HDX-MS Epitope Characterization for Evaluating Antibody Suitability for ELISA-based In Vitro Potency Testing of Vaccines

Abstract

Many current vaccine potency test methods relying on animal testing methods are aimed to be replaced by alternative methods, as per the 3Rs initiative. In vitro potency testing by sandwich ELISA has been implemented for several vaccines antigens, however, antibodies used as reagents in these assays must be carefully selected in order to ensure specificity and complementarity. Selected antibodies are further characterized to ensure that they also provide information on stability of the antigen as well as immunological activity (e.g. neutralization, binding inhibition, etc.). This study employs hydrogen deuterium exchange-mass spectrometry (HDX-MS) based epitope screening on antibody pairs to characterize that their target epitopes are spatially distinct, conformational in nature, and potentially located within immunogenic regions of the antigen. The epitope insights obtained from HDX-MS characterization are proposed as a useful tool to inform antibody selection for in vitro quantitation applications. Antibody pairs targeting epitopes suitable for in vitro quantitation of pertussis toxin (PTx), filamentous haemagglutinin (FHA) and tetanus toxoid (TTd) through sandwich ELISA are described.