title: Efficient large volume lentiviral vector production using flow electroporation.
abstract: Lentiviral vectors are beginning to emerge as a viable choice for human gene therapy. Here, we describe a method that combines the convenience of a suspension cell line with a scalable, nonchemically based, and GMP-compliant transfection technique known as flow electroporation (EP). Flow EP parameters for serum-free adapted HEK293FT cells were optimized to limit toxicity and maximize titers. Using a third generation, HIV-based, lentiviral vector system pseudotyped with the vesicular stomatitis glycoprotein envelope, both small- and large-volume transfections produced titers over 1×10(8) infectious units/mL. Therefore, an excellent option for implementing large-scale, clinical lentiviral productions is flow EP of suspension cell lines.
authors.affiliation: Department of Medical and Molecular Genetics, Indiana University School of Medicine , Indianapolis, IN 46202, USA.
authors.forename: Scott R
authors.initials: SR
authors.lastname: Witting
authors.forename: Lin-Hong
authors.initials: LH
authors.lastname: Li
authors.forename: Aparna
authors.initials: A
authors.lastname: Jasti
authors.forename: Cornell
authors.initials: C
authors.lastname: Allen
authors.forename: Kenneth
authors.initials: K
authors.lastname: Cornetta
authors.forename: James
authors.initials: J
authors.lastname: Brady
authors.forename: Rama
authors.initials: R
authors.lastname: Shivakumar
authors.forename: Madhusudan V
authors.initials: MV
authors.lastname: Peshwa
mesh_terms: D019149:Bioreactors; D002470:Cell Survival; D016895:Culture Media, Serum-Free; D003850:Deoxyribonuclease I / Q000378:metabolism; D018274:Electroporation; D005822:Genetic Vectors / Q000096:biosynthesis* / Q000235:genetics; D057809:HEK293 Cells; D006801:Humans; D016086:Lentivirus / Q000235:genetics*; D010957:Plasmids; D011994:Recombinant Proteins / Q000378:metabolism; D012212:Rheology; D014162:Transfection; D018116:Vesiculovirus / Q000737:chemistry / Q000235:genetics*; D014764:Viral Proteins / Q000737:chemistry / Q000235:genetics*
chemical_list: D016895:Culture Media, Serum-Free; D011994:Recombinant Proteins; D014764:Viral Proteins; D003850:Deoxyribonuclease I; C568813:dornase alfa