An update on the European Bioanalysis Forum recommendation on singlicate analysis for ligand binding assays:...

Duplicate measurements have long been used in regulated ligand-binding assays, particularly for pharmacokinetic assays. The European Bioanalysis Forum has previously shown that single-well analysis can be suitable for pharmacokinetic endpoints when supported by appropriate method development and validation. This paper extends that evaluation to biomarker and anti-drug antibody assays. A collaborative project involving multiple companies assessed whether single-well measurements provide outcomes comparable to duplicate formats across a wide range of assay types and platforms. The combined data showed strong agreement between both approaches and indicated that duplicate measurement seldom adds scientific value. The European Bioanalysis Forum therefore recommends a data-driven strategy in which replicate number is defined during method development and justified for the intended use of the data. Where performance supports it, single-well analysis should be considered the default approach.