Evaluating the impact of matrix effects on biomarker assay sensitivity

Abstract

Background: The accuracy of highly sensitive biomarker methods is often confounded by the presence of various circulating endogenous factors in samples causing matrix effects. Method: This article outlines two different biomarker methods: hepcidin enzyme-linked immunosorbent assay (ELISA) for which an orthogonal assessment of ELISA to liquid chromatography–tandem mass spectrometry was performed to examine the potential matrix effect, and sclerostin ELISA to evaluate the matrix effect. Results: Although the potential interfering effects of the endogenous hepcidin variants (prohepcidin and clipped) showed that these proteins had >30% immunoreactivity in ELISA, the hepcidin ELISA preferentially measures full-length hepcidin when the molar ratios of full-length to variants remain >1. The correlation of ELISA to liquid chromatography–tandem mass spectrometry results showed full-length hepcidin as the major form in diseased populations. Conclusion: A fit-for-for-purpose assessment of matrix effect/selectivity was also performed for each method. This article demonstrates the utility of a fit-for-purpose approach to assess the validity of biomarker methods in evaluating the interconnected parameters of matrix effects, sensitivity and selectivity.