Phosphofructokinase, Liver (PFKL) BioAssay™ ELISA Kit (Human)

The BioAssay™kit is a sandwich enzyme immunoassay for in vitro quantitative measurement of PFKL in in human tissue homogenates, cell lysates and other biological fluids.||Detection Range:|1.56-100ng/ml||Sensitivity:|0.59ng/ml||Specificity:|This assay has high sensitivity and excellent specificity for detection of Phosphofructokinase, Liver (PFKL). No significant cross-reactivity or interference between Phosphofructokinase, Liver (PFKL) and analogues was observed.||Precision:|Intra-Assay %CV: <10%|Inter-Assay %CV: <12%||Test Principle:|The microplate provided in this kit has been pre-coated with an antibody specific to PFKL. Standards or samples are then added to the appropriate microplate wells with a biotin-conjugated antibody specific to PFKL. 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 PFKL, 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 PFKL in the samples is then determined by comparing the O.D. of the samples to the standard curve.||Kit Components:|Pre-coated, ready to use 96-well strip plate|Standard, 2x vials|Detection Reagent A, 1x120ul|Detection Reagent B, 1x120ul|TMB Substrate, 1x9ml|Wash Buffer (30X): 1x20ml|Standard Diluent, 1x20ml|Assay Diluent A, 1x12ml|Assay Diluent B, 1x12ml|Stop Solution, 1x6ml||Storage and Stability:|Store Microtiter Strips, Standard, Detection Reagents and Positive Control at -20°C. After reconstitution of Positive Control, store at 4°C and use within 5 days. Store all the other components at 4°C. Unused kit is stable for 6 months after receipt. Once kit components are opened, it is highly recommended to use remaining reagents within 1 month provided this is within the expiration date of the kit. For maximum recovery of product, centrifuge the original vials after thawing and prior to removing the cap.||Assay Procedure Summary:|1. Prepare all reagents, samples and standards.|2. Add 100ul standard or sample to each well. Incubate 1 hour at 37°C.|3. Aspirate and add 100ul prepared Detection Reagent A. Incubate 1 hour at 37°C.|4. Aspirate and wash 3 times.|5. Add 100ul prepared Detection Reagent B. Incubate 30 minutes at 37°C.|6. Aspirate and wash 5 times.|7. Add 90ul Substrate Solution. Incubate 10-20 minutes at 37°C.|8. Add 50ul Stop Solution. Read at 450nm immediately.