Genome concentration, characterization, and integrity analysis of recombinant adeno-associated viral vectors using droplet digital PCR

ABSTRACT: Precise, reproducible characterization of AAV is critical for comparing preclinical results between laboratories and determining a safe and effective clinical dose for gene therapy applications. In this study, we systematically evaluated numerous parameters to produce a simple and robust ddPCR protocol for AAV characterization. The protocol uses a low ionic strength buffer containing Pluronic-F68 and polyadenylic acid to dilute the AAV into the ddPCR concentration range and a 10-minute thermal capsid lysis prior to assembling ddPCR reactions containing MspI. A critical finding is that the buffer composition affected the ITR concentration of AAV but not the ITR concentration of a double stranded plasmid, which has implications when using a theoretical, stoichiometric conversion factor to ob

SUBMITTER: Prantner A 

PROVIDER: S-EPMC9876284 | biostudies-literature | 2023 Jan

REPOSITORIES: biostudies-literature