    • MGC2865
    • PRAS
    • PRAS 40
    • PRAS40
    • Proline rich akt substrate
    • Proline rich Akt substrate 40 kDa
    • Proline-rich AKT1 substrate 1
    see all
  • Database links

Applications

Our Abpromise guarantee covers the use of ab210580 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
Sandwich ELISA Use at an assay dependent concentration.

Images

  • SimpleStep ELISA technology allows the formation of the antibody-antigen complex in one single step, reducing assay time to 90 minutes. Add samples or standards and antibody mix to wells all at once, incubate, wash, and add your final substrate. See protocol for a detailed step-by-step guide.

     

  • Background-subtracted data values (mean +/- SD) are graphed.

  • The concentrations of PRAS40 were measured in duplicate and interpolated from the PRAS40 standard curve and corrected for sample dilution. The interpolated dilution factor corrected values are plotted (mean +/- SD, n=2). The mean PRAS40 concentration was determined to be 6,256 pg/mL in NIH/3T3 extra, 1,480pg/mL in HELA extract, and 6,532pg/mL in PC12 extract samples.

Protocols

References

ab210580 has not yet been referenced specifically in any publications.

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