Immunoassays performed on chemistry instruments housed in the laboratory use antibodies to recognize and bind to specific kinds of drug molecules in a urine sample. A measurable signal is then produced in response to the binding of the drug (the drug serving as the antigen in the immunoassay). Detection of the signal is accomplished with the use of enzyme labels, along with their respective substrates. Examples of enzyme labels include glucose-6-phosphate dehydrogenase (G6PD), horseradish peroxidase, alkaline peroxidase, β-galactosidase, and luciferase. The enzyme labels can generate several signals depending on the substrate in which they are present. They can emit radiation, produce a color change, fluoresce under light, or emit light.
A known amount of antibody, labeled reagent drug, and substrate is added to a urine sample in a competitive immunoassay. The drug (or drug metabolite) in the sample competes with the labeled reagent drug for binding sites on the antibody to form antigen-antibody complexes. If no drug exists in the urine sample, the antibody binding sites are available to bind the labeled drug or metabolite, blocking the substrate from reacting with the enzyme. If the drug is present in the sample, it will bind to the antibody binding sites, leaving the labeled drug or metabolite available to react with the substrate and illicit a response measured as a change of absorbance by a spectrometer, fluorometer, or luminometer.
