Development of a generic transient transfection process at 100 L scale

Area of Science:

• Biotechnology
• Cell Biology
• Biopharmaceutical Manufacturing

Background:

• Transient gene expression in mammalian cells is crucial for recombinant protein production.
• Scaling up transient transfection processes presents challenges in maintaining efficiency and yield.
• Optimizing cell lines, media, and transfection parameters is key for large-scale biomanufacturing.

Purpose of the Study:

• To develop and optimize a scalable transient transfection process for recombinant IgG production.
• To establish a closed system for IgG production from cell seeding to protein capture.
• To achieve reproducible yields of high-quality IgG for in vitro and in vivo studies.

Main Methods:

• Utilized HEK293-EBNA cells and polyethylenimine (PEI) for transient transfection at 100 L scale.
• Optimized DNA:PEI ratio, investigated conditioned medium effects, and implemented a feed strategy.
• Employed continuous centrifugation for medium exchange and Streamline expanded bed chromatography for capture.

Main Results:

• Achieved reproducible production of >0.5 g recombinant IgG.
• Demonstrated process scalability from 10 L to 100 L, with yields of 5-8 mg/L IgG.
• Developed a closed process from seeding to protein capture, solving large-scale medium exchange challenges.

Conclusions:

• The developed transient transfection protocol is a robust and scalable method for recombinant IgG production.
• The optimized process allows for efficient capture and purification of IgG.
• This approach supports the consistent supply of recombinant IgG for preclinical and clinical research.