Affinity and Matrix Effects in Measuring Fish Plasma Vitellogenin Using Immunosorbent Assays: Considerations for Aquatic Toxicologists

Enzyme-linked immunosorbent assays (ELISAs) are important tools in aquatic toxicology and have become crucial in assessing exposure concentrations in the aquatic environment and acute physiological responses in exposed organisms. These assays utilize the inherent properties of antibodies to recognize and selectively bind a target molecule, while largely ignoring other molecules to provide semiquantitative values. A variety of methodologies to measure plasma vitellogenin using ELISAs have generated widely divergent data. Limitations of the ELISA method are known in the wider immunology field, though aquatic toxicologists may be less familiar with these limitations. We evaluated several mechanisms contributing to the divergent vitellogenin data in the literature. Antibody affinities and the matrix in which standard curves are constructed are possible error generators. These errors can be amplified by large sample dilutions necessary to fall within the standard curve. It is important for the aquatic toxicology research community to realize the limitations and understand the pitfalls of absolute plasma vitellogenin data in their studies.