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Effect of temperature, medium composition, and cell passage on production of herpes‐based viral vectors
Author(s) -
Wechuck James B.,
Ozuer Ali,
Goins William F.,
Wolfe Darren,
Oligino Thomas,
Glorioso Joseph C.,
Ataai Mohammad M.
Publication year - 2002
Publication title -
biotechnology and bioengineering
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.136
H-Index - 189
eISSN - 1097-0290
pISSN - 0006-3592
DOI - 10.1002/bit.10310
Subject(s) - titer , vector (molecular biology) , herpes simplex virus , biology , viral vector , virology , cell culture , virus , chemistry , gene , recombinant dna , biochemistry , genetics
Our work uses replication‐defective genomic herpes simplex virus type‐1 (HSV‐1)‐based vectors to transfer therapeutic genes into cells of the central nervous system and other tissues. Obtaining highly purified high‐titer vector stocks is one of the major obstacles remaining in the use of these vectors in gene therapy applications. We have examined the effects of temperature and media conditions on the half‐life of HSV‐1 vectors. The results reveal that HSV stability is 2.5‐fold greater at 33°C than at 37°C and is further stabilized at 4°C. Additionally, a significantly higher half‐life was measured for the vector in infection culture conditioned serum medium compared to fresh medium with or without serum. Synchronous infections incubated at 33°C produced 2‐fold higher amounts of vector than infected cells incubated at 37°C, but with a lag of 16–24 h. Vector production yielded 3‐fold higher titers and remained stable at peak levels for a longer period of time in cultures incubated at 33°C than 37°C. A pronounced negative effect of increased cell passage number on vector yield was observed. Vector production at 33°C yielded similar levels regardless of passage number but was reduced at 37°C as passage number increased. Together, these results contribute to improved methods for high‐titer HSV vector production. © 2002 Wiley Periodicals, Inc. Biotechnol Bioeng 79: 112–119, 2002.

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