What Is the Difference Between Direct and Indirect ELISA?

The difference between direct and indirect ELISA is that indirect ELISA requires the addition of a secondary antibody, while direct ELISA only uses a primary antibody. Indirect ELISA is considered to be more sensitive when detecting substances, but direct ELISA is a quicker, cheaper process that calls for less steps.

Direct ELISA, or enzyme-linked immunosorbent assay, begins with the absorption of an antigen onto a plate. A protein is then added to block all other possible binding sites. The first antibody is added, and it binds to any recognized antigen epitopes. Indirect ELISA continues the process by adding a second antibody, which binds to the first antibody. The first antibody is available only if it is bound to the antigen.

Both processes require numerous laboratory washes to remove additional substances. The ELISA process is important in the lab because it allows scientists to identify antigens. It is a popular, reliable way to test for diseases and food allergens.

Indirect ELISA is able to detect smaller amounts of an antigen, and it permits a single secondary antibody to be used with a plethora of different primary antibodies. Direct ELISA requires higher antigen amounts to make a detection, and is not as flexible, because only one antibody is used.