The hCG Quantitative Test is based on a solid phase enzyme-linked immunosorbent assays (ELISA). The assay system utilizes a mouse monoclonal anti-alpha-hCG antibody for solid phase (microtiter wells) immobilization and a mouse monoclonal anti-β-hCG antibody in the antibody-enzyme (horseradish peroxidase) conjugate solution. The test specimen (serum) is added to the alpha-hCG antibody coated microtiter wells and incubated with the Zero Buffer at room temperature for 30 minutes. If hCG is present in the specimen, it will combine with the antibody on the wells. The wells are then washed to remove any residual test specimen, and β-hCG monoclonal antibody labeled with horseradish peroxidase (conjugate) is added. The conjugate will bind immunologically to the hCG on the wells, resulting in the hCG molecules being sandwiched between the solid phase and enzyme-linked antibodies. After incubation at room temperature for 15 minutes, the wells are washed with water to remove unbound-labeled antibodies. A solution of TMB Reagent is added and incubated at room temperature for 20 minutes, resulting in the development of a blue color. The color development is stopped with the addition of Stop Solution, and the color is changed to yellow and measured spectrophotometrically at 450 nm
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