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Third generation Enzyme Immunoassay for the determination of Hepatitis B surface Antigen or HBsAg in human serum and plasma
A mouse monoclonal antibody specific for HBsAg is fixed to the surface of microwells. Patient’s serum/plasma is added to the microwell together with a second mouse monoclonal antibody, conjugated with Horseradish Peroxidase (HRP) and directed against a different epitope.
The specific immunocomplex, formed in the presence of HBsAg in the sample, is captured by the solid phase. At the end of the one-step incubation, microwells are washed to remove unbound serum proteins and HRP conjugate.
The chromogen/substrate is then added and, in the presence of captured HBsAg immunocomplex, the colorless substrate is hydrolyzed by the bound HRP conjugate to a colored end-product. After blocking the enzymatic reaction, its optical density is measured by an ELISA reader. The color intensity is proportional to the amount of HBsAg present in the sample.
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Hepatitis B 
