Viral quantitation: adeno-associated virus vector genome titer assay

Recombinant adeno-associated virus (AAV) is rapidly becoming the vector of choice for human gene therapy applications due to the ability to achieve prolonged gene expression in a variety of tissues without a significant immune response or toxicity. The most prevalent method for quantifying AAV vectors is currently quantitative PCR (qPCR). However, the reliance of this method on a standard curve and the potential impairment of DNA amplification efficiency from poor primer design, the presence of inhibitors, and template secondary structure can lead to variance and under-reporting of genome copy numbers. In this report, Martin Lock, Director, Process Development, Gene Therapy Program, University of Pennsylvania (PA, USA), discusses his laboratory’s comparison of qPCR and droplet digital PCR (ddPCR) for the absolute quantification of both single-stranded (ss) and self-complementary (sc) AAV genomes.