Routine batch quality testing for vaccine release, especially for potency determination, frequently involves the use of animals for both veterinary and human vaccines. This presentation gives an overview of our work during the VAC2VAC project, a collaboration between 23 partners funded by the EU, which aimed to minimize animal use in batch testing by developing immunoassays for potency evaluation of vaccines. Our research concentrated on creating a Luminex®-based multiplex immunoassay to monitor the consistency of antigen quantity and quality during the production of DTaP (Diphtheria, Tetanus, acellular Pertussis) vaccines. We employed well-defined monoclonal antibody pairs for the development and fine-tuning of the Luminex® assay, using both non-adsorbed and adsorbed antigens, as well as complete vaccine formulations from two manufacturers. The multiplex immunoassay showed high specificity, reproducibility, no cross-reactivity, and successfully identified qualitative and quantitative changes in formulations that were either overdosed, underdosed, or degraded by heat or H O . Moreover, the assessment of 11 primer vaccines from manufacturer A and 29 primer plus 28 booster vaccines from manufacturer B indicated acceptable batch-to-batch variability (CI: 82-122%) compared to a reference homologous batch, with the exception of the pertussis toxoid from manufacturer B. The findings of this study support the potential application of the immunoassays as an effective tool for DTaP vaccine quality control in the future.