title: Analysis of the production efficiency and titration of various recombinant adeno-associated viruses.
abstract: Recombinant adeno-associated virus type 2 (rAAV2) viral vector, a non-pathogenic human parvovirus, has recently emerged as a gene transfer vehicle for cancer gene therapy. To utilize rAAV2 properly and safely while carrying out preclinical and clinical studies, it is crucial to exactly titer the virus. We therefore compared biological infectious rAAV2 titers with physical titers of rAAV2 vectors encoding various transgenes with different sized viral genomes. Biological rAAV2 infectivity was assayed by measuring the number of virus particles able to transduce Hela cells using several detection methods, including X-gal staining and immunocytostaining. Physical titers of rAAV2 were determined using a commercially available rAAV2 particle-specific enzyme-linked immunosorbent assay. We found that total rAAV2 particle production was consistent within the limited size variations of the rAAV2 genome, regardless of the difference in transgenes. In contrast, the infectious titer of rAAV2 differed greatly, even for the same viruses, due to variation in the sensitivity of the relevant assays. Thus, the results suggest that both infectious virus titer and total virus particle should be precisely measured for rAAV2 vector utilized in each study.
authors.affiliation: Department of Microbiology, University of Ulsan College of Medicine, Seoul, Korea.
authors.forename: Young Ran
authors.initials: YR
authors.lastname: Nam
authors.forename: Sung Jin
authors.initials: SJ
authors.lastname: Kim
authors.forename: Boyoung
authors.initials: B
authors.lastname: Lee
authors.forename: Jin Woo
authors.initials: JW
authors.lastname: Chang
authors.forename: Chul-Hyen
authors.initials: CH
authors.lastname: Joo
authors.forename: Yoo Kyum
authors.initials: YK
authors.lastname: Kim
authors.forename: Heuiran
authors.initials: H
authors.lastname: Lee
mesh_terms: D004279:DNA, Viral / Q000737:chemistry*; D000229:Dependovirus / Q000235:genetics / Q000254:growth & development* / Q000472:pathogenicity*; D005822:Genetic Vectors / Q000502:physiology*; D006367:HeLa Cells; D006801:Humans; D007124:Immunoenzyme Techniques; D011995:Recombination, Genetic; D017186:Titrimetry; D014779:Virus Replication; D001616:beta-Galactosidase / Q000378:metabolism
chemical_list: D004279:DNA, Viral; D001616:beta-Galactosidase