Improving AAV vector yield in insect cells by modulating the temperature after infection

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DOIResolve DOI: http://doi.org/10.1002/bit.21364
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TypeArticle
Journal titleBiotechnology and Bioengineering
Volume97
Issue6
Pages15011509; # of pages: 9
SubjectAdenoviridae; animals; bio; biotechnology; cell culture techniques; cell line; cells; cytology; genetic enhancement; genetic vectors; genetics; methods; physiology; protein; proteins; spodoptera; temperature; virology; virus cultivation; insect cells; baculovirus; adeno-associated virus
AbstractVectors based on adeno-associated viruses (AAV) are sought for therapeutic gene delivery because of their ability to transduce a variety of tissues with no significant immunological response. Production using the baculovirus expression vector (BEV)/insect cell system has the potential to meet the needs for pre-clinical and clinical trials. In this co-infection system, three baculoviruses are used to produce the AAV vector. A strategy aimed at increasing encapsidation/maturation of the viral vector involved varying the temperature over the course of the process. Cultures were subjected to temperature changes at various times pre- and post-infection (up to 24 h postinfection). It was found that raising the culture temperature to 30°C at the time of infection nearly tripled the infectious titer. In fact, increasing the temperature to 30°C at any time in the process investigated resulted in an increase in titer. Also, raising the culture to 33°C or lowering the temperature to 24° or 21°C resulted in lower titers. The rise in infectious titer was also confirmed by an increase in DNase resistant particles (DRPs). Varying the temperature, however, did not affect the total amount of capsids significantly. Therefore increasing the culture temperature resulted in better encapsidation as determined by the ratio of capsids to DRPs to infectious particles. It is believed that an increase in early proteins and possibly a quicker cascade of baculovirus infection events resulted in this increased packaging efficiency.
Publication date
LanguageEnglish
AffiliationNRC Biotechnology Research Institute; National Research Council Canada
Peer reviewedNo
NRC number47801
47801
NPARC number3539249
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Record identifierdc06082c-920d-42b8-a2ad-b247b3ea6ad0
Record created2009-03-01
Record modified2016-05-09
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