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Chapters
- 00:05Introduction
- 00:44Generation of Baculovirus-infected TIPS Cells
- 03:05Production of AAV Vector
- 05:08Lysis of Cells and Release of AAV
- 06:07Purification of AAV Vector Using Affinity Column Chromatography System
- 08:03Concentration and Diafiltration of AAV Vector Using Tangential Flow Filtration (TFF)
- 08:54Results: Purification of Adeno-Associated Virus (AAV)2 Vector Using Baculovirus-Insect Cell Culture System
- 09:48Conclusion
In dit protocol wordt de AAV2-vector geproduceerd door het co-kweken van Spodoptera frugiperda (Sf9) insectencellen met baculovirus (BV)-AAV2-groen fluorescerend eiwit (GFP) of therapeutisch gen en BV-AAV2-rep-cap geïnfecteerde Sf9-cellen in suspensiecultuur. AAV-deeltjes komen vrij uit de cellen met behulp van detergent, worden geklaard, gezuiverd door affiniteitskolomchromatografie en geconcentreerd door tangentiële stromingsfiltratie.
