| Field | Specification |
|---|---|
| Target | |
| Alternative names | Tumor necrosis factor;Cachectin;TNF-alpha;Tumor necrosis factor ligand superfamily member 2;TNF-a;Tumor necrosis factor, membrane form;N-terminal fragment;NTF;Intracellular domain 1;ICD1;Intracellular domain 2;ICD2;C-domain 1;C-domain 2;Tumor necrosis factor, soluble form;TNF;TNFA, TNFSF2; |
| Gene ID | |
| UniProt # | |
| Reactivity | |
| Applications | |
| Immunogen | Expression system for standard: E.coli; Immunogen sequence: V77-L233 |
| Expression system | |
| Expression region | |
| Assay type | |
| Sample type(s) | Cell Culture Supernatant, Serum, Plasma |
| Sensitivity | |
| Detection range | |
| Assay time | |
| Storage | |
| Catalog no. (Mfr.) | |
| Main SKU |
Assay Principle
This sandwich ELISA quantifies human tumor necrosis factor (gene TNF) in cell culture supernatants, serum and plasma (heparin, EDTA, citrate). Strips are pre-coated with a capture antibody. After the sample is added, a biotinylated detection antibody binds the captured protein, and an avidin–biotin–peroxidase complex supplies the enzyme. TMB develops color in proportion to the bound target; the reaction is stopped and read at 450 nm, and concentrations are read from the standard curve.
Tumor necrosis factor (TNF, TNF-α) is a pro-inflammatory cytokine produced mainly by activated macrophages and T cells. Acting through the receptors TNFR1 and TNFR2, it activates NF-κB and MAP kinase pathways and can trigger apoptosis or necroptosis, driving fever and inflammation. TNF is a central target in rheumatoid arthritis and inflammatory bowel disease research and a standard readout in immunology studies.
Kit Components
| Anti-Human TNF Pre-coated 96-well strip microplate | 1 plate (96 wells) |
|---|---|
| Human TNF Standard | 2 vials, 10 ng/tube |
| Human TNF Biotinylated antibody (100x) | 100 µL |
| Avidin-Biotin-Peroxidase Complex (100x) | 100 µL |
| Sample Diluent | 30 mL |
| Antibody Diluent | 12 mL |
| Avidin-Biotin-Peroxidase Diluent | 12 mL |
| Color Developing Reagent (TMB) | 10 mL |
| Stop Solution | 10 mL |
| Wash Buffer (25x) | 20 mL |
| Adhesive plate sealers | 4 |
Also required (not supplied): microplate reader for 450 nm; incubator; automated plate washer (optional); precision pipettes for 0.5 µL through 1 mL volumes of aqueous solutions; multichannel pipettes (recommended for many samples); deionized or distilled water; 500 mL graduated cylinders; test tubes for dilution.
Kit components are not sold separately.
Performance Data
| Species | Human |
|---|---|
| Sample types | Cell culture supernatants, serum and plasma (heparin, EDTA, citrate) |
| Assay range | 15.6 pg/mL–1,000 pg/mL |
| Sensitivity | <1 pg/mL |
| Standard | Recombinant human TNF (Val77–Leu233), expressed in E. coli |
| Cross-reactivity | None detected against related proteins in supplier testing |
| Intra-assay CV | 4.8–7.4% (3 samples, n = 16 each) |
| Inter-assay CV | 5.4–7.4% (3 samples, n = 24 each) |
| Format | 96 wells, removable strips |
Sample dilution: start at 1:1. In the supplier’s tests, serum and plasma samples measured below the lowest standard.
Example standard curve (OD450, pg/mL): 0 = 0.01; 15.6 = 0.092; 31.2 = 0.152; 62.5 = 0.29; 125 = 0.512; 250 = 0.897; 500 = 1.375; 1000 = 2.04. Run a new standard curve on every plate.
Storage & Handling
Store at 4 °C for up to 6 months or −20 °C for up to 12 months. Avoid repeated freeze–thaw cycles. Ships on gel ice and tolerates up to 3 days at room temperature; freeze on arrival.
Safety
Sample data
| Concentration (pg/ml) | 0 | 15.6 | 31.2 | 62.5 | 125 | 250 | 500 | 1000 |
| O.D. | 0.01 | 0.092 | 0.152 | 0.29 | 0.512 | 0.897 | 1.375 | 2.04 |
Intra/inter assay consistency
| Intra-Assay Precision | Inter-Assay Precision | |||||
|---|---|---|---|---|---|---|
| Sample | 1 | 2 | 3 | 1 | 2 | 3 |
| n | 16 | 16 | 16 | 24 | 24 | 24 |
| Mean (pg/ml) | 16 | 82 | 201 | 14 | 75 | 186 |
| Standard deviation | 0.76 | 5.49 | 12.66 | 0.75 | 5.55 | 11.9 |
| CV (%) | 4.8% | 6.7% | 7.4% | 5.4% | 7.4% | 6.4% |
Kit components
Description|Quantity Pre-coated 96-well strip microplate|1 Standard|2 vials Biotinylated antibody (100x)|100ul Avidin-Biotin-Peroxidase Complex (100x)|100ul Sample Diluent|30ml Antibody Diluent|12ml Avidin-Biotin-Peroxidase Diluent|12ml Color Developing Reagent (TMB)|10ml Stop Solution|10ml Wash Buffer (25x)|20ml Adhesive plate sealers|4Materials required but not provided
- Microplate Reader capable of reading absorbance at 450nm.
- Incubator.
- Automated plate washer (optional).
- Pipettes and pipette tips capable of precisely dispensing 0.5 µl through 1 ml volumes of aqueous solutions.
- Multichannel pipettes are recommended for large amount of samples.
- Deionized or distilled water.
- 500ml graduated cylinders.
- Test tubes for dilution.
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