Rapid determination of adenoviral vector titers by quantitative real-time PCR.
Replication defective adenoviruses have been used as vectors in a variety of settings including gene transfer, gene manipulation, and functionality studies. A quantitative real-time PCR-based assay is described for rapid determination 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 quantitation 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 collection of adenoviral vectors.
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L
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Galapagos Genomics BV, Archimedesweg 4, PO Box 2048, 2301 CA Leiden, The Netherlands. libin@galapagos.nl
Bluyssen
H A
HA
De Raeymaeker
M
M
Laurysens
V
V
van der Beek
N
N
Pavliska
H
H
van Zonneveld
A J
AJ
Tomme
P
P
van Es
H H
HH
D000256:Adenoviridae / Q000302:isolation & purification*; D002460:Cell Line; D003199:Computer Systems; D005822:Genetic Vectors / Q000032:analysis*; D016133:Polymerase Chain Reaction / Q000379:methods; D014162:Transfection