Quantitative Assessment of the Physical Virus Titer and Purity by Ultrasensitive Flow Virometry

Abstract Rapid quantification of viruses is vital for basic research on viral diseases as well as biomedical application of virus‐based products. Here, we report the development of a high‐throughput single‐particle method to enumerate intact viral particles by ultrasensitive flow virometry, which detects single viruses as small as 27 nm in diameter. The nucleic acid dye SYTO 82 was used to stain the viral (or vector) genome, and a laboratory‐built nano‐flow cytometer (nFCM) was employed to simultaneously detect the side‐scatter and fluorescence signals of individual viral particles. Using the bacteriophage T7 as a model system, intact virions were completely discriminated from empty capsids and naked viral genomes. Successful measurement of the physical virus titer and purity was demonstrated for recombinant adenoviruses, which could be used for gene delivery, therapeutic products derived from phage cocktails, and infected cell supernatants for veterinary vaccine production.