How do primary and secondary antibodies work in immunoassays?

Immunoassays such as Enzyme-linked immunosorbent assay (ELISA) and Western blotting (WB) use antibodies to detect proteins and measure their concentration in a sample. Antibodies also facilitate the study of protein localizations and function. In direct immunoassays, a primary antibody recognizes and binds to an antigen. The interaction between antibody and antigen triggers a signal molecule, which enables detection. In indirect immunoassays, a secondary antibody recognizes and binds to a primary antibody already attached to an antigen. This complex interaction of the antigen, primary antibody and secondary antibody triggers the signal molecules, enabling detection.