Figure 4. A. For the production of lower amounts of protein people usually transfect cells with conventional reagents. Since
efficiencies are often low they add a selection agent (e.g., G418) and grow cells under these conditions for eight weeks.
Substantial amounts of protein are then produced by this pool of stably transfected cells. B. Transient transfection of a
cell line is significantly faster as no selection is required.
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Working with a stable clone is optimal for large-scale production of therapeutic proteins. This is not desirable during protein
characterization, functional validation, or optimization. Researchers need higher flexibility. Selecting stably transfected
cells takes time, however, and even if polyclonal batch cultures are used, several weeks are required to produce a milligram
of protein (Figure 4A). Thus, as a compromise, researchers often use proteins derived from simpler expression systems in spite
of the fact that they deliver proteins that do not correspond well to genuine human patterns.
Figure 5. Production of Human IgG1 Antibody in 1 x 108 Suspension CHO Cells After Nucleofection.
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Relief is on the way. Efficient transient gene transfer systems can speed up the process of producing milligram amounts of
protein.7 Using novel transfection technologies it only takes a few days (Figure 4B). As an example, 2 mg of a human IgG1 antibody
was produced in four days after Nucleofection of 1 x 108 suspension CHO cells (Figure 5).
Oliver Gresch, Ph.D., and Hans-Guenter Bruenker, Ph.D., amaxa GmbH, Nattermannallee 1, 50829 Cologne, Germany, phone: +49 221 99199 0, fax: +49 221 99199 111, oliver.gresch@amaxa.com
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References:
1. Roy N and Agarwal S. Therapeutic Protein Production – An Overview. In: Cooper E editor. Business Briefing: Future Drug
Discovery. London: World Markets Research Center Ltd; 2003 p. 79-82.
2. Thiel K. Biomanufacturing, from bust to boom...to bubble? Nature Biotechnol. 2004; 22:1365-1372.
3. Wurm F. Production of recombinant protein therapeutics in cultivated mammalian cells. Nature Biotechnol. 2004; 22:1393-1398.
4. Thomas C, Ehrhardt A, Kay M. Progress and problems with the use of viral vectors for gene therapy. Nature Reviews Genetics.
2003; 4:346-358.
5. Niidome T and Huang L. Gene therapy progress and prospects: nonviral vectors. Gene Therapy. 2002; 9:1647-1652.
6. Gresch O, Engel F, Nesic D, et al. New non-viral method for gene transfer into primary cells. Methods. 2004; 33:151-163.
7. Schlaeger E, Kitas E and Dorn A. SEAP expression in transiently transfected mammalian cells grown in serum-free suspension
culture. Cytotechnology. 2003. 42:47-55.
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