Enhanced Discriminability of Viral Vectors in Viscous Nanopores.

Abstract

Achieving safe and efficient gene therapy hinges upon the inspection of genomes enclosed within individual nano-carriers to mitigate potential health risks associated with empty or fragment-filled vectors. Here solid-state nanopore sensing is reported for identifications of intermediate adeno-associated virus (AAV) vectors in liquid. The method exploits the phenomenon of translocation slowdown induced by the viscosity of salt water-organic mixtures. This enables real-time ionic current measurements allowing precise tracking of the electroosmotic flow-driven motions of recombinant AAV vectors in a nanopore. The resulting ionic signals facilitate discrimination between replicative intermediates carrying ssDNA fragments and its full vector counterparts based on genome length-derived subtle nanometer differences in the viral diameters. This rapid and non-destructive means of genome analysis within virus capsids provides a promising avenue toward a robust methodology for ensuring the integrity of AAV vectors before administration.