TNF High Sensitivity ELISA Kit (Human)

Intra-assay Precision (Precision within an assay) : 3 samples with low, middle and high level High Sensitive Tumor Necrosis Factor Alpha (TNFa) were tested 20 times on one plate, respectively. Inter-assay Precision (Precision between assays) : 3 samples with low, middle and high level High Sensitive Tumor Necrosis Factor Alpha (TNFa) were tested on 3 different plates, 8 replicates in each plate. CV (%) = SD/meanX100 Intra-Assay: CV<10% Inter-Assay: CV<12%

APC1 protein; Cachectin; DIF; TNF, macrophage-derived; TNF, monocyte-derived; TNF-a; TNFA; TNF-alpha; TNFSF2; TNLG1F; tumor necrosis factor; tumor necrosis factor ligand 1F; tumor necrosis factor ligand superfamily member 2; tumor necrosis factor-alpha.

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. Upregulates 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:22517918) .

The test principle applied in this kit is Sandwich enzyme immunoassay. The microtiter plate provided in this kit has been pre-coated with an antibody specific to High Sensitive Tumor Necrosis Factor Alpha (TNFa) . Standards or samples are then added to the appropriate microtiter plate wells with a biotin-conjugated antibody specific to High Sensitive Tumor Necrosis Factor Alpha (TNFa) . Next, Avidin conjugated to Horseradish Peroxidase (HRP) is added to each microplate well and incubated. After TMB substrate solution is added, only those wells that contain High Sensitive Tumor Necrosis Factor Alpha (TNFa), biotin-conjugated antibody and enzyme-conjugated Avidin will exhibit a change in color. The enzyme-substrate reaction is terminated by the addition of sulphuric acid solution and the color change is measured spectrophotometrically at a wavelength of 450nm +/- 10nm. The concentration of High Sensitive Tumor Necrosis Factor Alpha (TNFa) in the samples is then determined by comparing the O.D. of the samples to the standard curve.

Reconstitution & Storage Instructions Western Blotting/Immunoblotting (WB/IB) Protocol Immunohistochemistry (IHC) Protocol Immunocytochemistry (ICC) Protocol Enzyme-Linked ImmunoSorbent Assay (ELISA) Protocol Blocking Peptide Competition Protocol (BPCP) Immunoprecipitation (IP) Protocol Antibody Array (AA) Protocol Reconstitution & Storage Instructions Reconstitution & Storage Instructions Western Blotting/Immunoblotting (WB/IB) Protocol Western Blotting/Immunoblotting (WB/IB) Protocol Immunohistochemistry (IHC) Protocol Immunohistochemistry (IHC) Protocol Immunocytochemistry (ICC) Protocol Immunocytochemistry (ICC) Protocol Enzyme-Linked ImmunoSorbent Assay (ELISA) Protocol Enzyme-Linked ImmunoSorbent Assay (ELISA) Protocol Blocking Peptide Competition Protocol (BPCP) Blocking Peptide Competition Protocol (BPCP) Immunoprecipitation (IP) Protocol Immunoprecipitation (IP) Protocol Antibody Array (AA) Protocol Antibody Array (AA) Protocol

Matrix Recovery range (%) Average (%) Serum (n=5) 88-96 91 EDTA Plasma (n=5) 82-95 88 Heparin Plasma (n=5) 80-101 86 Matrix Recovery range (%) Average (%) Matrix Recovery range (%) Average (%) Serum (n=5) 88-96 91 Serum (n=5) 88-96 91 EDTA Plasma (n=5) 82-95 88 EDTA Plasma (n=5) 82-95 88 Heparin Plasma (n=5) 80-101 86 Heparin Plasma (n=5) 80-101 86

Component Amount Anti-TNF Microplate 96 Wells (12 x 8 Well Strips) TNF Lyophilized Standard 2 100X Biotinylated TNF Detector Antibody 120 uL Avidin/HRP Conjugate 120 uL Standard Diluent 1 x 20 mL Detector Antibody Diluent 1 x 12 mL Conjugate Diluent 1 x 12 mL 30X Wash Buffer 1 x 20 mL TMB Substrate 1 x 9 mL Stop Solution 1 x 6 mL Component Amount Component Amount Anti-TNF Microplate 96 Wells (12 x 8 Well Strips) Anti-TNF Microplate 96 Wells (12 x 8 Well Strips) TNF Lyophilized Standard 2 TNF Lyophilized Standard 2 100X Biotinylated TNF Detector Antibody 120 uL 100X Biotinylated TNF Detector Antibody 120 uL Avidin/HRP Conjugate 120 uL Avidin/HRP Conjugate 120 uL Standard Diluent 1 x 20 mL Standard Diluent 1 x 20 mL Detector Antibody Diluent 1 x 12 mL Detector Antibody Diluent 1 x 12 mL Conjugate Diluent 1 x 12 mL Conjugate Diluent 1 x 12 mL 30X Wash Buffer 1 x 20 mL 30X Wash Buffer 1 x 20 mL TMB Substrate 1 x 9 mL TMB Substrate 1 x 9 mL Stop Solution 1 x 6 mL Stop Solution 1 x 6 mL

This assay has high sensitivity and excellent specificity for detection of Tumor Necrosis Factor Alpha (TNFa) . No significant cross-reactivity or interference between Tumor Necrosis Factor Alpha (TNFa) and analogues was observed.