TNF alpha BioAssay(TM) ELISA Kit, Human

Tumor necrosis factor-alpha (TNF-alpha, or TNF) is secreted by macrophages in response to inflammation, infection and cancer (1). Human Tumor Necrosis Factor (TNF) and Lymphotoxin (TNF-beta) are cytotoxic proteins which have similar biological activities and share 30% amino acid homology (2). TNF-alpha is produced by monocytes, which can stimulate endothelial cells to produce the multilineage growth factor granulocyte-macrophage colony-stimulating factor and extend the role of this immunoregulatory protein to the regulation of hematopoiesis in vitro (3). TNF is a soluble protein that causes damage to tumor cells but has no effect on normal cells. Human TNF has been purified to apparent homogeneity as a 17.3kD protein from HL-60 leukemia cells and has showed cytotoxic and cytostatic activities against various human tumor cell lines. The human TNF cDNA is 1585 base pairs in length and encodes a protein of 233 amino acids. The mature protein begins at residue 77, leaving a long leader sequence of 76 amino acids (4). TNF-alpha has been mapped to human chromosome 6 (2). Intended Use: TNF alpha BioAssay(TM) ELISA Kit, Human is a sandwich ELISA for quantitative detection of human TNF alpha in cell culture supernates, serum and plasma (heparin, EDTA, citrate). Range: 15.6-1000pg/ml Cell culture supernates, 7.8-500pg/ml Serum, plasma, Sensitivity: < 1pg/ml, Specificity: Natural and recombinant human TNFa, Crossreactivity: No detectable crossreactivity with other relevant proteins. Kit Components: 144065A: Microtiter Plate, 1x96 wells, *144065B: TNF alpha Standard, 2x1 vial, 144065C: Anti-human TNF alpha (Biotin), 1x130ul, USB Cat No Kit Component 143696: Avidin-Biotin-Peroxidase Complex (ABC), 1x130ul (1:100 dilution), 143697: Sample Diluent Buffer, 1x30ml, 143698: Antibody Diluent Buffer, 1x12ml, 143699: ABC Diluent Buffer, 1x12ml, 143700: TMB Color Developing Agent, 1x10ml, 143701: TMB Stop Solution, 2N H2SO4, 1x10ml, Assay Summary: 1. Add samples and standards and incubate the plate at 37°C for 90 minutes. Do not wash. , 2. Add biotinylated antibodies and incubate the plate at 37°C for 60 minutes. Wash plate 3 times with 0.01M TBS., 3. Add ABC working solution and incubate the plate at 37°C for 30 minutes. Wash plate 5times with 0.01M TBS., 4. Add TMB color developing agent and incubate the plate at 37°C in dark for 25-30 minutes., 5. Add TMB stop solution and read.. Storage and Stability: Store *144065B powder at 4°C. Once reconstituted store at 4°C for up to 12 hours or at -20°C for up to 48 hours. Store other components at 4°C. Stable for 6 months after receipt. For maximum recovery of product, centrifuge the original vial after thawing and prior to removing the cap.