Description
Intended Uses: This C5A ELISA kit is intended Laboratory for research use only.The Stop Solution changes the color from blue to yellow and the intensity of the color is measured at 450 nm using a spectrophotometer. In order to measure the concentration ofC5A in the sample, thisC5A ELISA Kit includes a set of calibration standards. The calibration standards are assayed at the same time as the samples and allow the operator to produce a standard curve of Optical Density versusC5A concentration. The concentration ofC5A in the samples is then determined by comparing the O.D. of the samples to the standard curve.
Principle of the Assay: This C5A enzyme linked immunosorbent assay applies a technique called a quantitative sandwich immunoassay. The microtiter plate provided in this kit has been pre-coated with a monoclonal antibody specific forC5A. Standards or samples are then added to the microtiter plate wells andC5A if present, will bind to the antibody pre-coated wells. In order to quantitatively determine the amount ofC5A present in the sample, a standardized preparation of horseradish peroxidase (HRP)-conjugated polyclonal antibody, specific forC5A are added to each well to "sandwich" theC5A immobilized on the plate. The microtiter plate undergoes incubation, and then the wells are thoroughly washed to remove all unbound components. Next, A and B substrate solution is added to each well. The enzyme (HRP) and substrate are allowed to react over a short incubation period.Only those wells that containC5A and enzyme-conjugated antibody will exhibit a change in colour. The enzyme-substrate reaction is terminated by the addition of a sulphuric acid solution and the colour change is measured spectrophotometrically at a wavelength of 450 nm