Abstract:
:As the use of adenoviral vectors in gene therapy protocols increases, there is a corresponding need for rapid, accurate, and reproducible titer methods. Multiple methods currently exist for determining titers of recombinant adenoviral vector, including optical absorbance, electron microscopy, fluorescent focus assay, and the "gold standard" plaque assay. This paper introduces a novel flow cytometric method for direct titer determination that relies on the expression of the green fluorescent protein (GFP), a tracking marker incorporated into several adenoviral vectors. This approach was compared to the plaque assay using 10(-4)- to 10(-6)-fold dilutions of a cesium-chloride-purified, GFP expressing adenovirus (AdEasy + GFP + GAL). The two approaches yielded similar titers: 3.25 +/- 1.85 x 10(9) PFU/mL versus 3.46 +/- 0.76 x 10(9) green fluorescent units/(gfu/mL). The flow cytometric method is complete within 24 h in contrast to the 7 x 10 days required by the plaque assay. These results indicate that the GFU/mL is an alternative functional titer method for fluorescent-tagged adenoviral vectors.
journal_name
Mol Biotechnoljournal_title
Molecular biotechnologyauthors
Hitt DC,Booth JL,Dandapani V,Pennington LR,Gimble JM,Metcalf Jdoi
10.1385/MB:14:3:197keywords:
subject
Has Abstractpub_date
2000-03-01 00:00:00pages
197-203issue
3eissn
1073-6085issn
1559-0305pii
MB:14:3:197journal_volume
14pub_type
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