(16a) Snagging mRNA in Wormlike Micelle Networks

The increased demand for mRNA vaccines requires new analytical methods to keep pace with manufacturing and quality control requirements. In multivalent vaccines, payloads of mRNA with similar but not identical lengths in the range of 2000-3000 bases must be independently dosed and checked carefully for purity. Because gel electrophoresis is slow and unable to discriminate mRNA in this length range, vaccine manufacturers rely on liquid chromatography for routine characterization, with attendant solvent and capital expense. Further, these methods are not practical for deployment in remote areas where vaccines may be subjected to intermittent lapses in proper storage.

Faster, higher-resolution electrophoretic separations of kilobase DNA or RNA can be accomplished using weakly overlapping networks of wormlike micelles made of mixtures of nonionic surfactant. These networks present a temporary gel to the migrating ssRNA, where pores in the gel continually dissolve and re-form due to micelle breakup and lateral diffusion processes. Using these materials, rapid, length-based separations can be realized up to a critical length where resolution is lost. In this “biased reptation” limit, the migration of the ssRNA is controlled by time scale of pore breakup rather than that of chain electrophoresis.

Here we demonstrate that attachment of 18-carbon hydrophobes and 10-100 base DNA oligomers gives large changes in the electrophoretic mobility of ssRNA in the biased reptation limit. These attachments locally hinder the transit of ssRNA through pores in the vicinity of the attachment, a process we refer to as “snagging.” In so doing, the migration of the entire kilobase chain is drastically altered by a seemingly small modification. In fact, attachments of oligomers longer than about 30 bases gives no additional mobility shift as they are longer than the transient pore size.

We present results confirming this snagging mechanism and present a scaling theory to describe the migration of tagged ssRNA in these wormlike micelle networks. We will also discuss applications of the method for mRNA vaccine analysis and virus quantitation in complex systems.