Abstract
This thesis project aims to develop new analytical methods to characterize vaccine formulations.The first objective was to evaluate the performance of capillary electrophoresis (CE) and Taylor's dispersion analysis (TDA) for the study of antigen-adjuvant interactions in vaccine products. As these interactions can have a significant impact on the immune response, the development of a vaccine candidate involves the precise study of antigen-adjuvant mixtures and their potential interactions. First, the study of the interactions between an anionic polymeric adjuvant (polyacrylic acid, SPA09), and a cationic vaccine antigen under development for the treatment of Staphylococcus aureus was carried out by frontal analysis continuous capillary electrophoresis (FACCE). The FACCE methodology provided direct access to the percentage of free antigens and to the average number of antigens bound per adjuvant. Which allowed to plot the adsorption isotherms and determine the interaction constants. TDA subsequently allowed to go further and determine the size of the antigen-adjuvant complexes under physiological conditions and also to access the interaction constants. The mean hydrodynamic radii (Rh) of the complexes obtained by TDA were confirmed by those measured by transmission electron microscopy (TEM) and the interaction parameters measured by TDA were of the same order of magnitude as those previously obtained by FACCE. Finally, capillary zone electrophoresis (CZE) allowed to study antigen/adjuvant interactions in the context of vaccines containing mixtures of several antigens. The methodology developed by CZE made it possible to separate three strains of inactivated poliovirus, without denaturing them, and to study their interaction with aluminum oxyhydroxide (AlOOH).The second objective of this thesis was to develop new analytical methods for the characterization of messenger RNA (mRNA) vaccine formulations. In this context, a method for analyzing the size and the size distribution of LNPs and mRNA has been developed by TDA. A comparison between the sizes obtained by TDA and by dynamic light scattering (DLS) was carried out and the two methods were found to be complementary. Thus, TDA methodology could be used to study different formulation parameters impacting the size of LNPs. Subsequently, TDA was also used to study the degradation of mRNA in the presence of ribonucleases (RNases) and to determine the encapsulation efficiency of mRNA within the LNPs as well as the size of the encapsulated mRNA. Finally, TDA also provided easy access to the viscosity of the LNP formulations and CE allowed the determination of the surface charge density of the LNPs.