Astana Biomed Group, an authorized Abcam distributor in Central Asia

Anti-ATG4B antibody [EPR16572] (ab199537)

Key features and details

  • Produced recombinantly (animal-free) for high batch-to-batch consistency and long term security of supply
  • Rabbit monoclonal [EPR16572] to ATG4B
  • Suitable for: WB, IP
  • Knockout validated
  • Reacts with: Rat, Human

Overview

  • Product name

    Anti-ATG4B antibody [EPR16572]
    See all ATG4B primary antibodies

  • Description

    Rabbit monoclonal [EPR16572] to ATG4B

  • Host species

    Rabbit

  • Tested Applications & Species

    Application Species
    IP
    Human
    WB
    Rat
    Human
    See all applications and species data

  • Immunogen

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

  • Positive control

    • WB: 293T, Ramos, HeLa, Jurkat and PC-12 whole cell lysates; Rat brain lysate. IP: Jurkat and Ramos whole cell lysates.

  • 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

Images

  • Western blot - Anti-ATG4B antibody [EPR16572] (ab199537)
    Western blot - Anti-ATG4B antibody [EPR16572] (ab199537)
    All lanes : Anti-ATG4B antibody [EPR16572] (ab199537) at 1/1000 dilution

    Lane 1 : Wild-type HeLa cell lysate
    Lane 2 : ATG4B knockout HeLa cell lysate

    Lysates/proteins at 20 µg per lane.

    Performed under reducing conditions.

    Predicted band size: 44 kDa
    Observed band size: 47 kDa
    why is the actual band size different from the predicted?



    Lanes 1 - 2: Merged signal (red and green). Green - ab199537 observed at 47 kDa. Red - loading control ab8245 (Mouse anti-GAPDH antibody [6C5]) observed at 37 kDa.

    ab199537 was shown to react with ATG4B in wild-type HeLa cells in Western blot with loss of signal observed in ATG4B knockout cell line ab265814 (ATG4B knockout cell lysate ab257364). Wild-type and ATG4B knockout HeLa cell lysates were subjected to SDS-PAGE. Membranes were blocked in 3 % milk in TBS-T (0.1 % Tween®) before incubation with ab199537 and ab8245 (Mouse anti-GAPDH antibody [6C5]) overnight at 4 °C at a 1 in 1000 dilution and a 1 in 20000 dilution respectively. Blots were incubated 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 in 20000 dilution for 1 h at room temperature before imaging.

  • Western blot - Anti-ATG4B antibody [EPR16572] (ab199537)
    Western blot - Anti-ATG4B antibody [EPR16572] (ab199537)

    Lane 1: Wild-type HAP1 cell lysate (20 µg)
    Lane 2: ATG4B knockout HAP1 cell lysate (20 µg)
    Lane 3: HeLa cell lysate (20 µg)
    Lane 4: Jurkat cell lysate (20 µg)
    Lanes 1 - 4: Merged signal (red and green). Green - ab199537 observed at 44 kDa. Red - loading control, ab8245, observed at 37 kDa.

    ab199537 was shown to specifically react with ATG4B when ATG4B knockout samples were used. Wild-type and ATG4B knockout samples were subjected to SDS-PAGE. ab199537 and ab8245 (loading control to GAPDH) were diluted 1/1000 and 1/10000 respectively and incubated overnight at 4°C. Blots were developed with Goat anti-Rabbit IgG H&L (IRDye® 800CW) preadsorbed ab216773 and Goat anti-Mouse IgG H&L (IRDye® 680RD) preadsorbed ab216776
    secondary antibodies at 1/10000 dilution for 1 hour at room temperature before imaging.

  • Western blot - Anti-ATG4B antibody [EPR16572] (ab199537)
    Western blot - Anti-ATG4B antibody [EPR16572] (ab199537)
    All lanes : Anti-ATG4B antibody [EPR16572] (ab199537) at 1/1000 dilution

    Lane 1 : 293T (Human epithelial cells from embryonic kidney) whole cell lysate
    Lane 2 : Ramos (Human Burkitt's lymphoma cell line) whole cell lysate
    Lane 3 : HeLa (Human epithelial cells from cervix adenocarcinoma) whole cell lysate
    Lane 4 : Jurkat (Human T cell leukemia cells from peripheral blood) whole cell lysate

    Lysates/proteins at 20 µg per lane.

    Secondary
    All lanes : Goat Anti-Rabbit IgG, (H+L), Peroxidase conjugated at 1/1000 dilution

    Predicted band size: 44 kDa
    Observed band size: 44 kDa


    Exposure time: 1 minute


    Blocking/Dilution buffer: 5% NFDM/TBST.

  • Western blot - Anti-ATG4B antibody [EPR16572] (ab199537)
    Western blot - Anti-ATG4B antibody [EPR16572] (ab199537)
    All lanes : Anti-ATG4B antibody [EPR16572] (ab199537) at 1/1000 dilution

    Lane 1 : Rat brain lysate
    Lane 2 : PC-12 (Rat adrenal gland pheochromocytoma) whole cell lysate

    Lysates/proteins at 10 µg per lane.

    Secondary
    All lanes : Goat Anti-Rabbit IgG, (H+L), Peroxidase conjugated at 1/1000 dilution

    Predicted band size: 44 kDa
    Observed band size: 44 kDa


    Exposure time: 1 minute


    Blocking/Dilution buffer: 5% NFDM/TBST.

  • Immunoprecipitation - Anti-ATG4B antibody [EPR16572] (ab199537)
    Immunoprecipitation - Anti-ATG4B antibody [EPR16572] (ab199537)

    ATG4B was immunoprecipitated from 1mg of Jurkat (Human T cell leukemia cells from peripheral blood) whole cell lysate with ab199537 at 1/50 dilution. Western blot was performed from the immunoprecipitate using ab199537 at 1/1000 dilution. Anti-Rabbit IgG (HRP), specific to the non-reduced form of IgG, was used as secondary antibody at 1/1500 dilution.

    Lane 1: Jurkat whole cell lysate 10ug (Input). Lane 2: ab199537 IP in Jurkat whole cell lysate. Lane 3: Rabbit monoclonal IgG (ab172730) instead of ab199537 in Jurkat whole cell lysate.

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

    Exposure time: 3 minutes.

  • Immunoprecipitation - Anti-ATG4B antibody [EPR16572] (ab199537)
    Immunoprecipitation - Anti-ATG4B antibody [EPR16572] (ab199537)

    ATG4B was immunoprecipitated from 1mg of Ramos (Human Burkitt's lymphoma cell line) whole cell lysate with ab199537 at 1/60 dilution. Western blot was performed from the immunoprecipitate using ab199537 at 1/500 dilution (Panel A) or ab154843 at 1/500 dilution (Panel B). Anti-Rabbit IgG (HRP), specific to the non-reduced form of IgG, was used as secondary antibody at 1/1500 dilution.

    Lane 1: Ramos whole cell lysate 10ug (Input). Lane 2: ab199537 IP in Ramos whole cell lysate. Lane 3: Rabbit monoclonal IgG (ab172730) instead of ab199537 in Ramos whole cell lysate.

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

    Exposure time: Panel A and B 30 seconds.

  • Anti-ATG4B antibody [EPR16572] (ab199537)
    Anti-ATG4B antibody [EPR16572] (ab199537)

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