Transfection Potency\nof Lipid Nanoparticles Containing\nmRNA Depends on Relative Loading Levels

When formulating mRNA into lipid nanoparticles (LNP),\nvarious copy\nnumbers of mRNA are encapsulated, leading to a distribution of mRNA\nloading levels within the LNPs. It is unclear whether the mRNA loading\nlevel affects the functional delivery of the message. Here we show\nthat depending on the mRNA loading level, LNPs exhibit distinct mass\ndensities and can be fractionated via ultracentrifugation. Upon fractionation,\nwe investigated if mRNA loading levels influence LNP sizing, lipid\ncomposition, and morphology. We further conducted <i>in vitro</i> and <i>in vivo</i> functional delivery of mRNA and found\nthat the LNP fraction with the highest mRNA loading levels was the\nleast transfection competent.