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RecombinantRabMAb

Recombinant Anti-Raptor antibody [EP539Y] (ab40768)

Reviews (3)Q&A (4)References (28)
Western blot - Anti-Raptor antibody [EP539Y] (ab40768)
  • Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-Raptor antibody [EP539Y] (ab40768)
  • Western blot - Anti-Raptor antibody [EP539Y] (ab40768)
  • Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-Raptor antibody [EP539Y] (ab40768)
  • Western blot - Anti-Raptor antibody [EP539Y] (ab40768)
  • Western blot - Anti-Raptor antibody [EP539Y] (ab40768)
  • Anti-Raptor antibody [EP539Y] (ab40768)

Key features and details

  • Produced recombinantly (animal-free) for high batch-to-batch consistency and long term security of supply
  • Rabbit monoclonal [EP539Y] to Raptor
  • Suitable for: WB, IHC-P
  • Reacts with: Mouse, Rat, Human

Conjugates logo Related conjugates and formulations

Carrier Free

Overview

  • Product name

    Anti-Raptor antibody [EP539Y]
    See all Raptor primary antibodies

  • Description

    Rabbit monoclonal [EP539Y] to Raptor

  • Host species

    Rabbit

  • Tested applications

    Suitable for: WB, IHC-Pmore details
    Unsuitable for: Flow Cyt,ICC/IF or IP

  • Species reactivity

    Reacts with: Mouse, Rat, Human

  • Immunogen

    Synthetic peptide within Human Raptor (C terminal). The exact sequence is proprietary.

  • Positive control

    • Normal human colon and Human cardiac muscle; mouse and rat brain tissue lysate HEK-293, HeLa, MCF-7 and A431 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.

    We are constantly working hard to ensure we provide our customers with best in class antibodies. As a result of this work we are pleased to now offer this antibody in purified format. We are in the process of updating our datasheets. The purified format is designated 'PUR' on our product labels. If you have any questions regarding this update, please contact our Scientific Support team.

Properties

Applications

The Abpromise guarantee

Our Abpromise guarantee covers the use of ab40768 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
WB (2)
1/1000 - 1/5000. Detects a band of approximately 140 kDa (predicted molecular weight: 149 kDa).

For unpurified use at 1/500. 
IHC-P
1/100. Perform heat mediated antigen retrieval with Tris/EDTA buffer pH 9.0 before commencing with IHC staining protocol.

See IHC antigen retrieval protocols.
Notes
WB
1/1000 - 1/5000. Detects a band of approximately 140 kDa (predicted molecular weight: 149 kDa).

For unpurified use at 1/500. 
IHC-P
1/100. Perform heat mediated antigen retrieval with Tris/EDTA buffer pH 9.0 before commencing with IHC staining protocol.

See IHC antigen retrieval protocols.
Application notes
Is unsuitable for Flow Cyt,ICC/IF or IP.

Target

  • Function

    Involved in the control of the mammalian target of rapamycin complex 1 (mTORC1) activity which regulates cell growth and survival, and autophagy in response to nutrient and hormonal signals; functions as a scaffold for recruiting mTORC1 substrates. 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.

  • Tissue specificity

    Highly expressed in skeletal muscle, and in a lesser extent in brain, lung, small intestine, kidney and placenta.

  • Sequence similarities

    Belongs to the WD repeat RAPTOR family.
    Contains 7 WD repeats.

  • Cellular localization

    Cytoplasm. Lysosome. Targeting to lysosomes depends on amino acid availability.
  • Information by UniProt

  • Database links

    • Alternative names

      • KIAA1303 antibody
      • KOG1 antibody
      • Mip1 antibody
      • P150 target of rapamycin (TOR) scaffold protein antibody
      • p150 target of rapamycin (TOR) scaffold protein containing WD repeats antibody
      • P150 target of rapamycin (TOR)-scaffold protein antibody
      • Raptor antibody
      • Regulatory associated protein of mTOR antibody
      • Regulatory associated protein of MTOR complex 1 antibody
      • Regulatory-associated protein of mTOR antibody
      • RPTOR antibody
      • RPTOR_HUMAN antibody
      see all

    Images

    • Western blot - Anti-Raptor antibody [EP539Y] (ab40768)
      Western blot - Anti-Raptor antibody [EP539Y] (ab40768)
      All lanes : Anti-Raptor antibody [EP539Y] (ab40768) at 1/5000 dilution

      Lane 1 : HEK-293 (human embryonic kidney) whole cell lysate
      Lane 2 : HeLa (human cervix adenocarcinoma) whole cell lysate
      Lane 3 : MCF-7 (human breast carcinoma) whole cell lysate

      Lysates/proteins at 20 µg per lane.

      Secondary
      All lanes : Goat Anti-Rabbit IgG H&L (HRP) (ab97051) at 1/20000 dilution

      Predicted band size: 149 kDa
      Observed band size: 140 kDa why is the actual band size different from the predicted?



      Blocking/Diluting buffer 5% NFDM /TBST
    • Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-Raptor antibody [EP539Y] (ab40768)
      Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-Raptor antibody [EP539Y] (ab40768)

      Immunohistochemical analysis of paraffin-embedded human cardiac muscle sections labelling Raptor with purified ab40768 at dilution of 1/100. The secondary antibody used was ab97051; a goat anti-rabbit IgG H&L (HRP) at dilution of 1/500. The sample was counterstained with hematoxylin. Antigen retrieval was performed using EDTA Buffer; pH 9.0. PBS was used instead of the primary antibody as the negative control and is shown in the inset.

    • Western blot - Anti-Raptor antibody [EP539Y] (ab40768)
      Western blot - Anti-Raptor antibody [EP539Y] (ab40768)
      Anti-Raptor antibody [EP539Y] (ab40768) at 1/5000 dilution + Rat brain tissue lysate at 20 mg/ml

      Secondary
      Goat Anti-Rabbit IgG H&L (HRP) (ab97051) at 1/20000 dilution

      Predicted band size: 149 kDa
      Observed band size: 140 kDa why is the actual band size different from the predicted?



      Blocking/Diluting buffer 5% NFDM /TBST
    • Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-Raptor antibody [EP539Y] (ab40768)
      Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-Raptor antibody [EP539Y] (ab40768)
      Immunohistochemical analysis of paraffin-embedded human colon using anti-Raptor ab40768 at 1/50 dilution.
    • Western blot - Anti-Raptor antibody [EP539Y] (ab40768)
      Western blot - Anti-Raptor antibody [EP539Y] (ab40768)
      Anti-Raptor antibody [EP539Y] (ab40768) at 1/1000 dilution + Mouse brain tissue lysate at 10 µg

      Secondary
      Goat Anti-Rabbit IgG H&L (HRP) (ab97051) at 1/20000 dilution

      Predicted band size: 149 kDa
      Observed band size: 140 kDa why is the actual band size different from the predicted?



      Blocking/Diluting buffer 5% NFDM /TBST
    • Western blot - Anti-Raptor antibody [EP539Y] (ab40768)
      Western blot - Anti-Raptor antibody [EP539Y] (ab40768)
      All lanes : Anti-Raptor antibody [EP539Y] (ab40768) at 1/500 dilution

      Lane 1 : 293 cell lysate
      Lane 2 : MCF-7 cell lysate
      Lane 3 : HeLa cell lysate
      Lane 4 : A431 cell lysate

      Lysates/proteins at 10 µg per lane.

      Predicted band size: 149 kDa
      Observed band size: 140 kDa why is the actual band size different from the predicted?
      Additional bands at: 21 kDa, 37 kDa. We are unsure as to the identity of these extra bands.

    • Anti-Raptor antibody [EP539Y] (ab40768)
      Anti-Raptor antibody [EP539Y] (ab40768)

    Protocols

    To our knowledge, customised protocols are not required for this product. Please try the standard protocols listed below and let us know how you get on.

    Click here to view the general protocols

    References (28)

    Publishing research using ab40768? Please let us know so that we can cite the reference in this datasheet.

    ab40768 has been referenced in 28 publications.

    • Song X  et al. Jian Pi Tiao Gan Yin alleviates obesity phenotypes through mTORC1/SREBP1 signaling in vitro and in vivo. Ann Transl Med 10:291 (2022). PubMed: 35433951
    • Wang C  et al. The ubiquitin ligase Nedd4-2 mediates the regulation of PepT2 by mTORC1 in bovine mammary epithelial cells. Anim Nutr 10:12-18 (2022). PubMed: 35601254
    • Ando T  et al. Combined Anticancer Effect of Plasma-Activated Infusion and Salinomycin by Targeting Autophagy and Mitochondrial Morphology. Front Oncol 11:593127 (2021). PubMed: 34150606
    • Zhang YY  et al. Calpain 6 inhibits autophagy in inflammatory environments: A preliminary study on myoblasts and a chronic kidney disease rat model. Int J Mol Med 48:N/A (2021). PubMed: 34435644
    • Lu X  et al. YAP Accelerates Notch-Driven Cholangiocarcinogenesis via mTORC1 in Mice. Am J Pathol 191:1651-1667 (2021). PubMed: 34129844
    View all Publications for this product

    Customer reviews and Q&As

    Show All Reviews Q&A
    Submit a review Submit a question

    1-7 of 7 Abreviews or Q&A

    Western blot abreview for Anti-Raptor antibody [EP539Y]

     Excellent
    Abreviews
    abreview image
    Application
    Western blot
    Sample
    Mouse Tissue lysate - whole (Muscle)
    Loading amount
    50 µg
    Specification
    Muscle
    Gel Running Conditions
    Reduced Denaturing
    Blocking step
    BSA as blocking agent for 1 hour(s) and 0 minute(s) · Concentration: 5% · Temperature: 25°C
    Read More
    The reviewer received a reward from Abcam’s Loyalty Program in thanks for submitting this Abreview and for helping the scientific community make better-informed decisions.

    Abcam user community

    Verified customer

    Submitted May 30 2012

    Western blot abreview for Anti-Raptor antibody [EP539Y]

     Good
    Abreviews
    abreview image
    Application
    Western blot
    Sample
    Mouse Tissue lysate - whole (Liver)
    Loading amount
    40 µg
    Specification
    Liver
    Gel Running Conditions
    Reduced Denaturing
    Blocking step
    BSA as blocking agent for 30 minute(s) · Concentration: 3% · Temperature: 25°C
    Read More
    The reviewer received a reward from Abcam’s Loyalty Program in thanks for submitting this Abreview and for helping the scientific community make better-informed decisions.

    Abcam user community

    Verified customer

    Submitted Mar 09 2012

    Immunoprecipitation abreview for Anti-Raptor antibody [EP539Y]

     Good
    Abreviews
    This product is known to not work in this application or species.
    abreview image
    Application
    Immunoprecipitation
    Sample
    Mouse Tissue lysate - whole (Liver)
    Total protein in input
    3000 µg
    Specification
    Liver
    Immuno-precipitation step
    Protein A
    Read More
    The reviewer received a reward from Abcam’s Loyalty Program in thanks for submitting this Abreview and for helping the scientific community make better-informed decisions.

    Abcam user community

    Verified customer

    Submitted Mar 09 2012

    Question

    Dear ,
    please find attach the filled questionnaire.
    thank you very much

    Read More
    Answer

    Thank you very much for taking the time to complete and send the questionnaire.

    The details you have kindly provided will provide us with vital information for our monitoring of product quality.

    Unfortunately none of these antibodies have been tested in frozen sections, and therefore they are not guaranteed in this application, nor do we have specific protocols tips to provide for it.

    However, there are some suggestions that may optimise the antibody performance, if you wish to give them a try:

    -I would recommend decreasing the permeabilization conditions. Both proteins are cytoplasmic; there is no need for such an aggressive permeabilization step. I would repeat the staining skipping this step and removing Triton from the blocking and the washing buffer as well as from the antibody diluent.

    -Use milder blocking conditions, using 5% of serum if possible from the same species as the secondary was raised in, to avoid cross reactivity.

    -Try using more concentrated dilutions, especially for Raptor antibody (1/50 – 1/25).

    If possible I would recommend trying both antibodies in paraffin embedded sections, as they are both guaranteed for it. The link to our protocols page is below, where you can find all the information regarding the different applications protocols, and very useful information for performing paraffin embedded immunostaining.

    https://www.abcam.com/index.html?pageconfig=popular_protocols

    Should the suggestions not improve the results, please do not hesitate to contact me again and I’ll be more than happy to help you further.

    Read More

    Question

    Dear Sr./ Mrs.,
    I am writing to you to discuss a problem we have just had with two
    antibodies purchased at your company, Rictor (AB104838) and Raptor
    (AB40768).
    We have just performed immunofluorescence staining on mouse cerebral
    tissue The staining did not work out and was similar to a negative
    control (without primary antibody and plus secondary antibody).
    The protocol that we used was the following:
    Sections were washed twice with PBS and incubated with NH4Cl 50 mM
    during 30 min. Tissue was permeated along 20 min by treatment with PBS
    containing 0.5% Triton X-100 and blocked during 1 h at room
    temperature in PBS plus 0.2% bovine serum albumin, 0.2% glycine, 0.2%
    sodium azide, 0.5% Triton X-100 and 5% normal goat serum. Slices were
    incubated overnight at 4°C with the corresponding primary antibodies
    in a buffer containing PBS plus 0.3% Triton X-100, 0.2% bovine serum
    albumin and 0.2% sodium azide. After primary antibody incubation,
    slices were washed in PBS twice for 10 min and incubated 2 h at room
    temperature with subtype-specific fluorescent secondary antibodies.
    After two consecutive 10 min-washes, slices were mounted with Mowiol
    on silane-coated slides.
    Primary antibodies were diluted 1:200 in the case of Rictor, and and
    1:100 in the case of Raptor, as recommended on the data sheet.
    We are not sure if we should follow a specific protocol to get the
    staining work out fine.
    Thank you very much,

    Read More
    Answer

    Thank you for contacting us.

    I am sorry to hear you are experiencing difficulties with one of our products. We take product complaints very seriously, and investigate every product that we feel may not be performing correctly.

    In order to better understand the problem, I would like to send you a questionnaire with some additional questions about the protocol used, as well as the order details. All this information is crucial to help us understand the possible causes of the problem. Any images are highly appreciated.

    Once we receive the completed questionnaire, we will look at the protocol and see if there are any suggestions we can make that may improve the results. If no suggestions can be made to the protocol, and the antibody was purchased within the last 6 months, it is covered by our Abpromise guarantee, and therefore I can offer you a free of charge replacement or a credit note for it.

    I look forward to receiving your reply. Please do not hesitate to contact us if you need any more advice or information.

    Read More

    Question

    I tested this antibody in WB using mouse cardiac tissue and saw no bands.  Can I please get a replacement with a different antibody?

    Read More
    Answer

    Thank you for contacting Abcam regarding ab40768. I am sorry this product did not perform as stated on the datasheet and for the inconvenience this has caused. As requested, I have issued a free of charge replacement. To check the status of the order please contact our Customer Service team and reference this number. Please note that this free of charge replacement vial is also covered by our Abpromise guarantee. Should you still be experiencing difficulties, or if you have any further questions, please do not hesitate to let us know. I wish you the best of luck with your research.  

    Read More

    Question

    While you were in your meeting I spoke to Bola who wanted your best recommendation for what dilution she should use for her secondary, while using ab40768 in Western Blotting, she also wanted your opinion if she should use milk for this. I told her I’ve been told we use BSA in characterization because milk contains other host proteins, but I’m no scientist.        

    Read More
    Answer

    Thank you for contacting us. The secondary antibodies can be used within range from 1/1000 to 1/50,000 dilution. The dilution range depends on the concentration of secondary antibody. We recommend starting with 1/1000, 1/2000, 1/5000 and 1/10,000 for optimization purpose. The dilution factor should also be recommended on vendor's datasheets of secondary antibodies which will give you more idea at what dilution to use. We use BSA in our own lab. You can use milk however if the background will be high then we may recommend switching to BSA. I hope this information is helpful to you. Please do not hesitate to contact us if you need any more advice or information. 

    Read More

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