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Monoclonal Anti-Tumor Necrosis Factor-α clone

28401 produced in mouse, purified immunoglobulin Catalog Number T6817 Product Description Monoclonal Anti-Tumor Necrosis Factor-α (TNF-α) (mouse IgG1) is produced in mouse using purified recombinant human tumor necrosis factor-α, expressed in E.

coli, as immunogen. The antibody is purified from ascites fluid using protein G affinity chromatography. Monoclonal Anti-Tumor Necrosis Factor-α recognizes recombinant human tumor necrosis factor by various immunochemical techniques including immunoblotting, immunohistochemistry, neutralization, and capture ELISA. Using the antibody as a capture antibody in human TNF-α sandwich ELISAs, there is less than 0.05% cross-reactivity with recombinant human TNF-β, recombinant mouse TNF-α, recombinant rat TNF-α, recombinant porcine TNF-α, recombinant human TNF RI, recombinant human TNF RII, recombinant mouse TNF RI, and recombinant mouse TNF RII. Tumor necrosis factor-α (TNF-α),1-5 also called cachectin, is a member of the TNF superfamily of cytokines. TNF-α is expressed as a

26 kDa membrane bound protein and is then cleaved by TNF-α converting enzyme (TACE) to release the soluble

17 kDa monomer, which forms homotrimers in circulation. Human and mouse TNF-α share ~79% amino acid sequence identity. TNF-α and the related molecule TNF-β (LT-α) share close structural homology with 28% amino acid sequence identity and both activate the same TNF receptors, TNF RI and TNF RII. Tumor necrosis factor-α plays roles in antitumor activity, immune modulation, inflammation, anorexia, cachexia, septic shock, viral replication, and hematopoiesis. It is expressed by a great variety of cells, with numerous inductive and suppressive agents. Primarily, macrophages produce TNF-α in response to immunological challenges such as bacteria (lipopoly- saccharides), viruses, parasites, mitogens, and other cytokines. Neutrophils, activated lymphocytes, NK cells, LAK cells, astrocytes, endothelial cells, smooth muscle cells, and some transformed cells also produce TNF-α. TNF-α is cytotoxic for many transformed cells (its namesake activity) but in normal diploid cells, it can stimulate proliferation (fibroblasts), differentiation (myeloid cells) or activation (neutrophils).5 TNF-α also shows antiviral effects against both DNA and RNA viruses and induces production of several other cytokines. Reagent Supplied as ~500 ?g of antiserum lyophilized from a 0.2 ?m filtered solution in phosphate buffered saline containing 5% trehalose. Preparation Instructions To one vial of lyophilized powder, add

1 mL of sterile phosphate buffered saline containing 0.1% human serum albumin or bovine serum albumin to produce a 0.5 mg/mL stock solution of antibody. Storage/Stability Prior to reconstitution, store at ?20 °C. Reconstituted product may be stored at 2-8 °C for up to one month. For prolonged storage, freeze in working aliquots at ?20 °C. Avoid repeated freezing and thawing. Procedure Capture ELISA: the antibody can be used as the capture antibody in a human TGF-α ELISA in combination with biotinylated, human TGF-α affinity purified polyclonal detection antibody. Using plates coated with

100 ?L/well of the capture antibody at

4 ?g/mL, in combination with

100 ?L/well of the detection antibody, an ELISA for sample volumes of

100 ?L can be obtained. To arrive at the optimal dose range for this ELISA, a two-fold dilution series of the protein standard starting with

1 ng/mL is suggested. Neutralization: the bioactivity was determined on mouse L929 cells6 using the murine L929 cytotoxicity assay. To measure this activity, recombinant human TNF-α is incubated with various concentrations (0.0001-10 ng/mL) of the antibody in a