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Updated: Feb 5, 2026

Production and Titering of Recombinant Adeno-associated Viral Vectors
Published on: November 27, 2011
Scalable Production and Purification of Adeno-Associated Viral Vectors (AAV)
Daniel Blessing1,2,3, Nicole Déglon4,5, Bernard L Schneider6
1Laboratory of Neurotherapies and Neuromodulation, Department of Clinical Neurosciences, Lausanne University Hospital, Lausanne, Switzerland. daniel.blessing@epfl.ch.
Abstract:
Here we describe methods for the production of adeno-associated viral (AAV) vectors by transient transfection of HEK293 cells grown in serum-free medium in orbital shaken bioreactors and the subsequent purification of vector particles. The protocol for expression of AAV components is based on polyethyleneimine (PEI) mediated transfection of a 2-plasmid system and is specified for production in milliliter to liter scales. After PEI and plasmid DNA (pDNA) complex formation the diluted cell culture is transfected without a prior concentration step or medium exchange. Following a 3-day batch process, cell cultures are further processed using different methods for lysis and recovery. Methods for the purification of viral particles are described, including iodixanol gradient purification, immunoaffinity chromatography, and ultrafiltration, as well as quantitative PCR to quantify vector titer.
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