Development of a multiplex immunoassay for simultaneous detection of antigens in DTaP vaccines

Routine batch quality testing before vaccine release, notably for potency evaluation, still relies on animal use for several animal and human vaccines. In this context, the VAC2VAC project is a public-private consortium of 22 partners funded by EU. Its main objective is to reduce the number of animal used for batch testing by developing immunoassays that could be implemented for routine potency assessment of vaccines. Our work focused on the development of a Luminex-based multiplex assay (four to six-plex assay depending on the number of pertussis antigens present in the formulation) to monitor the consistency of antigen quantity and quality throughout the production process of DTaP vaccines from two manufacturers (coded HuA and HuB). A first work was carried out through in-depth monoclonal antibody characterization to determine the best antibody pairs to use in the antibody-antigen sandwich approach. Selected antibody pairs were used for development and optimization of the Luminex assay with non-adsorbed antigens and, later, with complete vaccine formulation from HuA and HuB. All selected antibody pairs were tested as capture or detection antibody of the sandwich assay to determine the most favorable combination to use in the multiplex assay (better signal-to-noise ratio). As a proof of concept, the potency of 18 batches from HuB was compared using the multiplex assay.