Enzyme-linked Immunosorbent Assays (ELISAs) combine the specificity of antibodies with the sensitivity of simple enzyme assays, by using antibodies or antigens coupled to an easily-assayed enzyme. ELISAs can provide a useful measurement of antigen or antibody concentration. There are two main variations on this method: The ELISA can be used to detect the presence of antigens that are recognized by an antibody or it can be used to test for antibodies that recognize an antigen.
A general ELISA is a five-step procedure:
1) coat the microtiter plate wells with antigen;
2) block all unbound sites to prevent false positive results;
3) add primary antibody (e.g. rabbit monoclonal antibody) to the wells;
4) add secondary antibody conjugated to an enzyme (e.g. anti-mouse IgG);
5) reaction of a substrate with the enzyme to produce a colored product, thus indicating a positive reaction.