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 <i>in vivo</i>. 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 <i>in vivo</i> 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 <i>in vitro</i> transfection efficiencies, their <i>in vivo</i> 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.