The Human TNF alpha ELISA kit is a single-wash 90-min SimpleStep ELISA® used to quantify Human TNF alpha with a sensitivity of 4.32 pg/ml. The assay uses a simple Mix-Wash-Read protocol with just one incubation and wash step.Easy-to-use, rapid, sensitive human TNF alpha sandwich ELISA kit:- Colorimetric assay - 450nm readout; works on any plate reader- Different formats for different needs: 10x96 plates for bulk orders and 384-well for higher throughput- Broad sample compatibility – works with serum, plasma, cell culture supernatants, and other biological fluids
Colorimetric
Cerebral Spinal Fluid, Heparin Plasma, Citrate plasma, Cell culture supernatant, Serum, EDTA Plasma
Sandwich (quantitative)
Human
15.63 - 1000 pg/mL
1h 30m
= 4.32 pg/mL
Application | Reactivity | Dilution info | Notes |
---|---|---|---|
Application sELISA | Reactivity Reacts | Dilution info - | Notes - |
Select an associated product type
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. Impairs regulatory T-cells (Treg) function in individuals with rheumatoid arthritis via FOXP3 dephosphorylation. Up-regulates the expression of protein phosphatase 1 (PP1), which dephosphorylates the key 'Ser-418' residue of FOXP3, thereby inactivating FOXP3 and rendering Treg cells functionally defective (PubMed:23396208). Key mediator of cell death in the anticancer action of BCG-stimulated neutrophils in combination with DIABLO/SMAC mimetic in the RT4v6 bladder cancer cell line (PubMed:16829952, PubMed:22517918, PubMed:23396208). Induces insulin resistance in adipocytes via inhibition of insulin-induced IRS1 tyrosine phosphorylation and insulin-induced glucose uptake. Induces GKAP42 protein degradation in adipocytes which is partially responsible for TNF-induced insulin resistance (By similarity). Plays a role in angiogenesis by inducing VEGF production synergistically with IL1B and IL6 (PubMed:12794819). Promotes osteoclastogenesis and therefore mediates bone resorption (By similarity).The TNF intracellular domain (ICD) form induces IL12 production in dendritic cells.
TNFA, TNFSF2, TNF, Tumor necrosis factor, Cachectin, TNF-alpha, Tumor necrosis factor ligand superfamily member 2, TNF-a
The Human TNF alpha ELISA kit is a single-wash 90-min SimpleStep ELISA® used to quantify Human TNF alpha with a sensitivity of 4.32 pg/ml. The assay uses a simple Mix-Wash-Read protocol with just one incubation and wash step.Easy-to-use, rapid, sensitive human TNF alpha sandwich ELISA kit:- Colorimetric assay - 450nm readout; works on any plate reader- Different formats for different needs: 10x96 plates for bulk orders and 384-well for higher throughput- Broad sample compatibility – works with serum, plasma, cell culture supernatants, and other biological fluids
Colorimetric
Cerebral Spinal Fluid, Heparin Plasma, Citrate plasma, Cell culture supernatant, Serum, EDTA Plasma
Sandwich (quantitative)
Human
15.63 - 1000 pg/mL
1h 30m
Microplate
= 4.32 pg/mL
Sample | n | mean | SD | C.V. |
---|---|---|---|---|
Sample PBMC media | n 8 | mean - | SD - | C.V. 2.5 |
Sample | n | mean | SD | C.V. |
---|---|---|---|---|
Sample PBMC media | n 3 | mean - | SD - | C.V. 3.1 |
Sample type | Average % | Range |
---|---|---|
Sample type Cerebral Spinal Fluid | Average % = 83 | Range 80 - 88 % |
Sample type Cell culture supernatant | Average % = 110 | Range 105 - 113 % |
Sample type Serum | Average % = 88 | Range 83 - 93 % |
Sample type EDTA Plasma | Average % = 100 | Range 95 - 104 % |
Sample type Heparin Plasma | Average % = 98 | Range 89 - 110 % |
Sample type Citrate plasma | Average % = 91 | Range 86 - 97 % |
Blue Ice
+4°C
+4°C
+4°C
Human TNF alpha ELISA Kit ab181421 is a rapid single-wash 90-min ELISA kit to measure Human TNF alpha in cerebral spinal fluid (CSF), cell culture supernatant, serum and plasma. This SimpleStep ELISA® sensitivity is 4.32 pg/mL.
How the assay works
SimpleStep ELISA® technology employs capture antibodies conjugated to an affinity tag that is recognized by the monoclonal antibody used to coat our SimpleStep ELISA® plates. This approach to sandwich ELISA allows the formation of the antibody-analyte sandwich complex in a single step, significantly reducing assay time. See the SimpleStep ELISA® protocol summary in the image section for further details.
Assay protocol summary
Human TNF alpha ELISA Kit ab181421 protocol summary:
1. Mix : add samples / standards to the wells together with the capture and detector antibody cocktail. Incubate 1 hour at room temperature
2. Wash
3. Add TMB development solution - incubate for 10 min
4. Add Stop solution
5. Read the results on a plate reader at 450 nm
Alternative assay kits
Need to design your own immunoassay? We offer the antibody pair used in this kit in a BSA and Azide-free format, ready for conjugation:
- Human/Monkey TNF alpha Antibody Pair - BSA and Azide free Human/Monkey TNF alpha Antibody Pair - BSA and Azide free ab241791,
- Anti-TNF alpha antibody [EPR18565-29] - BSA and Azide free (Capture) Anti-TNF alpha antibody [EPR18565-29] - BSA and Azide free (Capture) ab242476,
- Anti-TNF alpha antibody [EPR18565-33] - BSA and Azide free (Detector) Anti-TNF alpha antibody [EPR18565-33] - BSA and Azide free (Detector) ab242978
Getting the best performance
ASSAY SPECIFICITY
This kit recognizes both native and recombinant human TNF alpha protein in serum, plasma, and cell culture supernatant samples only.
CROSS REACTIVITY
Recombinant proteins were prepared at 1000 pg/mL and assayed for cross reactivity. No cross reactivity was found for the following targets: - Human IL-2 - Human IL-4 - Human IL-1 alpha - Human IFN gamma - Human TNF beta - Human TNF R1.
INTERFERENCE
Recombinant Human TNF R1 was prepared at 1000 pg/mL and tested for interference. No interference with was observed.
SPECIES REACTIVITY
This kit recognizes human TNF alpha protein. Other species reactivity was determined by measuring 1000 pg/mL recombinant proteins of various species, interpolating the protein concentrations from the human standard curve, and expressing the interpolated concentrations as a percentage of the protein concentration assayed at the same dilution. Reactive species: Primate
Reactivity < 3% was determined for the following species: Mouse/Rat
CALIBRATION
This immunoassay is calibrated against a highly purified human TNF alpha. The NIBSC/WHO unclassified purified human TNF alpha preparation 12/154 was evaluated in this kit. The dose response curve of the unclassified standard TNF alpha parallels the SimpleStep standard curve. To convert sample values obtained with the SimpleStep Human TNF alpha kit to approximate NIBSC 12/154 International units, use the equation below.
NIBSC (12/154) approximate value (IU/mL) = 0.094 x SimpleStep Human TNF alpha value (pg/mL).
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.
Save your precious samples
Save your precious samples: our 384-well Human TNF alpha ELISA Kit only requires a maxmum of 12.5µL
How other researchers are using this kit
Human TNF alpha ELISA Kit ab181421 has been used to assess inflammation in different physio-pathological contexts such as obesity (1), coronary artery disease (2), and rheumatoid arthritis (3).
References: 1-Shaohong Yu et al. 2023, PMID: 38156278, 2-Yongshun Wang et al. 2023, PMID: 38054817, 3-Yaju Tan et al. 2024, PMID: 37997635
Related and recommended products
Human TNF alpha ELISA kit ab181421 is commonly used to measure TNF alpha as a marker of inflammation.
Other ELISA kits to measure inflammatory markers include:
- Human IL-6 ELISA kit Human IL-6 ELISA Kit ab178013
- Human IL-8 ELISA kit Human IL-8 ELISA Kit ab214030
- Human IL-1 beta ELISA kit Human IL-1 beta ELISA Kit ab214025
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.
This supplementary information is collated from multiple sources and compiled automatically.
Tumor necrosis factor alpha (TNF alpha) also known as TNF-a is a cytokine with a molecular weight of approximately 17 kDa. This protein is expressed mainly by macrophages though other cells like T-cells and natural killer cells also produce it. TNF alpha helps in regulating immune responses and inflammatory reactions. Its expression is important in triggering cell apoptosis and modulating immune cell activities. TNF alpha exists in membrane-bound and soluble forms both having biologic activity.
TNF alpha acts as an important regulator of inflammatory and immune responses. It is not part of a complex but interacts with TNF receptors to activate various signaling cascades. TNF alpha promotes leukocyte recruitment to sites of infection or injury by enhancing adhesion molecule expression on endothelial cells. Additionally it induces the production of other pro-inflammatory cytokines like IL-6 and IL-1 amplifying the immune response. Researchers often measure TNF alpha levels in samples using methods like ELISA or enzyme-linked immunosorbent assay and develop specific antibodies like anti-TNF alpha for experimental and therapeutic purposes.
TNF alpha participates in the NF-kB and MAPK pathways significant for mediating inflammatory and immune responses. In the NF-kB pathway TNF alpha activates transcription factors that control the expression of genes involved in inflammation and cell survival. TNF alpha also interacts with proteins such as the TNF receptor 1 (TNFR1) and receptor-associated factors (TRAFs) further relaying signals within these pathways. There is extensive research employing animal models using mouse TNF-alpha and anti-TNF reagents to dissect these pathways’ intricacies.
TNF alpha substantially affects conditions like rheumatoid arthritis and Crohn's disease both characterized by chronic inflammation. Elevated levels of TNF alpha drive the inflammatory processes central to these diseases leading to tissue damage. Other proteins like IL-6 also play critical roles in these disorders and together with TNF alpha create a pro-inflammatory environment. Therapeutic agents such as anti-TNF treatments aim to mitigate these effects by specifically targeting TNF alpha activity.
We are dedicated to supporting your work with high quality reagents and we are here for you every step of the way should you need us.
In the unlikely event of one of our products not working as expected, you are covered by our product promise.
Full details and terms and conditions can be found here:
Terms & Conditions.
Human TNF alpha standard curve comparison.
Standard Curve comparison between human TNF alpha SimpleStep ELISA kit and traditional ELISA kit from leading competitor. SimpleStep ELISA kit shows increased sensitivity.
Example of human TNF alpha standard curve in Sample Diluent NS.
The TNF alpha standard curve was prepared as described in Section 10. Raw data values are shown in the table. Background-subtracted data values (mean +/- SD) are graphed.
Interpolated concentrations of native TNF alpha in human PBMC cell culture supernatant samples.
PBMC cells were treated with 1.5% PHA-M for 46 hours in 10F RPMI. The concentrations of TNF alpha were measured in duplicates, interpolated from the TNF alpha standard curve and corrected for sample dilution. Undiluted samples are PHA-M treated PBMC cell culture supernatant 10%. The interpolated dilution factor corrected values are plotted (mean +/- SD, n=2). The mean TNF alpha concentration was determined to be 7326.31 pg/mL in PHA-M treated PBMC cell culture supernatant.
Interpolated concentrations of native TNF alpha PBMC cell culture supernatant samples.
PBMC cells were stimulated with 1.5% PHA-M or vehicle control in 10F RPMI and incubated for 46 hours. The concentrations of TNF alpha were measured in duplicate and interpolated from the TNF alpha standard curves. Undiluted samples are PHA-M stimulated PBMC cell culture supernatant 10% and unstimulated PBMC cell culture supernatant 10%. The interpolated dilution factor corrected values are plotted (mean +/- SD, n=2). The mean TNF alpha concentration was back calculated determined to be 7265.71 pg/mL in PHA-M stimulated PBMC cell culture supernatant and undetectable in the unstimulated PBMC control.
Linearity of dilution - native TNF alpha.
Native TNF alpha was measured in biological samples in a 2-fold dilution series. Sample dilutions are made in Sample Diluent NS.
Linearity of dilution - recombinant TNF alpha.
Recombinant TNF alpha was spiked biological samples and diluted in a 2-fold dilution series in Sample Diluent NS. Twenty individual healthy human female/male donors were measured in duplicate for the presence of TNF alpha. All values were below the detectable range of the assay
NL – Non-Linear
Human TNF alpha concentration was interpolated from the standard curve. Supernatants from cell culture samples were serially diluted and assessed by the Human TNF alpha ELISA kit (ab181421). Wild-type THP-1 and TNF alpha knockout THP-1 (Human TNF knockout THP-1 cell line ab273761) cells were assessed in duplicate (n=2). Cells were either treated with 100 ng/ml LPS for 16 h to induce expression of TNF alpha or not treated with LPS. Data are represented as the mean and error bars represent standard deviation."
Raw data value for example of TNF standard curve in Sample Diluent NS.
Raw data values are shown in the table
SimpleStep ELISA technology allows the formation of the antibody-antigen complex in one single step, reducing assay time to 90 minutes. Add samples or standards and antibody mix to wells all at once, incubate, wash, and add your final substrate. See protocol for a detailed step-by-step guide.
Please note: All products are 'FOR RESEARCH USE ONLY. NOT FOR USE IN DIAGNOSTIC OR THERAPEUTIC PROCEDURES'.
For licensing inquiries, please contact partnerships@abcam.com