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Related Experiment Video

Updated: Jul 17, 2026

Process Development for the Production and Purification of Adeno-Associated Virus (AAV)2 Vector using Baculovirus-Insect Cell Culture System

Process Development for the Production and Purification of Adeno-Associated Virus (AAV)2 Vector using Baculovirus-Insect Cell Culture System

Published on: January 13, 2022

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

Marc G Aucoin¹, Michel Perrier, Amine A Kamen

¹Biotechnology Research Institute, National Research Council of Canada, 6100 Royalmount Ave., Montréal, Quebec, Canada.

Biotechnology and Bioengineering

|February 3, 2007

Summary

Optimizing adeno-associated virus (AAV) vector production using the baculovirus expression vector (BEV)/insect cell system, researchers found that increasing culture temperature to 30°C significantly boosted infectious titer and encapsidation efficiency.

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Process Development for the Production and Purification of Adeno-Associated Virus (AAV)2 Vector using Baculovirus-Insect Cell Culture System

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Area of Science:

Biotechnology
Gene Therapy
Virology

Background:

Adeno-associated virus (AAV) vectors are promising for therapeutic gene delivery due to their broad tissue tropism and low immunogenicity.
The baculovirus expression vector (BEV)/insect cell system offers a scalable method for producing AAV vectors for clinical applications.

Purpose of the Study:

To optimize the production of therapeutic adeno-associated virus (AAV) vectors using a baculovirus expression vector (BEV)/insect cell system.
To investigate the impact of temperature variations on AAV vector encapsidation and infectious titer.

Main Methods:

A co-infection system utilizing three baculoviruses was employed for AAV vector production.
Culture temperatures were manipulated at various time points pre- and post-infection (up to 24 hours).
Infectious titers and DNase resistant particles (DRPs) were quantified to assess vector quality and quantity.

Main Results:

Raising the culture temperature to 30°C at the time of infection nearly tripled the infectious titer of AAV vectors.
Increased temperature to 30°C at any investigated stage enhanced AAV vector titer.
Higher temperatures (33°C) or lower temperatures (24°C, 21°C) resulted in reduced titers, while 30°C optimized encapsidation efficiency.

Conclusions:

Temperature optimization, specifically a shift to 30°C, significantly enhances adeno-associated virus (AAV) vector production in the baculovirus expression vector (BEV)/insect cell system.
This temperature-mediated increase in titer is attributed to improved encapsidation efficiency, as evidenced by the ratio of capsids to DNase resistant particles (DRPs).
The findings suggest that elevated early protein expression and accelerated baculovirus infection cascades at 30°C contribute to increased packaging efficiency.