Overview

  • Product name
    Human TNF alpha ELISA Kit, Fluorescent
    See all TNF alpha kits
  • Detection method
    Fluorescent
  • Precision
    Intra-assay
    Sample n Mean SD CV%
    Supernatant 8 2.3%
    Inter-assay
    Sample n Mean SD CV%
    Supernatant 3 5%
  • Sample type
    Cell culture supernatant, Serum, Heparin Plasma, EDTA Plasma, Citrate Plasma
  • Assay type
    Sandwich (quantitative)
  • Sensitivity
    2.4 pg/ml
  • Range
    3.91 pg/ml - 8000 pg/ml
  • Recovery

    Sample specific recovery
    Sample type Average % Range
    Serum 99.5 80% - 119%
    Cell culture media 100.97 95% - 109%
    Heparin Plasma 98 91% - 106%
    EDTA Plasma 102 95.7% - 110%
    Citrate Plasma 93.09 83.5% - 99%

  • Assay time
    1h 30m
  • Assay duration
    One step assay
  • Species reactivity
    Reacts with: Human
    Does not react with: Mouse
  • Product overview

    TNF-alpha in vitro CatchPoint SimpleStep ELISA (Enzyme-Linked Immunosorbent Assay) kit is designed for the quantitative measurement of TNF-alpha protein in human serum, plasma, and cell culture supernatants.


    This CatchPoint SimpleStep ELISA kit has been optimized for Molecular Devices Microplate Readers. Click here for a list of recommended Microplate Readers.
    If using a Molecular Devices’ plate reader supported by SoftMax® Pro software, a preconfigured protocol for these CatchPoint SimpleStep ELISA Kits is available with all the protocol and analysis settings at www.softmaxpro.org.


    The CatchPoint SimpleStep ELISA employs an affinity tag labeled capture antibody and a reporter conjugated detector antibody which immunocapture the sample analyte in solution. This entire complex (capture antibody/analyte/detector antibody) is in turn immobilized via immunoaffinity of an anti-tag antibody coating the well. To perform the assay, samples or standards are added to the wells, followed by the antibody mix. After incubation, the wells are washed to remove unbound material. CatchPoint HRP Development Solution containing the Stoplight Red Substrate is added. During incubation, the substrate is catalyzed by HRP generating a fluorescent product. Signal is generated proportionally to the amount of bound analyte and the intensity is measured in a fluorescence plater reader at 530/570/590 nm Excitation/Cutoff/Emission.

  • Notes

    TNF-alpha, also known as cachectin or TNFSF1A, is the prototypic ligand of the TNF superfamily which plays a central role in inflammation, apoptosis, proliferation, invasion, angiogenesis, metastasis and morphogenesis.   It is expressed on macrophages, endothelial, epithelial and tumor cells as a 26kDa transmembrane protein. TNF-alpha is cleaved by proteolytic processing into six chains: (1) TNF membrane form, (2) Intracellular domain 1, (3) Intracellular domain 2, (4) C-domain 1, (5) C-domain 2 and (6) TNF soluble form. Signaling from TNF-alpha differs depending on the type of ligand initiating the signaling event (intracellular, membrane or soluble). As an example, the membrane form of TNF-alpha appears to mediate anti-tumorigenic therapeutic responses whereas the soluble ligand is linked to inflammation and proliferation.

  • Tested applications
    Suitable for: Sandwich ELISAmore details
  • Platform
    Pre-coated microplate (12 x 8 well strips)

Properties

  • Storage instructions
    Store at +4°C. Please refer to protocols.
  • Components 1 x 96 tests
    100X Stoplight Red Substrate 1 x 120µl
    10X Wash Buffer PT (ab206977) 1 x 20ml
    500X Hydrogen Peroxide (H2O2, 3%) 1 x 50µl
    Antibody Diluent 5BI 1 x 6ml
    Plate Seals 1 unit
    Sample Diluent 50BP 1 x 20ml
    Sample Diluent NS 1 x 50ml
    SimpleStep Pre-Coated Black 96-Well Microplate 1 unit
    Stoplight Red Substrate Buffer 1 x 12ml
    Human TNFa Capture Antibody (Lyophilized) 2 vials
    Human TNFa Detector Antibody (Lyophilized) 2 vials
    Human TNFa Lyophilized Recombinant Protein 2 vials
  • Research areas
  • 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
  • Alternative names
    • APC1
    • APC1 protein
    • Cachectin
    • DIF
    • Differentiation inducing factor
    • Macrophage cytotoxic factor
    • Tnf
    • TNF superfamily, member 2
    • TNF, macrophage derived
    • TNF, monocyte derived
    • TNF-a
    • TNF-alpha
    • TNFA
    • TNFA_HUMAN
    • TNFSF2
    • Tumor necrosis factor
    • Tumor necrosis factor alpha
    • Tumor necrosis factor ligand superfamily member 2
    • Tumor Necrosis Factor, Membrane Form
    • Tumor necrosis factor, soluble form
    see all
  • Database links

Applications

Our Abpromise guarantee covers the use of ab229399 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
Sandwich ELISA Use at an assay dependent concentration.

Images

  • ELISA Protocol Summary
  • Background-subtracted data values (mean +/- SD) are graphed.

  • Human PBMCs were cultured in RPMI supplemented with 10% fetal calf serum, 2mM L-glutamine, 100 U/mL penicillin and 100 µg/mL streptomycin. Cells were cultured for 2 days at 37°C in the presence or absence of PHA. The concentrations of TNF-alpha were interpolated from the calibration curve and corrected for sample dilution. The mean TNF-alpha concentration was determined to be undetectable in basal media, 1.2 ng/mL in unstimulated PMBC supernatants and 4.7 ng/mL in stimulated PBMC supernatants.

    Ten individual healthy donors were evaluated for the presence of TNFalpha in serum using this assay. All samples had undetectable levels of TNF-alpha.

Protocols

References

ab229399 has not yet been referenced specifically in any publications.

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