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RecombinantRabMAb

Recombinant Anti-mTOR antibody [Y391] (ab32028)

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Reviews (4)Q&A (5)References (150)

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Western blot - Anti-mTOR antibody [Y391] (ab32028)
  • Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-mTOR antibody [Y391] (ab32028)
  • Immunohistochemistry (Frozen sections) - Anti-mTOR antibody [Y391] (ab32028)
  • Western blot - Anti-mTOR antibody [Y391] (ab32028)
  • Immunoprecipitation - Anti-mTOR antibody [Y391] (ab32028)
  • Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-mTOR antibody [Y391] (ab32028)
  • Immunoprecipitation - Anti-mTOR antibody [Y391] (ab32028)
  • Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-mTOR antibody [Y391] (ab32028)
  • Western blot - Anti-mTOR antibody [Y391] (ab32028)
  • Anti-mTOR antibody [Y391] (ab32028)

Key features and details

  • Produced recombinantly (animal-free) for high batch-to-batch consistency and long term security of supply
  • Rabbit monoclonal [Y391] to mTOR
  • Suitable for: WB, IP, IHC-P, IHC-Fr
  • Reacts with: Mouse, Rat, Human

Conjugates logo Related conjugates and formulations

Carrier Free

You may also be interested in

Protein
Product image
Recombinant Human mTOR protein (ab114179)
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Mouse mTOR ELISA Kit (ab206311)
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Human MTOR knockout HEK-293T cell line (ab255411)

View more associated products

Overview

  • Product name

    Anti-mTOR antibody [Y391]
    See all mTOR primary antibodies
  • Description

    Rabbit monoclonal [Y391] to mTOR
  • Host species

    Rabbit
  • Specificity

    Expression levels of the target protein vary with sample type and some optimisation may be required.

  • Tested applications

    Suitable for: WB, IP, IHC-P, IHC-Frmore details
    Unsuitable for: Flow Cyt or ICC/IF
  • Species reactivity

    Reacts with: Mouse, Rat, Human
  • Immunogen

    Synthetic peptide within Human mTOR aa 2400-2500 (C terminal). The exact sequence is proprietary.
    (Peptide available as ab193663)

  • Epitope

    ab32028 reacts with an epitope located in the C terminal region of mTOR.
  • Positive control

    • WB: Jurkat whole cell lysate (ab30128), HeLa, HaCaT and MDA-MB-231 cell lysates and rat brain tissue lysate. IHC-P: Human breast carcinoma, mouse testis and rat testis tissues. IP: Rat brain tissue lysate and HeLa whole cell lysate (ab150035). IHC-Fr: Human heart tissue sections.
  • 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

  • Form

    Liquid
  • Storage instructions

    Shipped at 4°C. Store at +4°C short term (1-2 weeks). Upon delivery aliquot. Store at -20°C. Avoid freeze / thaw cycle.
  • Storage buffer

    pH: 7.20
    Preservative: 0.01% Sodium azide
    Constituents: 59% PBS, 40% Glycerol, 0.05% BSA
  • Concentration information loading...
  • Purity

    Protein A purified
  • Clonality

    Monoclonal
  • Clone number

    Y391
  • Isotype

    IgG
  • Research areas

    • Cell Biology
    • Cell Cycle
    • Cell Cycle Inhibitors
    • Other
    • Epigenetics and Nuclear Signaling
    • DNA / RNA
    • DNA Damage & Repair
    • DNA Damage Response
    • DNA Damage Recognition
    • Cancer
    • Cell cycle
    • Cell cycle inhibitors
    • Other
    • Cardiovascular
    • Heart
    • Cardiogenesis
    • Transcription factors/regulators
    • Kits/ Lysates/ Other
    • Kits
    • ELISA Kits
    • ELISA Kits
    • Growth factors and hormones ELISA kits
    • Metabolism
    • Types of disease
    • Obesity
    • Neuroscience
    • Development
    • Neuroscience
    • Processes

Associated products

  • Alternative Versions

    • Anti-mTOR antibody [Y391] - BSA and Azide free (ab218525)
  • ChIP Related Products

    • ChIP Kit (ab500)
  • Compatible Secondaries

    • Goat Anti-Rabbit IgG H&L (Alexa Fluor® 488) (ab150077)
    • Goat Anti-Rabbit IgG H&L (DyLight® 488) preadsorbed (ab96899)
    • Goat Anti-Rabbit IgG H&L (HRP) (ab97051)
  • Isotype control

    • Rabbit IgG, monoclonal [EPR25A] - Isotype Control (ab172730)
  • Positive Controls

    • HeLa whole cell lysate (ab150035)
    • MCF7 whole cell lysate (ab29537)
    • HeLa whole cell lysate (ab29545)
    • Jurkat whole cell lysate (ab30128)
    • RAW 264.7 whole cell lysate (ab7187)
    • Jurkat whole cell lysate (ab7899)
  • Recombinant Protein

    • Recombinant Human mTOR protein (ab114179)

Applications

The Abpromise guarantee

Our Abpromise guarantee covers the use of ab32028 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 (3)
1/1000 - 1/5000. Detects a band of approximately 250 kDa (predicted molecular weight: 289 kDa).

This antibody detects non-specific bands and high background. It doesn't detect the target band in some mouse and rat tissues.

IP
1/50 - 1/100.
IHC-P (1)
1/400. Perform heat mediated antigen retrieval before commencing with IHC staining protocol.

See IHC antigen retrieval protocols.

IHC-Fr
1/200.
Notes
WB
1/1000 - 1/5000. Detects a band of approximately 250 kDa (predicted molecular weight: 289 kDa).

This antibody detects non-specific bands and high background. It doesn't detect the target band in some mouse and rat tissues.

IP
1/50 - 1/100.
IHC-P
1/400. Perform heat mediated antigen retrieval before commencing with IHC staining protocol.

See IHC antigen retrieval protocols.

IHC-Fr
1/200.
Application notes
Is unsuitable for Flow Cyt or ICC/IF.

Target

  • 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.
  • Target information above from: UniProt accession P42345 The UniProt Consortium
    The Universal Protein Resource (UniProt) in 2010
    Nucleic Acids Res. 38:D142-D148 (2010) .

    Information by UniProt
  • Database links

    • Entrez Gene: 2475 Human
    • Entrez Gene: 56717 Mouse
    • Entrez Gene: 56718 Rat
    • Omim: 601231 Human
    • SwissProt: P42345 Human
    • SwissProt: Q9JLN9 Mouse
    • SwissProt: P42346 Rat
    • Unigene: 338207 Human
    • Unigene: 21158 Mouse
    • Unigene: 11008 Rat
    see all
  • Alternative names

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

Images

  • Western blot - Anti-mTOR antibody [Y391] (ab32028)
    Western blot - Anti-mTOR antibody [Y391] (ab32028)
    All lanes : Anti-mTOR antibody [Y391] (ab32028) at 1/2000 dilution

    Lane 1 : MCF7 (Human breast adenocarcinoma epithelial cell) whole cell lysates in RIPA buffer
    Lane 2 : MCF7 (Human breast adenocarcinoma epithelial cell) whole cell lysates in HOT buffer
    Lane 3 : Rat brain lysates in RIPA buffer
    Lane 4 : Rat brain lysates in HOT buffer
    Lane 5 : HepG2 (Human hepatocellular carcinoma epithelial cell) whole cell lysates
    Lane 6 : Human fetal lung lysates

    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: 289 kDa
    Observed band size: 290 kDa why is the actual band size different from the predicted?


    Exposure time: 15 seconds


    Blocking and diluting buffer: 5% NFDM/TBST

  • Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-mTOR antibody [Y391] (ab32028)
    Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-mTOR antibody [Y391] (ab32028)

    Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) analysis of human breast carcinoma tissue labelling mTOR with purified ab32028 at a dilution of 1/400. Heat mediated antigen retrieval was performed using EDTA buffer pH 9. ab97051, a HRP-conjugated goat anti-rabbit IgG (H+L) was used as the secondary antibody (1/500). Negative control using PBS instead of primary antibody. Counterstained with hematoxylin.

  • Immunohistochemistry (Frozen sections) - Anti-mTOR antibody [Y391] (ab32028)
    Immunohistochemistry (Frozen sections) - Anti-mTOR antibody [Y391] (ab32028)

    IHC image of mTOR staining in a section of frozen normal human heart performed on a Leica BONDTM system using the standard protocol. The section was fixed in 10% paraformaldehyde (10 min) prior to staining. The section was incubated with ab32028, 1/200 dilution, for 15 mins at room temperature and detected using an HRP conjugated compact polymer system. DAB was used as the chromogen. The section was then counterstained with haematoxylin and mounted with DPX. The inset secondary-only control image is taken from an identical assay without primary antibody.

    For other IHC staining systems (automated and non-automated) customers should optimize variable parameters such as antigen retrieval conditions, primary antibody concentration and antibody incubation times.

  • Western blot - Anti-mTOR antibody [Y391] (ab32028)
    Western blot - Anti-mTOR antibody [Y391] (ab32028)
    Lanes 1-7 : Anti-mTOR antibody [Y391] (ab32028)
    Lane 8 : Anti-mTOR antibody [Y391] (ab32028) at 1/1000 dilution

    Lane 1 : Rat brain tissue lysate with 5% NFDM/TBST
    Lane 2 : Rat heart tissue lysate with 5% NFDM/TBST
    Lane 3 : Rat liver tissue lysate with 5% NFDM/TBST
    Lane 4 : Rat spleen tissue with 5% NFDM/TBST
    Lane 5 : Mouse brain tissue lysate with 5% NFDM/TBST
    Lane 6 : Mouse heart tissue lysate with 5% NFDM/TBST
    Lane 7 : Mouse kidney tissue lysate with 5% NFDM/TBST
    Lane 8 : Mouse liver tissue lysate with 5% NFDM /TBST

    Lysates/proteins at 20 µg per lane.

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

    Predicted band size: 289 kDa
    Observed band size: 289 kDa



    Exposure time:

    Lane 1: 10 seconds
    Lane 2-8: 180 seconds

    This antibody detects non-specific bands and high background. It doesn’t detect the target band in some mouse and rat tissues.

  • Immunoprecipitation - Anti-mTOR antibody [Y391] (ab32028)
    Immunoprecipitation - Anti-mTOR antibody [Y391] (ab32028)

    ab32028 (purified) at a dilution of 1/100 immunoprecipitating mTOR in HeLa whole cell lysate.

    Lane 1 (input): HeLa whole cell lysate (10µg)

    Lane 2 (+): ab32028 + HeLa whole cell lysate.

    Lane 3 (-): Rabbit monoclonal IgG (ab172730) instead of ab32028 in HeLa whole cell lysate.

    For western blotting, ab131366 VeriBlot for IP Detection Reagent (HRP) was used for detection (1/1000).

    Blocking buffer and concentration: 5% NFDM/TBST.

    Diluting buffer and concentration: 5% NFDM /TBST.

  • Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-mTOR antibody [Y391] (ab32028)
    Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-mTOR antibody [Y391] (ab32028)

    Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) analysis of mouse testis tissue labelling mTOR with purified ab32028 at a dilution of 1/400. Heat mediated antigen retrieval was performed using EDTA buffer pH 9. ab97051, a HRP-conjugated goat anti-rabbit IgG (H+L) was used as the secondary antibody (1/500). Negative control using PBS instead of primary antibody. Counterstained with hematoxylin.

  • Immunoprecipitation - Anti-mTOR antibody [Y391] (ab32028)
    Immunoprecipitation - Anti-mTOR antibody [Y391] (ab32028)

    ab32028 (purified) at a dilution of 1/100 immunoprecipitating mTOR in rat brain tissue lysate.

    Lane 1 (input): Rat brain tissue lysate (10µg)

    Lane 2 (+): ab32028 + rat brain tissue lysate.

    Lane 3 (-): Rabbit monoclonal IgG (ab172730) instead of ab32028 in rat brain tissue lysate.

    For western blotting, ab131366 VeriBlot for IP Detection Reagent (HRP) was used for detection (1/1000).

    Blocking buffer and concentration: 5% NFDM/TBST.

    Diluting buffer and concentration: 5% NFDM /TBST.

  • Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-mTOR antibody [Y391] (ab32028)
    Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-mTOR antibody [Y391] (ab32028)

    Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) analysis of rat testis tissue labelling mTOR with purified ab32028 at a dilution of 1/400. Heat mediated antigen retrieval was performed using EDTA buffer pH 9. ab97051, a HRP-conjugated goat anti-rabbit IgG (H+L) was used as the secondary antibody (1/500). Negative control using PBS instead of primary antibody. Counterstained with hematoxylin.

  • Western blot - Anti-mTOR antibody [Y391] (ab32028)
    Western blot - Anti-mTOR antibody [Y391] (ab32028)
    All lanes : Anti-mTOR antibody [Y391] (ab32028) at 1/1000 dilution

    Lane 1 : HeLa (Human cervix adenocarcinoma epithelial cell) whole cell lysate with 5% NFDM/TBST
    Lane 2 : HaCaT (Human skin keratinocyte) whole cell lysate with 5% NFDM/TBST
    Lane 3 : MDA-MB-231 (Human breast adenocarcinoma epithelial cell) whole cell lysate with 5% NFDM/TBST
    Lane 4 : Mouse thymus (8-10 weeks) tissue lysate with 5% NFDM/TBST
    Lane 5 : Mouse lung (8-10 weeks) tissue lysate with 5% NFDM/TBST
    Lane 6 : Rat thymus tissue lysate with 5% NFDM/TBST
    Lane 7 : Rat lung tissue lysate with 5% NFDM/TBST

    Lysates/proteins at 20 µg per lane.

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

    Predicted band size: 289 kDa
    Observed band size: 289 kDa



    Exposure time:

    Lane 1-3: 100 seconds
    Lane 4-7: 180 seconds

    This antibody detects non-specific bands and high background. It doesn’t detect the target band in some mouse and rat tissues.

  • Anti-mTOR antibody [Y391] (ab32028)
    Anti-mTOR antibody [Y391] (ab32028)

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

Datasheets and documents

  • SDS download

  • Datasheet download

    Download

References (150)

Publishing research using ab32028? Please let us know so that we can cite the reference in this datasheet.

ab32028 has been referenced in 150 publications.

  • Qiu X  et al. Metformin alleviates ß-glycerophosphate-induced calcification of vascular smooth muscle cells via AMPK/mTOR-activated autophagy. Exp Ther Med 21:58 (2021). PubMed: 33365058
  • Gong L  et al. The co-treatment of rosuvastatin with dapagliflozin synergistically inhibited apoptosis via activating the PI3K/AKt/mTOR signaling pathway in myocardial ischemia/reperfusion injury rats. Open Med (Wars) 15:47-57 (2021). PubMed: 33385063
  • Rong JH  et al. Lobaplatin Enhances Radioactive 125I Seed-Induced Apoptosis and Anti-Proliferative Effect in Non-Small Cell Lung Cancer by Suppressing the AKT/mTOR Pathway. Onco Targets Ther 14:289-300 (2021). PubMed: 33469307
  • Zhang S  et al. Metformin-Induced MicroRNA-34a-3p Downregulation Alleviates Senescence in Human Dental Pulp Stem Cells by Targeting CAB39 through the AMPK/mTOR Signaling Pathway. Stem Cells Int 2021:6616240 (2021). PubMed: 33505470
  • Zeng X  et al. Inhibition of mitochondrial and cytosolic calpain attenuates atrophy in myotubes co-cultured with colon carcinoma cells. Oncol Lett 21:124 (2021). PubMed: 33552245
View all Publications for this product

Customer reviews and Q&As

Show All Reviews Q&A
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1-9 of 9 Abreviews or Q&A

Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) abreview for Anti-mTOR antibody [Y391]

Good
Abreviews
Abreviews
abreview image
Application
Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections)
Sample
Human Tissue sections (prostate cancer)
Antigen retrieval step
Heat mediated - Buffer/Enzyme Used: EDTA, pH9, 100C 20 min
Permeabilization
No
Specification
prostate cancer
Blocking step
H2O2 as blocking agent for 10 minute(s) · Concentration: 3% · Temperature: 25°C
Fixative
Formaldehyde
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 Apr 24 2015

Western blot abreview for Anti-mTOR antibody [Y391]

Excellent
Abreviews
Abreviews
Application
Western blot
Sample
Human Cell lysate - whole cell (HUVEC)
Gel Running Conditions
Reduced Denaturing (gradient 4-15%)
Loading amount
45 µg
Treatment
FGF2 50 ng/ml for 5 minutes
Specification
HUVEC
Blocking step
Milk as blocking agent for 1 hour(s) and 0 minute(s) · Concentration: 5% · Temperature: RT°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 Sep 17 2008

Western blot abreview for Anti-mTOR antibody [Y391] - ChIP Grade

Poor
Abreviews
Abreviews
Application
Western blot
Sample
Mouse Tissue lysate - whole (ventricular tissue lysates)
Loading amount
100 µg
Specification
ventricular tissue lysates
Gel Running Conditions
Reduced Denaturing (4 ~ 12% gradient NuPAGE)
Blocking step
Milk as blocking agent for 1 hour(s) and 0 minute(s) · Concentration: 1% · Temperature: 22°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 Jul 05 2008

Western blot abreview for Anti-mTOR antibody [Y391] - ChIP Grade

Average
Abreviews
Abreviews
abreview image
Application
Western blot
Sample
Human Cell lysate - whole cell (Keratinocyte)
Loading amount
15 µg
Specification
Keratinocyte
Gel Running Conditions
Reduced Denaturing (10% gel)
Blocking step
BSA as blocking agent for 1 hour(s) and 0 minute(s) · Concentration: 10% · 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.

Prof. Paul Townsend

Verified customer

Submitted Jun 26 2008

Question

I am attempting to immunoprecipitate mTOR and known interacting proteins (specifically Raptor and Rictor) with ab32028. I am successfully pulling down mTOR (as evidenced by Western blotting), but it is very inefficient and I am not seeing interacting proteins. Could I receive some advice/assistance on immunoprecipitation with this antibody? Please let me know what other info you would like from me about my protocol.

Read More

Abcam community

Verified customer

Asked on Aug 09 2012

Answer

Thank you for your reply.

As requested, I am sending you a new vial of mTOR antibody, ab2732 to use in your IP experiments, since the original vial of antibody was not working as well as it should have. Your new order number, for tracking purposes only, is *.

If there is anything else I can help you with, or if you have any issues with the new vial of the antibody, then please let me know.

Read More

Abcam Scientific Support

Answered on Aug 09 2012

Question

I am attempting to immunoprecipitate mTOR and known interacting proteins (specifically Raptor and Rictor) with ab32028. I am successfully pulling down mTOR (as evidenced by Western blotting), but it is very inefficient and I am not seeing interacting proteins. Could I receive some advice/assistance on immunoprecipitation with this antibody? Please let me know what other info you would like from me about my protocol.

Read More

Abcam community

Verified customer

Asked on Aug 08 2012

Answer

Thank you for your patience whilst I was looking in to this matter.

I checked the concentration for this lot and it is around 1.758 mg/ml. So, I don't believe it is a concentration issue.

However, I did notice that IP testing was only done on Human sample here and not mouse sample. Mouse cross-reactivity was only determined by WB only. Therefore, it could be that, although the antibody is IP'ing the mouse mTOR, the affinity of the antibody to the mouse version may be lower than that to human and that might explain why you are not seeing the co-IP of your other proteins of interest.
To help try and resolve this situation, I don’t think that any protocol tips would prove to be successful; therefore I would like to suggest that I send ab2732 as a free of charge replacement:

https://www.abcam.com/mTOR-antibody-ab2732.html



If you are interested in receiving ab2732, then please let me know, or if there is anything else that I can help you with to resolve this issue. If you could also provide either the Abcam order number or the PO# used to purchase the antibody, that would be very helpful.



I look forward to your reply.

Read More

Abcam Scientific Support

Answered on Aug 08 2012

Question

I am attempting to immunoprecipitate mTOR and known interacting proteins (specifically Raptor and Rictor) with ab32028. I am successfully pulling down mTOR (as evidenced by Western blotting), but it is very inefficient and I am not seeing interacting proteins. Could I receive some advice/assistance on immunoprecipitation with this antibody? Please let me know what other info you would like from me about my protocol.

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Abcam community

Verified customer

Asked on Aug 06 2012

Answer

Thank you for contacting Abcam.

I am sorry that you are having issues with ab32028 in IP. The antibody is covered under our Abpromise for six months and is guaranteed to work in IP on mouse, rat and human samples . If we cannot resolve the issue you are having with the antibody then I would be happy to either send a replacement antibody or to process a refund.

To be able to help you futher, would you be able to send me some information on the protocol that you have been using with ab32028 eg how much antibody you are using, length of incubation, type of samples etc. Also, if possible, it would be very helpful if you could also supply an image of the results that you are getting with the antibody.

I look forward to your reply and helping to resolve this issue.

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Abcam Scientific Support

Answered on Aug 06 2012

Question

Thanks again for getting back to me. Are these other mTOR antibodies
tested to work in rat by abcam? If so can you please send me a few of
their catalog numbers so I can look more into it? Sorry to be picky
about this but we've already tested a bunch of slides already
on this current antibody so I need to be as efficient as possible with
any other ones we test.

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Abcam community

Verified customer

Asked on May 12 2012

Answer

Thank you for your email.
I have heard back regarding ab32028:
The lab tested WB on C6 and PC-12 cell lysates as well as on rat brain tissue lysates. They did not test rat tissues in IHC-P.
As soon as I have more information for the other antibodies, I will let you know.
I hope this information is of help to you. Please do not hesitate to contact us if you need any more advice or information.
Use our products? Submit an Abreview. Earn rewards!
https://www.abcam.com/abreviews

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Abcam Scientific Support

Answered on May 12 2012

Question

Thanks again for getting back to me. Are these other mTOR antibodies
tested to work in rat by abcam? If so can you please send me a few of
their catalog numbers so I can look more into it? Sorry to be picky
about this but we've already tested a bunch of slides already
on this current antibody so I need to be as efficient as possible with
any other ones we test.

Read More

Abcam community

Verified customer

Asked on May 11 2012

Answer

Thank you for your reply.
The following mTOR antibodies have been tested in rat as well as IHC-P:
ab32028, ab51089, ab59299, and ab59350.
The antibody ab32028 has been tested in the lab on IHC-P. I am not certain that rat tissue was used, though. A publication used rat samples: Gant JC et al. Disrupting function of FK506-binding protein 1b/12.6 induces the Ca²+-dysregulation aging phenotype in hippocampal neurons. J Neurosci 31:1693-703 (2011). Rat. PubMed: 21289178
ab51089, ab59299, ab59350: IHC-P was tested, but I have no rat data at hand.
I will contact the lab to check what rat tissue/samples the antibodies were tested on and let you know.
I hope this information is helpful to you. Please do not hesitate to contact us if you need any more advice or information.
Use our products? Submit an Abreview. Earn rewards!
https://www.abcam.com/abreviews

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Abcam Scientific Support

Answered on May 11 2012

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