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Validated using a knockout cell lineRecombinantRabMAb

Recombinant Anti-YTHDF3 antibody [EPR21912-3] (ab220161)

Reviews (3) Submit a question References (9)
Western blot - Anti-YTHDF3 antibody [EPR21912-3] (ab220161)
  • Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-YTHDF3 antibody [EPR21912-3] (ab220161)
  • Immunohistochemistry (Frozen sections) - Anti-YTHDF3 antibody [EPR21912-3] (ab220161)
  • Flow Cytometry (Intracellular) - Anti-YTHDF3 antibody [EPR21912-3] (ab220161)
  • Immunoprecipitation - Anti-YTHDF3 antibody [EPR21912-3] (ab220161)
  • Western blot - Anti-YTHDF3 antibody [EPR21912-3] (ab220161)
  • Western blot - Anti-YTHDF3 antibody [EPR21912-3] (ab220161)
  • Immunohistochemistry (Frozen sections) - Anti-YTHDF3 antibody [EPR21912-3] (ab220161)
  • Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-YTHDF3 antibody [EPR21912-3] (ab220161)
  • Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-YTHDF3 antibody [EPR21912-3] (ab220161)
  • Anti-YTHDF3 antibody [EPR21912-3] (ab220161)

Key features and details

  • Produced recombinantly (animal-free) for high batch-to-batch consistency and long term security of supply
  • Rabbit monoclonal [EPR21912-3] to YTHDF3
  • Suitable for: Flow Cyt (Intra), IP, WB, IHC-Fr, IHC-P
  • Knockout validated
  • Reacts with: Mouse, Rat, Human

Conjugates logo Related conjugates and formulations

Carrier Free

Overview

  • Product name

    Anti-YTHDF3 antibody [EPR21912-3]
    See all YTHDF3 primary antibodies

  • Description

    Rabbit monoclonal [EPR21912-3] to YTHDF3

  • Host species

    Rabbit

  • Tested applications

    Suitable for: Flow Cyt (Intra), IP, WB, IHC-Fr, IHC-Pmore details
    Unsuitable for: ICC/IF

  • Species reactivity

    Reacts with: Mouse, Rat, Human

  • Immunogen

    Recombinant fragment. This information is proprietary to Abcam and/or its suppliers.

  • Positive control

    • WB: Wild-type mESC whole cell lysate. GST-tagged human YTHDF3 recombinant protein. Fresh HT-1080, HeLa, NIH/3T3, PC-12 whole cell lysate. IHC-P: Mouse and rat cerebrum tissue. Human colon tissue. IHC-Fr: Mouse and rat cerebrum tissue. Flow Cyt (intra): HeLa cells. IP: HeLa whole 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.

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 long term. Avoid freeze / thaw cycle.

  • Storage buffer

    pH: 7.2
    Preservative: 0.01% Sodium azide
    Constituents: 40% Glycerol (glycerin, glycerine), 0.05% BSA, PBS
  • Concentration information loading...

  • Purity

    Protein A purified

  • Clonality

    Monoclonal

  • Clone number

    EPR21912-3

  • Isotype

    IgG

  • Research areas

Applications

The Abpromise guarantee

Our Abpromise guarantee covers the use of ab220161 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 (Intra)
1/60.
IP
1/30.
WB (1)
1/1000. Detects a band of approximately 73 kDa (predicted molecular weight: 64 kDa).
IHC-Fr
1/100.
IHC-P (1)
Use at an assay dependent concentration. Perform heat mediated antigen retrieval with Tris/EDTA buffer pH 9.0 before commencing with IHC staining protocol.

Use at 1/500 (mouse, rat) and 1/100 (human) dilution.
Notes
Flow Cyt (Intra)
1/60.
IP
1/30.
WB
1/1000. Detects a band of approximately 73 kDa (predicted molecular weight: 64 kDa).
IHC-Fr
1/100.
IHC-P
Use at an assay dependent concentration. Perform heat mediated antigen retrieval with Tris/EDTA buffer pH 9.0 before commencing with IHC staining protocol.

Use at 1/500 (mouse, rat) and 1/100 (human) dilution.
Application notes
Is unsuitable for ICC/IF.

Target

Images

  • Western blot - Anti-YTHDF3 antibody [EPR21912-3] (ab220161)
    Western blot - Anti-YTHDF3 antibody [EPR21912-3] (ab220161)
    All lanes : Anti-YTHDF3 antibody [EPR21912-3] (ab220161) at 1/1000 dilution

    Lane 1 : Wild-type mESC (mouse embryo stem cell) whole cell lysate
    Lane 2 : YTHDF3 knockout mESC whole cell lysate

    Lysates/proteins at 20 µg per lane.

    Secondary
    All lanes : Goat Anti-Rabbit IgG H&L (HRP) (ab97051) at 1/100000 dilution

    Predicted band size: 64 kDa
    Observed band size: 73 kDa why is the actual band size different from the predicted?


    Exposure time: 59 seconds


    The wild-type and YTHDF3 knockout cell lysates were kindly provided by an anonymous collaborator.

    ab220161 was shown to specifically react with YTHDF3 in wild-type mESC cells as signal was lost in YTHDF3 knockout cells. Wild-type and YTHDF3 knockout samples were subjected to SDS-PAGE. ab220161 and ab181602 (Rabbit anti-GAPDH loading control) were incubated 1 hour at room temperature at 1/1000 dilution and 1/200,000 dilution respectively. Blots were developed with Goat Anti-Rabbit IgG, (H+L), Peroxidase conjugated (ab97051) secondary antibody at 1/100,000 dilution for 1 hour at room temperature before imaging.

    The blot was developed on a BIO-RAD® ChemiDoc™ MP instrument using the ECL technique.

    Blocking/Dilution buffer: 5% NFDM/TBST.

  • Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-YTHDF3 antibody [EPR21912-3] (ab220161)
    Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-YTHDF3 antibody [EPR21912-3] (ab220161)

    Immunohistochemical analysis of paraffin-embedded human colon tissue labeling YTHDF3 with ab220161 at 1/100 dilution, followed by a ready to use Goat Anti-Rabbit IgG H&L (HRP). Cytoplasmic staining on human colon (PMID:29103884, 28250115) is observed. Counterstained with hematoxylin.

    Secondary antibody only control: Used PBS instead of primary antibody, secondary antibody is a ready to use Goat Anti-Rabbit IgG H&L (HRP).

    Perform heat mediated antigen retrieval using ab93684 (Tris/EDTA buffer, pH 9.0).

  • Immunohistochemistry (Frozen sections) - Anti-YTHDF3 antibody [EPR21912-3] (ab220161)
    Immunohistochemistry (Frozen sections) - Anti-YTHDF3 antibody [EPR21912-3] (ab220161)

    Immunohistochemical analysis of 4% PFA-fixed, 0.2% Triton X-100 permeabilized frozen mouse cerebrum tissue labeling YTHDF3 with ab220161 at 1/100 dilution (green), followed by ab150077 AlexaFluor®488 Goat anti-Rabbit secondary at a 1/1,000 dilution. Cytoplasmic and nuclear staining in mouse cerebrum (PMID:29103884, 28250115) is observed. Counterstained with DAPI (blue).

    Secondary antibody only control: Used PBS instead of primary antibody, secondary antibody is ab150077 AlexaFluor®488 Goat anti-Rabbit used at a 1/1,000 dilution.

    Heat mediated antigen retrieval using sodium citrate buffer (10 mM citrate pH 6.0 and 0.05% Tween-20).

  • Flow Cytometry (Intracellular) - Anti-YTHDF3 antibody [EPR21912-3] (ab220161)
    Flow Cytometry (Intracellular) - Anti-YTHDF3 antibody [EPR21912-3] (ab220161)

    Intracellular flow cytometric analysis of 4% paraformaldehyde-fixed, 90% methanol permeabilized HeLa (human cervix adenocarcinoma epithelial cell) cell line labeling YTHDF3 with ab220161 at 1/60 (red) compared with a Rabbit monoclonal IgG (ab172730) (black) and an unlabeled control (cells without incubation with primary antibody and secondary antibody) (blue). Goat anti rabbit IgG (Alexa Fluor® 488, ab150077), at 1/2000 dilution was used as the secondary antibody. 

     

     

  • Immunoprecipitation - Anti-YTHDF3 antibody [EPR21912-3] (ab220161)
    Immunoprecipitation - Anti-YTHDF3 antibody [EPR21912-3] (ab220161)

    YTHDF3 was immunoprecipitated from 0.35 mg HeLa (Human cervix adenocarcinoma epithelial cell) whole cell lysate with ab220161 at 1/30 dilution. Western blot was performed from the immunoprecipitate using ab220161 at 1/1000 dilution. VeriBlot for IP Detection Reagent (HRP) (ab131366), was used at 1/5000 dilution.

    Lane 1: HeLa whole cell lysate 10 µg (Input).
    Lane 2: ab220161 IP in HeLa whole cell lysate.
    Lane 3: Rabbit monoclonal IgG (ab172730) instead of ab220161 in HeLa whole cell lysate.

    Blocking/Dilution buffer: 5% NFDM/TBST.
    Exposure time: 15 seconds.

    Lysate were made freshly and used in IP test immediately to minimize protein degradation. Incubation time was 2h.

  • Western blot - Anti-YTHDF3 antibody [EPR21912-3] (ab220161)
    Western blot - Anti-YTHDF3 antibody [EPR21912-3] (ab220161)
    All lanes : Anti-YTHDF3 antibody [EPR21912-3] (ab220161) at 1/1000 dilution

    Lane 1 : HT-1080 (human fibrosarcoma epithelial cell) whole cell lysate (fresh lysate)
    Lane 2 : HeLa (human cervix adenocarcinoma epithelial cell) whole cell lysate (fresh lysate)
    Lane 3 : NIH/3T3 (mouse embryonic fibroblast) whole cell lysate (fresh lysate)
    Lane 4 : PC-12 (rat adrenal gland pheochromocytoma) whole cell lysate (fresh lysate)
    Lane 5 : HeLa (human cervix adenocarcinoma epithelial cell) whole cell lysate
    Lane 6 : NIH/3T3 (mouse embryonic fibroblast) whole cell lysate
    Lane 7 : PC-12 (rat adrenal gland pheochromocytoma) whole cell lysate

    Lysates/proteins at 20 µg per lane.

    Secondary
    All lanes : Goat Anti-Rabbit IgG H&L (HRP) (ab97051) at 1/100000 dilution

    Predicted band size: 64 kDa



    Lysate should be made freshly and used in WB immediately to minimize protein degradation (lane1-4). Lane 5-7 are the lysates from same cell lines but have experienced freeze-thaw cycles.

    Blocking/Dilution buffer: 5% NFDM/TBST.

    Exposure times: Lanes 1-4: 37 secs; Lanes 5-7: 92 secs.

  • Western blot - Anti-YTHDF3 antibody [EPR21912-3] (ab220161)
    Western blot - Anti-YTHDF3 antibody [EPR21912-3] (ab220161)
    All lanes : Anti-YTHDF3 antibody [EPR21912-3] (ab220161) at 1/5000 dilution

    Lane 1 : GST-tagged human YTHDF1 recombinant protein 20 ng
    Lane 2 : GST-tagged human YTHDF2 recombinant protein 20 ng
    Lane 3 : GST-tagged human YTHDF3 recombinant protein 20 ng

    Secondary
    All lanes : Goat Anti-Rabbit IgG H&L (HRP) (ab97051) at 1/100000 dilution

    Predicted band size: 64 kDa


    Exposure time: 10 seconds


    The YTHDF recombinant proteins were kindly provided by an anonymous collaborator.

    Blocking/Dilution buffer: 5% NFDM/TBST.

  • Immunohistochemistry (Frozen sections) - Anti-YTHDF3 antibody [EPR21912-3] (ab220161)
    Immunohistochemistry (Frozen sections) - Anti-YTHDF3 antibody [EPR21912-3] (ab220161)

    Immunohistochemical analysis of 4% PFA-fixed, 0.2% Triton X-100 permeabilized frozen rat cerebrum tissue labeling YTHDF3 with ab220161 at 1/100 dilution (green), followed by ab150077 AlexaFluor®488 Goat anti-Rabbit secondary at a 1/1,000 dilution. Cytoplasmic and nuclear staining in rat cerebrum (PMID:29103884, 28250115) is observed. Counterstained with DAPI (blue).

    Secondary antibody only control: Used PBS instead of primary antibody, secondary antibody is ab150077 AlexaFluor®488 Goat anti-Rabbit used at a 1/1,000 dilution.

    Heat mediated antigen retrieval using sodium citrate buffer (10 mM citrate pH 6.0 and 0.05% Tween-20).

  • Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-YTHDF3 antibody [EPR21912-3] (ab220161)
    Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-YTHDF3 antibody [EPR21912-3] (ab220161)

    Immunohistochemical analysis of paraffin-embedded rat cerebrum tissue labeling YTHDF3 with ab220161 at 1/500 dilution, followed by a ready to use Goat Anti-Rabbit IgG H&L (HRP). Cytoplasmic staining on neurons of rat cerebrum (PMID:29103884, 28250115) is observed. Counterstained with hematoxylin.

    Secondary antibody only control: Used PBS instead of primary antibody, secondary antibody is a ready to use Goat Anti-Rabbit IgG H&L (HRP).

    Perform heat mediated antigen retrieval using ab93684 (Tris/EDTA buffer, pH 9.0).

  • Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-YTHDF3 antibody [EPR21912-3] (ab220161)
    Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-YTHDF3 antibody [EPR21912-3] (ab220161)

    Immunohistochemical analysis of paraffin-embedded mouse cerebrum tissue labeling YTHDF3 with ab220161 at 1/500 dilution, followed by a ready to use Goat Anti-Rabbit IgG H&L (HRP). Cytoplasmic staining on neurons of mouse cerebrum (PMID:29103884, 28250115) is observed. Counterstained with hematoxylin.

    Secondary antibody only control: Used PBS instead of primary antibody, secondary antibody is a ready to use Goat Anti-Rabbit IgG H&L (HRP).

    Perform heat mediated antigen retrieval using ab93684 (Tris/EDTA buffer, pH 9.0).

  • Anti-YTHDF3 antibody [EPR21912-3] (ab220161)
    Anti-YTHDF3 antibody [EPR21912-3] (ab220161)

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 (9)

Publishing research using ab220161? Please let us know so that we can cite the reference in this datasheet.

ab220161 has been referenced in 9 publications.

  • Sun Z  et al. RNA demethylase ALKBH5 inhibits TGF-ß-induced EMT by regulating TGF-ß/SMAD signaling in non-small cell lung cancer. FASEB J 36:e22283 (2022). WB ; Human . PubMed: 35344216
  • You A  et al. TTC22 promotes m6A-mediated WTAP expression and colon cancer metastasis in an RPL4 binding-dependent pattern. Oncogene N/A:N/A (2022). WB ; Human . PubMed: 35798874
  • Tao L  et al. FTO modifies the m6A level of MALAT and promotes bladder cancer progression. Clin Transl Med 11:e310 (2021). PubMed: 33634966
  • Xu H  et al. YTHDF2 alleviates cardiac hypertrophy via regulating Myh7 mRNA decoy. Cell Biosci 11:132 (2021). PubMed: 34266473
  • Ding H  et al. GNAS promotes inflammation-related hepatocellular carcinoma progression by promoting STAT3 activation. Cell Mol Biol Lett 25:8 (2020). WB ; Human . PubMed: 32123532
  • Jurczyszak D  et al. HIV protease cleaves the antiviral m6A reader protein YTHDF3 in the viral particle. PLoS Pathog 16:e1008305 (2020). WB ; Human . PubMed: 32053707
  • Jin D  et al. m6A demethylase ALKBH5 inhibits tumor growth and metastasis by reducing YTHDFs-mediated YAP expression and inhibiting miR-107/LATS2-mediated YAP activity in NSCLC. Mol Cancer 19:40 (2020). PubMed: 32106857
  • Jian D  et al. METTL14 aggravates endothelial inflammation and atherosclerosis by increasing FOXO1 N6-methyladeosine modifications. Theranostics 10:8939-8956 (2020). PubMed: 32802173
  • Jin D  et al. m6A mRNA methylation initiated by METTL3 directly promotes YAP translation and increases YAP activity by regulating the MALAT1-miR-1914-3p-YAP axis to induce NSCLC drug resistance and metastasis. J Hematol Oncol 12:135 (2019). WB ; Human . PubMed: 31818312

Customer reviews and Q&As

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1-3 of 3 Abreviews or Q&A

ChIP abreview for Anti-YTHDF3 antibody [EPR21912-3]

 Inconclusive
Abreviews
Application
ChIP
Sample
Human Cell lysate - whole cell (Lung)
Specification
Lung
Detection step
Other
Type
Cross-linking (X-ChIP)
Duration of cross-linking step: 10 minute(s) and 0 second(s)
Specification of the cross-linking agent: 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 May 07 2021

Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) abreview for Anti-YTHDF3 antibody [EPR21912-3]

 Excellent
Abreviews
abreview image
Application
Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections)
Sample
Mouse Tissue sections (ovary)
Antigen retrieval step
Heat mediated - Buffer/Enzyme Used: Citrate antigen repair solution
Permeabilization
Yes - PBST (0.5% Triton X-100 in PBS)
Specification
ovary
Blocking step
BSA as blocking agent for 1 hour(s) and 30 minute(s) · Concentration: 5% · Temperature: 25°C
Fixative
Paraformaldehyde
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. Bin Shen

Verified customer

Submitted Nov 27 2019

Western blot abreview for Anti-YTHDF3 antibody [EPR21912-3]

 Excellent
Abreviews
abreview image
Application
Western blot
Sample
Mouse Tissue lysate - whole (spleen)
Gel Running Conditions
Reduced Denaturing (10%)
Loading amount
30 µg
Specification
spleen
Blocking step
Milk as blocking agent for 1 hour(s) and 30 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.

PROF. Bin Shen

Verified customer

Submitted Nov 27 2019

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