Human Chorionic Gonadotropin ELISA - Free Beta-HCG

Enzyme Immunoassay for the Determination of Free Beta-Subunit of Chorionic Gonadotropin (Free Beta-hCG) in Serum

FOR RESEARCH USE ONLY. NOT FOR USE IN DIAGNOSTIC PROCEDURES

SUMMARY
Human chorionic gonadotropin can be used as a tumor marker, as its ? subunit is secreted by some cancers including seminoma, choriocarcinoma, germ cell tumors, hydatidiform mole formation, teratoma with elements of choriocarcinoma, and islet cell tumor. For this reason a positive result in males can be a test for testicular cancer. The normal range for men is between 0-5 mIU/mL. Combined with alpha-fetoprotein, ?-HCG is an excellent tumor marker for the monitoring of germ cell tumors.

PRINCIPLE OF THE TEST
The free Beta-hCG ELISA test is based on the principle of a solid phase enzyme-linked immunosorbent assay.1 The assay system utilizes a unique monoclonal antibody directed against a distinct antigenic determinant on the free Beta-hCG. Mouse monoclonal anti-free-Beta-hCG antibody is used for solid phase immobilization (on the microtiter wells). A goat anti whole hCG antibody is in the antibody-enzyme (horseradish peroxidase) conjugate solution. The test sample is allowed to react sequentially with the two antibodies, resulting in the free Beta-hCG molecules being sandwiched between the solid phase and enzyme-linked antibodies. After two separate 30 minute incubations at 37 °C, the wells are washed with water to remove unbound labeled antibodies. A solution of TMB Reagent is added and incubated for 20 minutes, resulting in the development of a blue color. The color development is stopped with the addition of Stop Solution changing the color to yellow. The concentration of Beta-hCG is directly proportional to the color intensity of the test sample. Absorbance is measured spectrophotometrically at 450 nm.

EXAMPLE OF STANDARD CURVE
Results of a typical standard run with optical density readings at 450 nm shown in the Y axis against Human Chorionic Gonadotropin concentrations shown in the X axis. This standard curve is for the purpose of illustration only, and should not be used to calculate unknowns. Each user should obtain his or her own data and standard curve.