Critical considerations in the immunochemical detection and quantitation of antigenic biomarkers

The formation of covalent adducts as a result of the interaction of metabolically activated chemicals with host macromolecules is a common critical event in mutagenic, carcinogenic, and immunologic phenomena. Because of their antigenicity and their immunogenicity, covalent adducts may be detected using sensitive immunochemical techniques. The immunochemical approaches to biomonitoring and molecular dosimetry of DNA damage are particularly attractive because they allow sensitive quantitation of specific DNA adducts present in small samples and do not rely on the use of radiolabeled adducts. Two examples of biomarker immunoassay development are presented: an avidin/biotin-amplified ELISA for the major DNA adduct of the human bladder carcinogen 4-aminobiphenyl (ABP), and a particle concentration fluorescent immunoassay (PCFIA) for the major protein adduct associated with toxicity by the prototype hepatotoxin acetaminophen. The examples illustrate critical steps in the development of biomarker immunoassays which include selection of the relevant adduct, preparation of an appropriate immunogen, immunization, characterization of antisera, and development of application-specific sample processing techniques for biomarker quantitation. Immunochemical procedures may be combined with other analytical techniques to form hybrid systems which take advantage of both the antigenicity and the physical or chemical properties of a biomarker to achieve greater specificity and/or sensitivity. The future usefulness of these new tools of molecular epidemiology will depend on a compound-by-compound validation of methods and critical evaluation of the biologic importance of the particular antigenic biomarker as an indicator of exposure and as an indicator of risk.