Astana Biomed Group, an authorized Abcam distributor in Central Asia

Anti-TNF alpha antibody [EPR19147] - Low endotoxin, Azide free (ab222500)

Key features and details

  • Produced recombinantly (animal-free) for high batch-to-batch consistency and long term security of supply
  • Rabbit monoclonal [EPR19147] to TNF alpha - Low endotoxin, Azide free
  • Suitable for: IP, ICC/IF, ELISA, WB
  • Knockout validated
  • Reacts with: Mouse, Human, Recombinant fragment

Overview

  • Product name

    Anti-TNF alpha antibody [EPR19147] - Low endotoxin, Azide free
    See all TNF alpha primary antibodies

  • Description

    Rabbit monoclonal [EPR19147] to TNF alpha - Low endotoxin, Azide free

  • Host species

    Rabbit

  • Tested applications

    Suitable for: IP, ICC/IF, ELISA, WBmore details

  • Species reactivity

    Reacts with: Mouse, Human, Recombinant fragment

  • Immunogen

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

  • Positive control

    • WB: TPA pretreated THP-1 treated with 100 ng/ml LPS for 7 hours and TPA pretreated THP-1 treated with 100 ng/ml LPS for 4 hours, then added 1 µg/ml BFA for 3 hours whole cell lysates; TPA pretreated RAW 264.7, TPA pretreated RAW 264.7 treated with 100 ng/ml LPS for 7 hours and TPA pretreated RAW 264.7 treated with 100 ng/ml LPS for 4 hours, then added 1 µg/ml BFA for 3 hours whole cell lysates. IP: TPA pretreated THP-1 treated with 100 ng/ml LPS for 4 hours, then added 1 µg/ml BFA for 3 hours cell lysate.

  • General notes

    ab222500 is a carrier-free antibody designed for use in antibody labeling, including fluorochromes, metal isotopes, oligonucleotides, enzymes.

     

    Our Low endotoxin, azide-free formats have low endotoxin level (≤ 1 EU/ml, determined by the LAL assay) and are free from azide, to achieve consistent experimental results in functional assays.

    Our RabMAb® technology is a patented hybridoma-based technology for making rabbit monoclonal antibodies. For details on our patents, please refer to RabMAb® patents.

    Reproducibility is key to advancing scientific discovery and accelerating scientists’ next breakthrough.

    Abcam is leading the way with our range of recombinant antibodies, knockout-validated antibodies and knockout cell lines, all of which support improved reproducibility.

    We are also planning to innovate the way in which we present recommended applications and species on our product datasheets, so that only applications & species that have been tested in our own labs, our suppliers or by selected trusted collaborators are covered by our Abpromise™ guarantee.

    In preparation for this, we have started to update the applications & species that this product is Abpromise guaranteed for.

    We are also updating the applications & species that this product has been “predicted to work with,” however this information is not covered by our Abpromise guarantee.

    Applications & species from publications and Abreviews that have not been tested in our own labs or in those of our suppliers are not covered by the Abpromise guarantee.

    Please check that this product meets your needs before purchasing. If you have any questions, special requirements or concerns, please send us an inquiry and/or contact our Support team ahead of purchase. Recommended alternatives for this product can be found below, as well as customer reviews and Q&As.

Properties

Images

  • Western blot - Anti-TNF alpha antibody [EPR19147] - Low endotoxin, Azide free (ab222500)
    Western blot - Anti-TNF alpha antibody [EPR19147] - Low endotoxin, Azide free (ab222500)
    All lanes : Anti-TNF alpha antibody [EPR19147] (ab183218) at 1/1000 dilution

    Lane 1 : Wild-type THP-1 Brefeldin A (ab120299) treated (5 µg/ml, 4 h) cell lysate
    Lane 2 : Wild-type THP-1 LPS treated (100 ng/ml, 16 h) and Brefeldin A (ab120299) treated (5 µg/ml, 4 h) cell lysate
    Lane 3 : TNF alpha knockout THP-1 Brefeldin A (ab120299) treated (5 µg/ml, 4 h) cell lysate
    Lane 4 : TNF alpha knockout THP-1 LPS treated (100 ng/ml, 16 h) and Brefeldin A (ab120299) treated (5 µg/ml, 4 h) cell lysate
    Lane 5 : U937 PMA treated (10 mM, 2 days) plus 16 h no treatment and Brefeldin A (ab120299) treated (5 µg/ml, 4 h) cell lysate
    Lane 6 : U937 PMA treated (10 mM, 2 days) and LPS treated (1 µg/ml, 16 h) plus Brefeldin A (ab120299) treated (5 µg/ml, 4 h) cell lysate

    Lysates/proteins at 30 µg per lane.

    Performed under reducing conditions.

    Observed band size: 26 kDa
    why is the actual band size different from the predicted?



    This data was developed using the same antibody clone in a different buffer formulation (ab183218).

    Lanes 1 - 6: Merged signal (red and green). Green - ab183218 observed at 26 kDa. Red - loading control ab7291 (Mouse anti-Alpha Tubulin [DM1A]) observed at 55 kDa.

    ab183218 was shown to react with TNF alpha in THP-1 wild-type cells in Western blot with loss of signal observed in TNF knockout sample. Wild-type and TNF knockout THP-1 cell lysates were subjected to SDS-PAGE. Membranes were blocked in fluorescent western blot (TBS-based) blocking solution before incubation with ab183218 and ab7291 (Mouse anti-Alpha Tubulin [DM1A]) 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.

  • Immunocytochemistry/ Immunofluorescence - Anti-TNF alpha antibody [EPR19147] - Low endotoxin, Azide free (ab222500)
    Immunocytochemistry/ Immunofluorescence - Anti-TNF alpha antibody [EPR19147] - Low endotoxin, Azide free (ab222500)

    Immunocytochemistry/ Immunofluorescence analysis of RAW 264.7(Mouse Abelson murine leukemia virus-induced tumor macrophage) treated with 100ng/ml LPS for 7 h and 1µg/ml BFA for the last 3h cells labeling TNF alpha with purified ab183218 at 1/5000 dilution (0.09 µg/ml). Cells were fixed in 4% paraformaldehyde and permeabilized with 0.1% Triton X-100. Cells were counterstained with Ab195889 Anti-alpha Tubulin antibody [DM1A] - Microtubule Marker (Alexa Fluor® 594) 1/200 (2.5 µg/ml). Goat anti rabbit IgG (Alexa Fluor® 488, ab150077) was used as the secondary antibody at 1/1000 dilution (2 µg/ml) dilution. DAPI was used as nuclear counterstain. PBS instead of the primary antibody was used as the secondary antibody only control.

    This data was developed using the same antibody clone in a different buffer formulation containing PBS, BSA, glycerol, and sodium azide (ab183218).

  • ELISA - Anti-TNF alpha antibody [EPR19147] - Low endotoxin, Azide free (ab222500)
    ELISA - Anti-TNF alpha antibody [EPR19147] - Low endotoxin, Azide free (ab222500)

    ELISA using ab183218 between 0.1 and 1000 ng/ml to detect human TNF alpha peptide. Secondary used was Alkaline Phosphatase AffiniPure Goat Anti-Rabbit IgG (H+L) 1/2000.

    This data was developed using the same antibody clone in a different buffer formulation containing PBS, BSA, glycerol, and sodium azide (ab183218).

  • Immunoprecipitation - Anti-TNF alpha antibody [EPR19147] - Low endotoxin, Azide free (ab222500)
    Immunoprecipitation - Anti-TNF alpha antibody [EPR19147] - Low endotoxin, Azide free (ab222500)

    TNF alpha was immunoprecipitated from 0.35 mg of TPA pretreated THP-1 (Human monocytic leukemia cell line) whole cell lysate treated with 100 ng/ml LPS for 4 hours, then added 1 μg/ml BFA for 3 hours with ab183218 at 1/40 dilution. Western blot was performed from the immunoprecipitate using ab183218 at 1/1000 dilution. VeriBlot for IP Detection Reagent (HRP) (ab131366), was used for detection at 1/1000 dilution.

    Lane 1: TPA pretreated THP-1 whole cell lysate treated with 100 ng/ml LPS for 4 hours, then added 1 μg/ml BFA for 3 hours 10µg (Input).

    Lane 2: ab183218 IP in TPA pretreated THP-1 whole cell lysate treated with 100 ng/ml LPS for 4 hours, then added 1 μg/ml BFA for 3 hours.

    Lane 3: Rabbit IgG,monoclonal [EPR25A]-Isotype Control (ab172730) instead of ab183218 in TPA pretreated THP-1 whole cell lysate treated with 100 ng/ml LPS for 4 hours, then added 1 μg/ml BFA for 3 hours.

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

    Exposure time: 1 second.

    This data was developed using the same antibody clone in a different buffer formulation containing PBS, BSA, glycerol, and sodium azide (ab183218).

  • Anti-TNF alpha antibody [EPR19147] - Low endotoxin, Azide free (ab222500)
    Anti-TNF alpha antibody [EPR19147] - Low endotoxin, Azide free (ab222500)

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