Question

In the ELISA procedure, first a protein sample is immobilized to a plastic well. Then, the antibody to the protein of interest is added, incubated, and the excess is washed out. Next, the detection antibody (which binds to the primary antibody) is added, incubated, and the excess is washed out. The last step is to add the detection dye (which binds to the detection antibody) to the wells and allow it to incubate. What do you predict the result would look like if you added the detection antibody first, then the antibody to the protein of interest

Answer 1

In this ELISA procedure, a sample of protein is immobilized in the plastic well and then antibody for this protein added and incubated, and cleared out. In the given setting detection, the antibody is conjugated with the protein of interest and if the substrate molecule added to the reaction it will produce a colored product by reacting with the enzyme.

No reaction will take place in the well in case of not adding secondary antibody or detection antibody to the reaction and no colored product will be received.