Pni formulation buffer

The circRNAs were encapsulated with lipid nanoparticles (LNPs) according to a previously described process (Ickenstein and Garidel, 2019 (link)). First, the circRNA was diluted with PNI Formulation Buffer (Precision NanoSystems, #NWW0043) to a final concentration of 170 μg/ml. Then, the lab-prepared or commercial LNP (Precision NanoSystems) were mixed with the circRNA solution at the volume ratio of 1:3 through the Ignite NxGen Cartridge (Precision NanoSystems, #NIT0002) using NanoAssemblr Ignite (Precision NanoSystems). Then the LNP-circRNA formulations were diluted 40-fold with 1×PBS buffer (pH 7.2∼7.4) and concentrated by ultrafiltration with Amicon® Ultra Centrifugal Filter Unit (Millipore). The concentration and encapsulation rate of circRNAs were measured by the Quant-it RiboGreen RNA Assay Kit (Invitrogen, #R11490). The size of LNP-circRNA particles was measured using dynamic light scattering on a Malvern Zetasizer Nano-ZS 300 (Malvern). Samples were irradiated with a red laser, and scattered light was detected. The results were analyzed to obtain an autocorrelation function using the software Zetasizer V7.13.