Production of Adeno-Associated Virus Vectors in Cell Stacks for Preclinical Studies in Large Animal Models

This protocol for production of AAV viral vectors is cost-effective and yields high purity, high titer, research-grade AAV for use in preclinical, large animal models. This technique uses adherent HEK293 cells in cell culture chambers, which is time efficient and less hands-on, allowing for fewer disruptions to the cells. This protocol can greatly aid in viral vector production for the field of gene therapy, and can also be used for the production of other AAV serotypes that also bind heparan sulfate.

To begin, collect HEK293 cells from the culture when it reaches 80%confluency. Aspirate the media before gently washing the cell culture plate with three milliliters of PBS. Then aspirate PBS and add three milliliters of trypsin.