human TNF-a ELISA kit (96 Wells)

The prototype ligand of the TNF superfamily, TNF-?/TNFSF1A, is a pleiotropic cytokine that plays a central role in inflammation and apoptosis (1-4). Human cells known to express TNF-? include B cells, colonic columnar epithelial cells, NK and CD3+CD56+ hepatic natural T cells, macrophages, monocytes and monocyte-derived dendritic cells, CD4+ and CD8+ T cells, mast cells, neutrophils, keratinocytes, plasma cells, and adipocytes.
It is synthesized as a 26 kDa, type II transmembrane protein that is 233 amino acids (aa) in length (4, 5). It contains a 30 aa cytoplasmic domain, a 26 aa transmembrane segment, and a 177 aa extracellular region (6, 7). TNF-? is assembled intracellularly to form a transmembrane, non-covalently-linked homotrimeric protein. The 157 aa residue soluble form of TNF-? (sTNF-? is released from the C-terminus of the transmembrane protein through the activity of TNF-?-converting enzyme (TACE), a membrane-bound disintegrin metalloproteinase (8, 9).
TNF-? is reported to promote inflammatory cell infiltration by upregulating leukocyte adhesion molecules on endothelial cells, serve as a chemotactic agent for monocytes, and activate phagocyte killing mechanisms (10). Deficiencies in either TNF-? or its receptors can increase susceptibility to infection by intracellular pathogens (11 - 12). TNF- may also play a role in lymphoid tissue development. Knockout mice lack splenic B cell follicles and the ability to form germinal centers (13, 14). Other potential physiological roles for TNF-? and its receptors include regulating the differentiation of hematopoietic stem and progenitor cells (15 - 17).
TNF-? has been implicated in a number of pathophysiological processes. It is associated with unregulated pro-inflammatory activity and is thought to be a critical mediator of endotoxin-induced septic shock (18). Cachexia (or whole body wasting) has also been associated with long-term circulating TNF-?. Other disorders with potential TNF-? involvement include asthma), type 2 diabetes, CrohnпїЅs disease, and rheumatoid arthritis.

For the quantitative determination of human tumor necrosis factor(TNF-?) concentrations in cell culture supernates, serum, and plasma.

Principle of the assay: This assay employs the quantitative sandwich enzyme immunoassay technique. A monoclonal antibody specific for TNF-? has been pre-coated onto a microplate. Standards and samples are pipetted into the wells and any TNF-? present is bound by the immobilized antibody. Following incubation unbound samples are removed during a wash step, and then a detection antibody specific for TNF-? is added to the wells and binds to the combination of capture antibody- TNF-? in sample. Following a wash to remove any unbound combination, and enzyme conjugate is added to the wells. Following incubation and wash steps a substrate is added. A coloured product is formed in proportion to the amount of TNF-? present in the sample. The reaction is terminated by addition of acid and absorbance is measured at 450nm. A standard curve is prepared from seven TNF-? standard dilutions and TNF-? sample concentration determined.

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