ELISA

1) One capture antibody from the coated microwell
2) The standards and the blood plasma sample is added and any target proteins present binds to the bound antibody
3) After the well is washed, a detecting mix is added, which includes biotin-conjugated detection antibodies and Streptavidin-HRP. The biotinylated antibodies are specific for a different epitope of the 8 antigens and will bind to any target proteins bound in the well (the target protein is “sandwiched” between the two antibodies).
4) Streptavidin-HRP will bind as well, as streptavidin binds with high affinity to biotin.
5) The chemiluminescent substrate is added which reacts with HRP to produce a signal, that is proportionate to the amount of protein bound in the well.