Determination of Lentiviral Infectious Titer by a Novel Droplet Digital PCR Method.

Abstract

Lentivirus is one of the best vehicles in delivering exogenous genes for therapeutics. Prior to application, it is very important to determine the infectious titer, which measures only mature virus capable of infecting target cells. Quantitative polymerase chain reaction (PCR) and fluorescence-activated cell sorting are commonly used for determination of infectious titer. This study introduces a new method based on Droplet Digital PCR (ddPCR), a recently developed PCR technology that quantifies the absolute amount of target DNA in the reaction. In this study, the dynamic range, Limit of Quantification (LOQ), and data acceptance criteria for ddPCR are defined against lentiviral sequence. ddPCR performance is also compared to established FACS and qPCR methods. This work not only demonstrates the feasibility of ddPCR in determining lentiviral infectious titer, but provides a detailed method that can be easily adapted by the scientific community.