The CanAg ProGRP EIA is a solid-phase, one-step, non-competitive immunoassay based on antibodies specific for different epitopes specifically expressed in ProGRP.
Calibrators, controls or unknown samples are incubated together with affinity purified biotinylated Anti-ProGRP polyclonal antibody and horseradish peroxidase (HRP) labelled Anti-ProGRP Monoclonal antibody E146 in Streptavidin coated microtiter strips.
After washing, buffered Substrate/Chromogen reagent (hydrogen peroxide and 3, 3', 5, 5' tetra-methylbenzidine) is added to each well and the enzyme reaction is allowed to proceed.
During the enzyme reaction a blue colour will develop if antigen is present. The intensity of the colour is proportional to the amount of ProGRP present in the samples. The colour intensity is determined in a microplate spectrophotometer at 450 nm after addition of Stop Solution.
Calibration curves are constructed for each assay by plotting absorbance value versus the concentration for each calibrator. The ProGRP concentrations of unknown samples are then read from the calibration curve.