Multiple antibodies for RNA:DNA hybrids conjugated with alkaline phosphatase are added. These labeled antibodies attach to the hybrids and form sandwiches. The three components of each sandwich are: capture antibody: hybrid : labeled antibodies.
The microplate is washed to remove any unbound alkaline phosphatase-labeled antibodies. Since multiple labeled antibodies attach to each hybrid, the signal is amplified 3000 fold.
