Astana Biomed Group, an authorized Abcam distributor in Central Asia

Anti-WTAP antibody [EPR18744] (ab195380)

Key features and details

  • Produced recombinantly (animal-free) for high batch-to-batch consistency and long term security of supply
  • Rabbit monoclonal [EPR18744] to WTAP
  • Suitable for: ICC/IF, IP, Flow Cyt, IHC-P, WB
  • Knockout validated
  • Reacts with: Human

Overview

  • Product name

    Anti-WTAP antibody [EPR18744]
    See all WTAP primary antibodies

  • Description

    Rabbit monoclonal [EPR18744] to WTAP

  • Host species

    Rabbit

  • Tested Applications & Species

    Application Species
    Flow Cyt
    Human
    ICC/IF
    Human
    IHC-P
    Human
    IP
    Human
    WB
    Human
    See all applications and species data

  • Immunogen

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

  • Positive control

    • WB: Jurkat, K562, HeLa and HepG2 whole cell lysates; human kidney, thymus and lung lysates. IHC-P: Human endometrium and liver cancer tissues. ICC/IF: Jurkat and HeLa cells. Flow Cyt: Jurkat cells. IP: Jurkat 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: PBS, 40% Glycerol (glycerin, glycerine), 0.05% BSA
  • Concentration information loading...

  • Purity

    Protein A purified

  • Clonality

    Monoclonal

  • Clone number

    EPR18744

  • Isotype

    IgG

  • Research areas

Images

  • Western blot - Anti-WTAP antibody [EPR18744] (ab195380)
    Western blot - Anti-WTAP antibody [EPR18744] (ab195380)

    Lane 1: Wild-type HAP1 whole cell lysate (20 µg)
    Lane 2: WTAP knockout HAP1 whole cell lysate (20 µg)
    Lane 3: MOLT-4 whole cell lysate (20 µg)
    Lane 4: K562 whole cell lysate (20 µg)

    Lanes 1 - 4: Merged signal (red and green). Green - ab195380 observed at 50 kDa. Red - loading control, ab18058, observed at 130 kDa.

    ab195380 was shown to specifically react with WTAP in wild-type HAP1 cells as signal was lost in WTAP knockout cells. Wild-type and WTAP knockout samples were subjected to SDS-PAGE. Ab195380 and ab18058 (Mouse anti-Vinculin loading control) were incubated overnight at 4°C at 1/1000 dilution and 1/20000 dilution respectively. Blots were developed with Goat anti-Rabbit IgG H&L (IRDye® 800CW) preabsorbed ab216773 and Goat anti-Mouse IgG H&L (IRDye® 680RD) preabsorbed ab216776 secondary antibodies at 1/10000 dilution for 1 hour at room temperature before imaging.

  • Immunocytochemistry/ Immunofluorescence - Anti-WTAP antibody [EPR18744] (ab195380)
    Immunocytochemistry/ Immunofluorescence - Anti-WTAP antibody [EPR18744] (ab195380)

    Immunofluorescent analysis of 4% paraformaldehyde-fixed, 0.1% Triton X-100 permeabilized Jurkat (Human T cell leukemia cell line from peripheral blood) cells labeling WTAP with ab195380 at 1/500 dilution, followed by Goat Anti-Rabbit IgG (Alexa Fluor® 488) (ab150077) secondary antibody at 1/1000 dilution (green). Confocal image showing nuclear staining on Jurkat cell line. The nuclear counter stain is DAPI (blue). 

    Tubulin is detected with Anti-alpha Tubulin mouse MAb (ab7291) at 1/1000 dilution, followed by Goat Anti-Mouse IgG H&L (Alexa Fluor® 594) (ab150120) secondary antibody  at 1/1000 dilution (red).

    The negative controls are as follows:
    -ve control 1: ab195380 at 1/500 dilution, followed by Goat Anti-Mouse IgG H&L (Alexa Fluor® 594) (ab150120) secondary antibody at 1/1000 dilution.
    -ve control 2: Anti-alpha Tubulin mouse MAb (ab7291) at 1/1000 dilution, followed by Goat Anti-Rabbit IgG H&L (Alexa Fluor® 488) (ab150077) secondary antibody at 1/1000 dilution.

  • Western blot - Anti-WTAP antibody [EPR18744] (ab195380)
    Western blot - Anti-WTAP antibody [EPR18744] (ab195380)
    All lanes : Anti-WTAP antibody [EPR18744] (ab195380) at 1/1000 dilution

    Lane 1 : Jurkat (Human T cell leukemia cell line from peripheral blood) whole cell lysate
    Lane 2 : K562 (Human chronic myelogenous leukemia cell line from bone marrow ) whole cell lysate
    Lane 3 : HeLa (Human epithelial cell line from cervix adenocarcinoma) whole cell lysate
    Lane 4 : HepG2 (Human liver hepatocellular carcinoma cell line) 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

    Developed using the ECL technique.

    Predicted band size: 44 kDa
    Observed band size: 50 kDa
    why is the actual band size different from the predicted?


    Exposure time: 3 minutes


    Blocking/Dilution buffer: 5% NFDM/TBST.

    The observed molecular weight is consistent with what has been described in the literature (PMID: 17088532).

  • Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-WTAP antibody [EPR18744] (ab195380)
    Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-WTAP antibody [EPR18744] (ab195380)

    Immunohistochemical analysis of paraffin-embedded human endometrium tissue labeling WTAP with ab195380 at 1/100 dilution, followed by Goat Anti-Rabbit IgG H&L (HRP) (ab97051) at 1/500 dilution. Nuclear staining on human endometrium is observed. Counter stained with Hematoxylin.

    Secondary antibody only control: Used PBS instead of primary antibody, secondary antibody is Goat Anti-Rabbit IgG H&L (HRP) (ab97051) at 1/500 dilution.

    Perform heat mediated antigen retrieval with Tris/EDTA buffer pH 9.0 before commencing with IHC staining protocol.

  • Flow Cytometry - Anti-WTAP antibody [EPR18744] (ab195380)
    Flow Cytometry - Anti-WTAP antibody [EPR18744] (ab195380)

    Flow cytometric analysis of 4% paraformaldehyde-fixed Jurkat (Human T cell leukemia cell line from peripheral blood) cells labeling WTAP with ab195380 at 1/600 dilution (red) compared with a rabbit monoclonal IgG isotype control (ab172730; black) and an unlabelled control (cells without incubation with primary antibody and secondary antibody; blue). Goat Anti-Rabbit IgG (Alexa Fluor® 488) at 1/2000 dilution was used as the secondary antibody.

  • Western blot - Anti-WTAP antibody [EPR18744] (ab195380)
    Western blot - Anti-WTAP antibody [EPR18744] (ab195380)
    All lanes : Anti-WTAP antibody [EPR18744] (ab195380) at 1/1000 dilution

    Lane 1 : Human kidney lysate
    Lane 2 : Human thymus lysate
    Lane 3 : Human lung lysate

    Lysates/proteins at 10 µg per lane.

    Secondary
    All lanes : Goat Anti-Rabbit IgG Peroxidase Conjugate, specific to the non-reduced form of IgG at 1/100000 dilution

    Developed using the ECL technique.

    Predicted band size: 44 kDa
    Observed band size: 50 kDa why is the actual band size different from the predicted?


    Exposure time: 3 minutes


    Blocking/Dilution buffer: 5% NFDM/TBST.

    The observed molecular weight is consistent with what has been described in the literature (PMID: 17088532).

  • Immunocytochemistry/ Immunofluorescence - Anti-WTAP antibody [EPR18744] (ab195380)
    Immunocytochemistry/ Immunofluorescence - Anti-WTAP antibody [EPR18744] (ab195380)

    Immunofluorescent analysis of 4% paraformaldehyde-fixed, 0.1% Triton X-100 permeabilized HeLa (Human epithelial cell line from cervix adenocarcinoma) cells labeling WTAP with ab195380 at 1/500 dilution, followed by Goat Anti-Rabbit IgG (Alexa Fluor® 488) (ab150077) secondary antibody at 1/1000 dilution (green). Confocal image showing nuclear staining on HeLa cell line. The nuclear counter stain is DAPI (blue). 

    Tubulin is detected with Anti-alpha Tubulin mouse MAb (ab7291) at 1/1000 dilution, followed by Goat Anti-Mouse IgG H&L (Alexa Fluor® 594) (ab150120) secondary antibody at 1/1000 dilution (red).

    The negative controls are as follows:
    -ve control 1: ab195380 at 1/500 dilution, followed by Goat Anti-Mouse IgG H&L (Alexa Fluor® 594) (ab150120) secondary antibody at 1/1000 dilution.
    -ve control 2: Anti-alpha Tubulin mouse MAb (ab7291) at 1/1000 dilution, followed by Goat Anti-Rabbit IgG (Alexa Fluor® 488) (ab150077) secondary antibody at 1/1000 dilution.

  • Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-WTAP antibody [EPR18744] (ab195380)
    Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-WTAP antibody [EPR18744] (ab195380)

    Immunohistochemical analysis of paraffin-embedded human liver cancer tissue labeling WTAP with ab195380 at 1/100 dilution, followed by Goat Anti-Rabbit IgG H&L (HRP) (ab97051) at 1/500 dilution. Nuclear staining on human liver cancer is observed. Counter stained with Hematoxylin.

    Secondary antibody only control: Used PBS instead of primary antibody, secondary antibody is Goat Anti-Rabbit IgG H&L (HRP) (ab97051) at 1/500 dilution.

    Perform heat mediated antigen retrieval with Tris/EDTA buffer pH 9.0 before commencing with IHC staining protocol.

  • Immunoprecipitation - Anti-WTAP antibody [EPR18744] (ab195380)
    Immunoprecipitation - Anti-WTAP antibody [EPR18744] (ab195380)

    WTAP was immunoprecipitated from 0.35 mg of Jurkat (Human T cell leukemia cell line from peripheral blood) whole cell lysate with ab195380 at 1/40 dilution. Western blot was performed from the immunoprecipitate using ab195380 at 1/1000 dilution. VeriBlot for IP Detection Reagent (HRP) (ab131366), was used for detection at 1/10000 dilution.

    Lane 1: Jurkat whole cell lysate 10µg (Input).

    Lane 2: ab195380 IP in Jurkat whole cell lysate.

    Lane 3: Rabbit monoclonal IgG (ab172730) instead of ab195380 in Jurkat whole cell lysate.

    Blocking and dilution buffer and concentration: 5% NFDM/TBST.

    Exposure time: 30 seconds.

  • Anti-WTAP antibody [EPR18744] (ab195380)
    Anti-WTAP antibody [EPR18744] (ab195380)

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