The CanAg CA19-9 EIA is a solid-phase, non-competitive immunoassay based upon the direct sandwich technique.
Calibrators, controls and patient samples are incubated together with a biotinylated Anti-CA19-9 monoclonal antibody (MAb) C192 (derived from mice) in streptavidin coated microstrips.
CA19-9 present in calibrators or samples is adsorbed to the streptavidin coated micro- strips by the biotinylated Anti-CA19-9 MAb during the incubation.
The strips are then washed and incubated with HRP labeled Anti-CA19-9 MAb C192. After washing, buffered Substrate/Chromogen reagent (hydrogen peroxide and 3, 3’, 5, 5’ tetra-methyl-benzidine) is added to each well and the enzyme reaction is allowed to proceed.
During the enzyme reaction a blue color will develop if anti-gen is present. The intensity of the color is proportional to the amount of CA19-9 present in the samples.The color intensity is determined in a microplate spectrophotometer at 620 nm (or optionally at 405 nm after addition of Stop Solution).
Calibration curves are constructed for each assay by plotting absorbance value versus the concentration for each calibrator. The CA19-9 concentrations of patient samples are then read from the calibration curve.