Recombinant
RabMAb

Recombinant Anti-TNF alpha antibody [EPR19147] (ab183218)

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Overview

  • Product name

    Anti-TNF alpha antibody [EPR19147]
    See all TNF alpha primary antibodies

  • Description

    Rabbit monoclonal [EPR19147] to TNF alpha

  • Host species

    Rabbit

  • Tested applications

    Suitable for: WB, IP, ELISA, ICC/IFmore details

  • Species reactivity

    Reacts with: Mouse, Human

  • Immunogen

    Synthetic peptide within Human TNF alpha aa 1-100. The exact sequence is proprietary.
    Database link: P01375

  • 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

    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

Applications

Our Abpromise guarantee covers the use of ab183218 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 1/1000. Detects a band of approximately 26, 14 kDa (predicted molecular weight: 25 kDa).
IP 1/40.
ELISA Use at an assay dependent concentration.
ICC/IF 1/5000.

Target

  • Function

    Cytokine that binds to TNFRSF1A/TNFR1 and TNFRSF1B/TNFBR. It is mainly secreted by macrophages and can induce cell death of certain tumor cell lines. It is potent pyrogen causing fever by direct action or by stimulation of interleukin-1 secretion and is implicated in the induction of cachexia, Under certain conditions it can stimulate cell proliferation and induce cell differentiation.

  • Involvement in disease

    Genetic variations in TNF are a cause of susceptibility psoriatic arthritis (PSORAS) [MIM:607507]. PSORAS is an inflammatory, seronegative arthritis associated with psoriasis. It is a heterogeneous disorder ranging from a mild, non-destructive disease to a severe, progressive, erosive arthropathy. Five types of psoriatic arthritis have been defined: asymmetrical oligoarthritis characterized by primary involvement of the small joints of the fingers or toes; asymmetrical arthritis which involves the joints of the extremities; symmetrical polyarthritis characterized by a rheumatoidlike pattern that can involve hands, wrists, ankles, and feet; arthritis mutilans, which is a rare but deforming and destructive condition; arthritis of the sacroiliac joints and spine (psoriatic spondylitis).

  • Sequence similarities

    Belongs to the tumor necrosis factor family.

  • Post-translational
    modifications

    The soluble form derives from the membrane form by proteolytic processing.
    The membrane form, but not the soluble form, is phosphorylated on serine residues. Dephosphorylation of the membrane form occurs by binding to soluble TNFRSF1A/TNFR1.
    O-glycosylated; glycans contain galactose, N-acetylgalactosamine and N-acetylneuraminic acid.

  • Cellular localization

    Secreted and Cell membrane.
  • Information by UniProt

  • Database links

    • Alternative names

      • APC1 antibody
      • APC1 protein antibody
      • Cachectin antibody
      • DIF antibody
      • Differentiation inducing factor antibody
      • Macrophage cytotoxic factor antibody
      • Tnf antibody
      • TNF superfamily member 2 antibody
      • TNF superfamily, member 2 antibody
      • TNF, macrophage derived antibody
      • TNF, monocyte derived antibody
      • TNF-a antibody
      • TNF-alpha antibody
      • TNFA antibody
      • TNFA_HUMAN antibody
      • TNFSF2 antibody
      • Tumor necrosis factor (TNF superfamily member 2) antibody
      • Tumor necrosis factor alpha antibody
      • Tumor necrosis factor antibody
      • Tumor necrosis factor ligand superfamily member 2 antibody
      • Tumor Necrosis Factor, Membrane Form antibody
      • Tumor necrosis factor, soluble form antibody
      see all

    Images

    • Western blot - Anti-TNF alpha antibody [EPR19147] (ab183218)
      Western blot - Anti-TNF alpha antibody [EPR19147] (ab183218)
      All lanes : Anti-TNF alpha antibody [EPR19147] (ab183218) at 1/2000 dilution

      Lane 1 : TPA pretreated THP-1 (Human monocytic leukemia cell line) whole cell lysate
      Lane 2 : TPA pretreated THP-1 (Human monocytic leukemia cell line) whole cell lysate treated with 100 ng/ml LPS for 7 hours
      Lane 3 : 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

      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: 25 kDa
      Observed band size: 14,26 kDa
      why is the actual band size different from the predicted?


      Exposure time: 3 minutes


      Blocking/Dilution buffer: 5% NFDM/TBST.

      The 14kDa band probably is the degradable inactive fragment that has been decribed by literature (PMID: 9933416).

    • Immunocytochemistry/ Immunofluorescence - Anti-TNF alpha antibody [EPR19147] (ab183218)
      Immunocytochemistry/ Immunofluorescence - Anti-TNF alpha antibody [EPR19147] (ab183218)

      Immunocytochemistry/ Immunofluorescence analysis of RAW264.7(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.

    • ELISA - Anti-TNF alpha antibody [EPR19147] (ab183218)
      ELISA - Anti-TNF alpha antibody [EPR19147] (ab183218)
      ELISA using ab183218 between 0.1 and 1000 ng/ml to detect TNF alpha peptide. Secondary used was Alkaline Phosphatase AffiniPure Goat Anti-Rabbit IgG (H+L) 1/2000.
    • Western blot - Anti-TNF alpha antibody [EPR19147] (ab183218)
      Western blot - Anti-TNF alpha antibody [EPR19147] (ab183218)
      All lanes : Anti-TNF alpha antibody [EPR19147] (ab183218) at 1/1000 dilution

      Lane 1 : TPA pretreated RAW 264.7 (Mouse macrophage cell line transformed with Abelson murine leukemia virus) whole cell lysate
      Lane 2 : TPA pretreated RAW 264.7 (Mouse macrophage cell line transformed with Abelson murine leukemia virus) whole cell lysate treated with 100 ng/ml LPS for 7 hours
      Lane 3 : TPA pretreated RAW 264.7 (Mouse macrophage cell line transformed with Abelson murine leukemia virus) whole cell lysate treated with 100 ng/ml LPS for 4 hours, then added 1 µg/ml BFA for 3 hours

      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: 25 kDa
      Observed band size: 14,26,33 kDa why is the actual band size different from the predicted?


      Exposure time: 3 seconds


      Blocking/Dilution buffer: 5% NFDM/TBST.

      The 33kDa band probably is the dimeric intermediate form that has been described in the literature (PMID: 9933416).

    • Immunoprecipitation - Anti-TNF alpha antibody [EPR19147] (ab183218)
      Immunoprecipitation - Anti-TNF alpha antibody [EPR19147] (ab183218)

      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.

    References

    This product has been referenced in:

    • Liang Z & Ren C Emodin attenuates apoptosis and inflammation induced by LPS through up-regulating lncRNA TUG1 in murine chondrogenic ATDC5 cells. Biomed Pharmacother 103:897-902 (2018). WB ; Mouse . Read more (PubMed: 29710506) »
    • Yang QC  et al. A murine model of dry eye induced by topical administration of erlotinib eye drops. Int J Mol Med 41:1427-1436 (2018). Read more (PubMed: 29286080) »
    See all 3 Publications for this product

    Publishing research using ab183218? Please let us know so that we can cite the reference in this datasheet.

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