The efficacy of RNA therapeutics depends on multiple factors, including delivery efficiency, RNA design, and innate immune responses. Delivery efficiency determines how effectively RNA reaches target cells. RNA design (sequence, structure, and chemical modifications) influences RNA stability and translation. Innate immune responses can be triggered by the RNA itself or delivery vehicles. In vaccines, innate immune activation can be advantageous for promoting immunogenicity, but in other contexts, it may compromise efficacy through inflammation, reduced transgene expression, or cell death. Optimizing these factors is critical for achieving robust transgene expression and therapeutic success.

This Methods Collection focuses on experimental strategies to measure, optimize, and modulate delivery, RNA design, and innate immune responses in RNA therapeutics. Topics of interest include protocols for RNA and lipid nanoparticle quality control, transfection and RNA sequence optimization, and bioassays to assess and modulate innate immune activation and transgene expression. By showcasing both established and innovative approaches, this collection provides practical guidance to help researchers avoid common pitfalls and implement robust, reproducible protocols in RNA therapeutics.