title: Scale-up of the adenovirus expression system for the production of recombinant protein in human 293S cells.
abstract: Human 293S cells, a cell line adapted to suspension culture, were grown to 5 x 10(6) cells/mL in batch with calcium-free DMEM. These cells, infected with new constructions of adenovirus vectors, yielded as much as 10 to 20% recombinant protein with respect to the total cellular protein content. Until recently, high specific productivity of recombinant protein was limited to low cell density infected cultures of no more than 5 x 10(5) cells/mL. In this paper, we show with a model protein, Protein Tyrosine Phosphatase 1C, how product yield can be maintained at high cell densities of 2 x 10(6) cells/mL by a medium replacement strategy. This allows the production of as much as 90 mg/L of active recombinant protein per culture volume. Analysis of key limiting/inhibiting medium components showed that glucose addition along with pH control can yield the same productivity as a medium replacement strategy at high cell density in calcium-free DMEM. Finally, the above results were reproduced in 3L bioreactor suspension culture thereby establishing the scalability of this expression system. The process we developed is used routinely with the same success for the production of various recombinant proteins and viruses.
authors.affiliation: Institut de recherche en biotechnologie, CNRC, Montréal, Québec, Canada.
authors.forename: A
authors.initials: A
authors.lastname: Garnier
authors.forename: J
authors.initials: J
authors.lastname: Côté
authors.forename: I
authors.initials: I
authors.lastname: Nadeau
authors.forename: A
authors.initials: A
authors.lastname: Kamen
authors.forename: B
authors.initials: B
authors.lastname: Massie
mesh_terms: D000260:Adenoviruses, Human; D001709:Biotechnology / Q000295:instrumentation / Q000379:methods; D002455:Cell Division; D002460:Cell Line; D002470:Cell Survival; D003470:Culture Media; D046508:Culture Techniques / Q000295:instrumentation / Q000379:methods; D004591:Electrophoresis, Polyacrylamide Gel; D005822:Genetic Vectors; D006801:Humans; D007668:Kidney; D007700:Kinetics; D017027:Protein Tyrosine Phosphatases / Q000096:biosynthesis* / Q000302:isolation & purification; D011994:Recombinant Proteins / Q000096:biosynthesis* / Q000302:isolation & purification; D013997:Time Factors; D014162:Transfection
chemical_list: D003470:Culture Media; D011994:Recombinant Proteins; D017027:Protein Tyrosine Phosphatases