Rapid determination of adenoviral vector titers by quantitative real-time PCR

Replication defective adenoviruses have been used as vectors in a variety o f settings including gene transfer, gene manipulation, and functionality st udies. A quantitative real-time PCR-based assay is described for rapid dete rmination of physical titers of recombinant adenovirus vectors. This method is based on amplification of a 77 bp fragment located near the left end of the adenovirus type 5 genome. Evaluation of this method demonstrated that it is simple, sensitive and reproducible, and has a dynamic range of quanti tation over 5 logs. This assay is applicable to purified adenovirus as well as vectors prepared by simple cell lysis procedure, requiring only a small amount of starting material. The simplicity and short turn-around time of this assay should facilitate rapid titer determination for a large collecti on of adenoviral vectors. (C) 2001 Elsevier Science B.V. All rights reserve d.