Overview

  • Product name

    Mouse mTOR ELISA Kit
    See all mTOR kits
  • Detection method

    Colorimetric
  • Precision

    Intra-assay
    Sample n Mean SD CV%
    cell lysate 8 = 3.1%
    Inter-assay
    Sample n Mean SD CV%
    cell lysate 3 = 6.9%
  • Sample type

    Cell culture supernatant, Cell culture extracts, Tissue Extracts, Tissue Homogenate
  • Assay type

    Sandwich (quantitative)
  • Sensitivity

    16.7 pg/ml
  • Range

    39.06 pg/ml - 2500 pg/ml
  • Recovery

    Sample specific recovery
    Sample type Average % Range
    Cell culture media 97 94% - 101%

  • Assay time

    1h 30m
  • Assay duration

    One step assay
  • Species reactivity

    Reacts with: Mouse, Rat, Human
  • Product overview

    Abcam’s mTOR in vitro SimpleStep ELISA® (Enzyme-Linked Immunosorbent Assay) kit is designed for the quantitative measurement of mTOR protein in mousecell culture supernatants, cell and tissue extracts.

    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.

    Sensitivity:

    Samples in Sample Diluent NS: 16.7pg/mL

    Samples in 1X Cell Extraction Buffer PTR: 30.7pg/mL

  • Notes

    Mammalian target of rapamycin (mTOR) is a serine/threonine protein kinase part of two distinct signaling complexes, mTORC1 and mTORC2. These two complexes share four proteins (mTOR, mLST8, DEPTOR, Tti1/tel2), with only mTORC1 containing Raptor and PRAS40 and mTORC2 containing Rictor, mSin1 and Protor1/2. The complex mTORC1 (rapamycin sensitive complex) coordinates inputs from growth factors, stress, energy status, oxygen and amino acids levels to control processes such as protein and lipid synthesis and autophagy. The complex mTORC2 is insensitive to nutrients and rapamycin, but it responds to insulin signaling. It also controls ion transport and cell shape by targeting serum/glucocorticoid protein kinase (SGK1) and protein kinase (PKC-α) respectively.

    The canonical regulation of mTORC1 occurs through the TSC/Rheb pathway which receives signals from AKT, AMPK and IKKβ to activate the complex. Phosphorylation of mTOR at Ser2448 is carried out directly by AKT kinase as well as p70S6 kinase acting as a feedback signal. Phosphorylation at this site is a biomarker for the activation state of the PI-3 kinase pathway as well as the activation status of mTOR. Activation of mTOR leads to phosphorylation of PRAS40, raptor and DEPTOR and the consequential activation of mTORC1. Deregulated signaling of mTOR has been implicated in diseases such as cancer, metabolic syndrome, neurodegeneration and aging. Constitutive activation of PI3K-mTORC1 signaling in cancer cells inhibits autophagy, deregulates protein synthesis via 4E-BP1/eIF4E and increases de novo lipid synthesis via SREBP1. Similarly mTOR signaling is a key factor in the regulation of tissue metabolism in the normal and nutrient overload state affecting the hypothalamus, adipose tissue, the liver, skeletal muscle and pancreas.Notably, rat and human mTOR are 99.5% and 98.9% identical to mouse mTOR, 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 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
    ab221824 - Antibody Diluent 4BI 1 x 6ml
    Mouse mTOR Capture Antibody (lyophilized) 1 vial
    Mouse mTOR Detector Antibody (lyophilized) 1 vial
    Mouse mTOR Lyophilized Recombinant Protein 2 vials
    Plate Seals 1 unit
    Sample Diluent NS 1 x 50ml
    SimpleStep Pre-Coated 96-Well Microplate (ab206978) 1 unit
    Stop Solution 1 x 12ml
    TMB Development Solution 1 x 12ml
  • Research areas

  • Function

    Kinase subunit of both mTORC1 and mTORC2, 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 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-421', which then promotes protein synthesis by phosphorylating PDCD4 and targeting it for degradation. Phosphorylates MAF1 leading to attenuation of its RNA polymerase III-repressive function. mTORC2 is also activated by growth. factors, but seems to be nutrient-insensitive. mTORC2 seems to function upstream of Rho GTPases to regulate the actin cytoskeleton, probably by activating one or more Rho-type guanine nucleotide exchange factors. mTORC2 promotes the serum-induced formation of stress-fibers or F-actin. mTORC2 plays a critical role in AKT1 'Ser-473' phosphorylation, which may facilitate the phosphorylation of the activation loop of AKT1 on 'Thr-308' by PDK1 which is a prerequisite for full activation. mTORC2 regulates the phosphorylation of SGK1 at 'Ser-422'. mTORC2 also modulates the phosphorylation of PRKCA on 'Ser-657'.
  • Tissue specificity

    Expressed in numerous tissues, with highest levels in testis.
  • Sequence similarities

    Belongs to the PI3/PI4-kinase family.
    Contains 1 FAT domain.
    Contains 1 FATC domain.
    Contains 7 HEAT repeats.
    Contains 1 PI3K/PI4K domain.
  • Post-translational
    modifications

    Autophosphorylated; when part of mTORC1 or mTORC2.
  • Cellular localization

    Endoplasmic reticulum membrane. Golgi apparatus membrane. Mitochondrion outer membrane. Lysosome. Cytoplasm. Nucleus > PML body. Shuttles between cytoplasm and nucleus. Accumulates in the nucleus in response to hypoxia (By similarity). Targeting to lysosomes depends on amino acid availability and RRAGA and RRAGB.
  • Information by UniProt
  • Alternative names

    • dJ576K7.1 (FK506 binding protein 12 rapamycin associated protein 1)
    • FK506 binding protein 12 rapamycin associated protein 1
    • FK506 binding protein 12 rapamycin associated protein 2
    • FK506 binding protein 12 rapamycin complex associated protein 1
    • FK506-binding protein 12-rapamycin complex-associated protein 1
    • FKBP rapamycin associated protein
    • FKBP12 rapamycin complex associated protein
    • FKBP12-rapamycin complex-associated protein
    • FKBP12-rapamycin complex-associated protein 1
    • FLJ44809
    • FRAP
    • FRAP1
    • FRAP2
    • Mammalian target of rapamycin
    • Mechanistic target of rapamycin
    • mTOR
    • MTOR_HUMAN
    • OTTHUMP00000001983
    • RAFT1
    • Rapamycin and FKBP12 target 1
    • Rapamycin associated protein FRAP2
    • Rapamycin target protein
    • Rapamycin target protein 1
    • RAPT1
    • Serine/threonine-protein kinase mTOR
    see all
  • Database links

Applications

Our Abpromise guarantee covers the use of ab206311 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.

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

  • Native mouse mTOR was measured in 600 μg/mL NIH 3T3 cell extractand 200 μg/mL C2C12 cell extractdiluted in a 2-fold dilution series in 1X Cell Extraction Buffer PTR. Native rat mTOR was measured in 200 μg/mL PC-12 cell extractdiluted in a 2-fold dilution series in 1X Cell Extraction Buffer PTR. The concentrations of mouse and rat mTOR were measured in duplicate and interpolated from the mousemTOR standard curve and corrected for sample dilution. The interpolated dilution factor corrected values are graphed (mean +/- SD).

  • Recombinant mouse mTOR was spiked into 10% RPMI culture media and diluted in a 2-fold dilution series in Sample Diluent NS. The concentrations of mTOR were measured in duplicate and interpolated from the mouse mTOR standard curve and corrected for sample dilution. The interpolated dilution factor corrected values are graphed (mean +/- SD).

Protocols

References

ab206311 has not yet been referenced specifically in any publications.

Customer reviews and Q&As

mTOR elisa kit

Excellent Excellent 5/5 (Ease of Use)
Abreviews
mTOR ELISA was performed on whole macrophages sorted from MC38 tumors excised from B6 mice. Total of 20 ug of protein was loaded in each well in the ELISA.

black bar: non-obese mice
teal bar: obese mice

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Verified customer

Submitted Aug 12 2019

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