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Monitoring of the Adenovirus Production Process by Flow Cytometry
Author(s) -
Sandhu Kalbinder Singh,
AlRubeai Mohamed
Publication year - 2008
Publication title -
biotechnology progress
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 0.572
H-Index - 129
eISSN - 1520-6033
pISSN - 8756-7938
DOI - 10.1021/bp070198s
Subject(s) - flow cytometry , recombinant dna , viral vector , green fluorescent protein , cell culture , genetic enhancement , biology , reporter gene , adenoviridae , dna , virus , microbiology and biotechnology , computational biology , gene , virology , gene expression , genetics
Adenovirus (Ad) has become the vector of choice for gene therapy clinical protocols worldwide; it is the only viral vector to date that has been licensed for use in a gene therapy treatment. There is, however, a need to develop a simple, reliable at‐line method to monitor the production of virus and recombinant proteins (r‐proteins) that have no intrinsic reporter properties. Here we utilize flow cytometry to measure cell size, granularity, and DNA content in a single‐step analysis and to correlate these parameters to the production of a type‐5 Ad (Ad5) expressing the recombinant green fluorescent protein (GFP). Clear correlations between these parameters and productivity are made, with forward scatter and DNA content showing the highest correlation coefficients, 0.9 and 0.83 for virus production and r‐protein production, respectively. Measuring these parameters requires little or no processing of the cells from culture to analysis. These parameters have been used successfully to monitor, at‐line, the amount of Ad and r‐protein product in a 293‐Ad system.