Evaluation of mRNA Transfection Reagents for mRNA Delivery and Vaccine Efficacy via Intramuscular Injection in Mice

The selection of an effective delivery carrier is crucial to assessing mRNA-based vaccines and therapeutics in vivo. Although lipid nanoparticles (LNPs) are commonly used for mRNA delivery, the LNP-mRNA formulation process is laborious and time-consuming and requires a high-cost microfluidic device. Instead, mixing with commercial reagents may simplify mRNA transfection into cells. However, their potential as in vivo carriers in intramuscular vaccination in mouse models remains unclear. In this study, we used three types of commercial RNA transfection reagents, MessengerMAX (MAX; liposome), TransIT-mRNA (IT; cationic polymer), and Invivofectamine (IVF; LNP), to produce nanoparticles directly by pipetting. The particle characteristics and mRNA delivery efficacy of the mRNA-transfection reagent mixtures were analyzed. Additionally, immune responses to vaccine efficacy and protective immunity of the mRNA mixtures as vaccine antigens were evaluated in a mouse model. Although MAX and IT showed high in vitro transfection efficiencies, their in vivo performances were limited. In contrast, IVF exhibited notable particle stability and homogeneity, making it a promising delivery carrier. Intramuscular IVF injection significantly enhanced both innate and adaptive immune responses with a robust systemic protein expression. Notably, when using SARS-CoV-2 Spike mRNA, IVF showed robust humoral immune responses, including production of IgG and neutralizing antibodies, thereby resulting in complete protection against SARS-CoV-2 infection. Therefore, these findings position IVF as an accessible and efficient mRNA carrier for evaluating mRNA vaccines and therapeutic efficacy in basic research.