Chapter 2 - Immunoassay design and issues of interferences

Abstract

Currently, hundreds of immunoassays are commercially available and routinely used in clinical laboratory analysis. They help in the diagnosis and monitoring of various diseases, including allergy, anemia, autoimmune, cardiovascular, diabetes, metabolic, endocrine, and cancer. Immunoassays could be broadly classified as competitive immunoassay (for small molecules (where a single antibody against the analyte is used or sandwich immunoassay (for large molecules) where two specific antibodies targeting different parts of the analyte molecule are used. In addition, immunoassay format can be either homogenous where no separation between bound and free analyte is needed before measuring the signal or heterogenous format where bound form is separated from the free from before measuring the analyte signal. Various commercial assay platforms include cloned enzyme donor immunoassay (CEDIA), enzyme multiplied immunoassay technique (EMIT), luminescent oxygen channeling immunoassay (LOCI), kinetic interaction of microparticles in solution (KIMS), etc. In addition, issues of monoclonal verses polyclonal antibody have also been addressed. In general, monoclonal antibody-based immunoassays are more specific for analytes compared to polyclonal antibody-based assays. However, immunoassays also suffer from interferences from both structurally related and unrelated molecules. Interferences of drug metabolite with assay intended for parent drug are a common problem. Moreover, other sources of interference such as heterophilic antibody and rheumatoid factors mostly in sandwich immunoassays are also discussed.