NFkB (Nuclear Factor Kappa B) BioAssay(TM) ELISA Kit (Human)

The NFkB (Nuclear Factor Kappa B) BioAssay(TM) ELISA Kit (Human) is a quantitative sandwich assay for the detection of NFkB in human serum, plasma and other biological fluids. Cell lysates and tissue homogenates, though not tested, may potentially be used as samples. Detection Range: 0.313-20ng/ml, Sensitivity: <0.188ng/ml, Precision: Intra-Assay CV: <8%, Inter-Assay CV: <10%, Assay Principle: The microtiter plate provided in this kit has been pre-coated with an antibody specific to NFkB. Standards and samples are added to the appropriate microtiter plate wells followed by a biotin-conjugated antibody specific to NFkB. Streptavidin conjugated to horseradish peroxidase (HRP) is added to each microplate well and incubated. After TMB substrate solution is added, only those wells that contain NFkB, biotin-conjugated antibody and enzyme-conjugated streptavidin complex will exhibit a change in color. The enzyme-substrate reaction is terminated by the addition of sulfuric acid solution and the color change is measured spectrophotometrically at a wavelength of 450nm ± 10nm. The concentration of NFkB in the sample is then determined by comparing the O.D. of the sample to the standard curve. Kit Components: *357596A: Microtiter Strips, 8x12 wells., *357596B: Standard, 2 vials, 357596C: Sample/Standard Dilution Buffer, 1x20ml, 357596D: Antibody-Biotin (Concentrated), 1x120ul, 357596E: Antibody Dilution Buffer, 1x10ml, *357596F: Streptavidin-HRP Conjugate (SABC), 1x120ul, 357596G: SABC Dilution Buffer, 1x10ml, *357596H: TMB Substrate, 1x10ml, 357596J: Stop Solution, 1x10ml, 357596K: Wash Buffer (25X), 1x30ml, Storage and Stability: Store unopened *357596A at 4ºC, store at -20ºC once opened. Store unopened *357596B at 4°C, once reconstituted store at 4°C for up to 12 hours or at -20°C for up to 48 hours. Store *357596F and *357596H in the dark at 4°C. Store all other components at 4°C. Kit is stable for 6 months after receipt. For maximum recovery of product, centrifuge the original vials after thawing and prior to removing the cap. Assay Summary: 1. Wash plate 2 times before adding standards, samples and control (zero) to wells., 2. Add 100ul standard or sample to each well and incubate for 90 minutes at 37°C , 3. Aspirate the liquid and wash plate 3 times., 3. Add 100ul Antibody-Biotin working solution to each well and incubate for 60 minutes at 37°C, 4. Aspirate the liquid and wash plate 3 times., 5. Add 100ul SABC working solution to each well. Incubate for 30 minutes at 37°C, 6. Aspirate the liquid and wash plate 5 times., 7. Add 90ul TMB substrate. Incubate 15-30 minutes at 37°C, 8. Add 50ul Stop Solution. Read at 450nm immediately., 9. Calculate results.