Description(s):
Alpha-fetoprotein (AFP) is a plasma protein produced by the liver and the yolk sac during fetal development. AFP is believed to be the fetal analog of serum albumin and it acts as a carrier protein for steroids, bilirubin, fatty acids, and retinoids. In addition to its function as a transport protein, AFP can exert immunosuppressive activity, regulate cell proliferation and apoptosis, initiate intracellular signaling, and contribute to cell invasion. Low levels of AFP in the maternal serum are associated with a higher incidence of Down syndrome, while higher levels of AFP are an indicator of spina bifida and anencephaly. Because of this, maternal AFP serum level is commonly used to screen for Down syndrome and neural tube defects. Furthermore, some tumor types have been found to secrete AFP including hepatoblastoma and hepatocellular carcinoma. Thus, AFP can be used as a tumor marker to monitor a patient’s response to a treatment such as chemotherapy.
Principle
The wells of the plate are coated with capture antibody specific to AFP. In this assay, the test sample initially reacts with the solid phase capture antibody, resulting in AFP being bound to the well. The wells are then washed to remove unbound proteins, and biotin-linked antibodies are added to bind to the immobilized AFP. After washing away the unbound antibodies, Streptavidin-HRP conjugate is added to form a complex with the antibody-bound AFP. After incubation, the wells are washed to remove unbound Streptavidin-HRP conjugate. TMB substrate is then added and forms a blue color when the HRP-linked antibodies are detected. The reaction is then terminated with Stop Solution, which changes the color from blue to yellow. The AFP concentration in each well is directly proportional to its color intensity and can be quantified by measuring its optical density at 450 nm (OD450) in a microplate reader.
