Apolipoprotein H (Apo-H) ELISA

The principle of the double antibody sandwich ELISA is represented in Figure 1. In this assay the apolipoprotein H (Apo-H) present in samples reacts with the anti-Apo-H antibodies, which have been adsorbed to the surface of polystyrene microtiter wells. After the removal of unbound proteins by washing, anti-Apo-H antibodies conjugated with horseradish peroxidase (HRP) are added. These enzyme-labeled antibodies form complexes with the previously bound Apo-H. Following another washing step, the enzyme bound to the immunosorbent is assayed by the addition of a chromogenic substrate, 3,3’,5,5’-tetramethylbenzidine (TMB). The quantity of bound enzyme varies directly with the concentration of Apo-H in the sample tested. Thus, the absorbance, at 450 nm, is a measure of the concentration of Apo-H in the test sample. The quantity of Apo-H in the test sample can be interpolated from the standard curve constructed from the standards and corrected for sample dilution.