Journal: Human Gene Therapy Methods
Article Title: Sodium Chloride Enhances Recombinant Adeno-Associated Virus Production in a Serum-Free Suspension Manufacturing Platform Using the Herpes Simplex Virus System
Figure Lengend Snippet: Characterization of rAAV vector made in Expi293F suspension system. (A and B) rAAV9-GFP was produced by rHSV infection in adherent HEK293 cells grown in 10-chamber CellSTACKs (CS10) under optimal conditions (approximately 3 × 10 9 cells) either in DMEM containing 5% FBS alone ( n = 4) or supplemented with 60 m M sodium chloride ( n = 6), or in a 3-liter suspension culture containing 3 × 10 9 Expi293F cells ( n = 5). Total vector genome (A) and transducing unit (B) yields are shown. Total purified vector genomes are represented relative to units produced per cell (C) and per liter of medium (D) . (E) The vector genome-to-transducing unit (VG/TU) ratio was calculated for each sample, and the averaged values for each group are shown. (F) Coomassie staining of rAAV9-GFP final stocks produced in HEK293 CS10 in the absence of salt supplementation (lane 1), or supplemented with sodium chloride (lane 2), or produced in Expi293F 3-liter cultures supplemented with sodium chloride (lanes 3–7). A total of 10 11 VG was loaded per well. * p
Article Snippet: Cell lines Suspension BHK21C13-2P (BHK) cells were obtained from Sigma-Aldrich (St. Louis, MO) and maintained in Dulbecco's modified Eagle's medium (DMEM; HyClone/GE Healthcare Life Sciences, Logan, UT) supplemented with 5% FBS (Corning, Corning, NY) and 1% antibiotic/antimycotic (Gibco/Thermo Fisher Scientific, Waltham, MA).
Techniques: Plasmid Preparation, Produced, Infection, Purification, Staining