Fig. 2: The development of affinity-based magnetic isolation of LNPcor complexes.

a Schematic illustration of the ultrafast affinity-based, 96-well isolation method. Anti-PEG antibody-conjugated magnetic beads capture LNPcor from plasma containing free protein, extracellular vesicles and lipoprotein particles. Non-specifically bound biomolecules are removed with a series of wash steps. The LNPcor is eluted by alternating the pH. Up to 96 samples can be harvested in parallel within ~45 min. Illustration is generated with BioRender. b The design of experiment (DoE) space of LNPcor capture (epitope), wash and elution. As one of the LNPs’ structural components, PEG existing on the LNPs surface was employed here as a primary “endogenous” affinity tag, avoiding the need to add any additional surface antigens that might modify the LNPs’ surface properties and affect the biomolecular corona formation. To preserve the corona content and the LNPs themselves it was necessary to avoid commonly used detergents during washing and elution steps. A moderate change of pH was an effective and relatively mild way to disassociate LNPcor from antibodies. Nevertheless, a pH lower than the pKa of cationic ionizable lipid included in the LNPs (i.e., MC3, pKa = 6.44) usually results in LNP disassembly. The circles indicate where the majority of LNPs were detected. Only when combining an antibody against PEG backbone with PBS washing and basic elution conditions, were the majority of LNPs was identified in final elution. c The recovery ratio of LNPcor in terms of particle number, rhodamine labeled lipids (lipid) and Cy5 labeled mRNA (mRNA). Mean value (black bar) derived from independent experimental replicates (n = 2), with individual values indicated. Source data are provided as a Source Data file.