The 3M Allergen Protein ELISA Kits utilize a sandwich ELISA. The target protein present in the sample react with the specific antibodies, which have been adsorbed to the surface of polystyrene microtiter wells. After the removal of unbound proteins by washing, antibodies conjugated with horseradish peroxidase (HRP) are added. This enzyme-labeled antibody forms complexes with the previously bound target protein.
Following a second washing step, the enzyme bound to the immunosorbent is detected by the addition of a chromogenic substrate, 3,3’,5,5’-tetramethylbenzidine (TMB). The color development from this enzymatic reaction varies directly with the concentration of target protein in the sample tested; thus, the absorbance, at 450 nm, is a measure of the concentration of the target protein in the test sample.
The quantity of protein in the test sample can be extrapolated from the standard curve, constructed from standards of known concentration, and adjusted to consider the sample dilution.