Each test kit contains antibody-coated wells with antibodies specific to the kit’s target substance.

First, samples and controls are added to their respective wells. During an incubation, the target substance binds to the antibodies. The wells are washed to remove all unbound materials.

An enzyme conjugate (antibody chemically linked with an enzyme) is added to all wells. During an incubation, conjugate binds to the already bound target substance, forming a “sandwich.” The more target substance in the wells, the more conjugate that binds in the wells.

The wells are washed again to remove all unbound conjugate.

A substrate, which changes color in the presence of the conjugate, is added to the wells. During an incubation, color develops in proportion to the amount of conjugate in the wells. The more conjugate, the more color, and the more target substance that is detected.