Immunoradiometric assay for quantitative determination of Chorionic Gonadotropin (hCG) in human serum or plasma
Human Chorionic Gonadotropin (hCG) is a glycoprotein (molecular weight: 38000 Dalton) with approximately 30% carbohydrate content and 9% of sialic acid; it is secreted during pregnancy, by syncytiotrophoblast cells of the placenta. The molecule is composed of two subunits known as αand β. The α-subunit shares structural homology with pituitary glycoprotein hormones (FSH, LH and TSH). The β-subunit is characterized by a sequence of 30 amino acids at the C-terminal, which are specific for each hormone. In this way, unique biological as well as immunological activity is conferred to each hormone. Thus, a specific hCG assay is only possible by means of antibodies directed against the β-subunit. The most important function of hCG is to sustain the corpus luteum in producing Estradiol and Progesterone, thereby supporting pregnancy, in the male fetus in addition, it stimulates Leydig cells to produce Testosterone, thus contributing to sexual differentiation.
The present method is based upon two anti-βhCG monoclonal antibodies recognizing two different epitopes of the molecule. One antibody is adsorbed to the solid phase (coated tube), the other-labeled with Iodine-125 is used as tracer, and the sample to be tested is first incubated in the coated tube. Following this incubation and after an aspirating / washing cycle, the labeled antibody is added to each coated tube, where it is bound to the solid phase by means of the antigen in the standards and (if present) in samples. The amount of binding will be directly proportional to the antigen concentration. After tracer incubation, another washing / aspirating cycle will remove any liquid reagent residue. The radioactivity in the tubes is measured in a gamma counter.