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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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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

DOI:
10.3791/62829-v

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10:31 min

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January 13, 2022

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, , ,
1Division of Experimental Hematology and Cancer Biology,Cincinnati Children’s Hospital Medical Center, 2University of Cincinnati College of Medicine, 3Raymond G. Perelman Center for Cellular and Molecular Therapeutics,The Children’s Hospital of Philadelphia

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

Summary

Automatic Translation

Automatic Translation

In this protocol, AAV2 vector is produced by co-culturing Spodoptera frugiperda (Sf9) insect cells with baculovirus (BV)-AAV2-green fluorescent protein (GFP) or therapeutic gene and BV-AAV2-rep-cap infected Sf9 cells in suspension culture. AAV particles are released from the cells using detergent, clarified, purified by affinity column chromatography, and concentrated by tangential flow filtration.

Tags

Keywords: Adeno-associated VirusAAVBaculovirusInsect Cell CultureSf9 CellsTIPS CellsVirus ProductionVirus PurificationFlow CytometryCryopreservation

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