F8 (Coagulation Factor VIII) BioAssay(TM) ELISA Kit (Human)

The F8 (Coagulation Factor VIII) BioAssay(TM) ELISA Kit (Human) is a quantitative sandwich assay for the detection of F8 in human plasma. 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 F8. Standards and samples are added to the appropriate microtiter plate wells followed by a biotin-conjugated antibody specific to F8. 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 F8, 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 F8 in the sample is then determined by comparing the O.D. of the sample to the standard curve. Kit Components: 355545A: Microtiter Strips, 8x12 wells., 355545B: Standard, 2 vials, 355545C: Sample/Standard Dilution Buffer, 1x20ml, 355545D: Antibody-Biotin (Concentrated), 1x120ul, 355545E: Antibody Dilution Buffer, 1x10ml, 355545F: Streptavidin-HRP Conjugate (SABC), 1x120ul, 355545G: SABC Dilution Buffer, 1x10ml, 355545H: TMB Substrate, 1x10ml, 355545J: Stop Solution, 1x10ml, 355545K: Wash Buffer (25X), 1x30ml, Storage and Stability: Store unopened 355545A at 4ºC, store at -20ºC once opened. Store unopened 355545B at 4°C, once reconstituted store at 4°C for up to 12 hours or at -20°C for up to 48 hours. Store 355545F and 355545H protected from light 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 2 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.