High cell density transient transfection of CHO cellsfor TGF-beta1 expression

Abdalla A. Elshereef, André Jochums Antonina Lavrentieva,Lena Stuckenberg, Thomas Scheper Dörte Solle
Source: Eng Life Sci.
Publication Date: (2019)
Issue: 9:
Research Area:
Drug Discovery
Culture Media:


High cell densities for transient transfection with polyethyleneimine (PEI) can be used
for rapid and maximal production of recombinant proteins. High cell densities can be
obtained by different cultivation systems, such as batch or perfusion systems. Herein,
densities up to 18 million cells/mL were obtained by centrifugation for transfection
evaluation. PEI transfection efficiency was easily determined by transfected enhanced
green fluorescence protein (EGFP) reporter plasmid DNA (pDNA). A linear correlation
between fluorescence intensity and transfection efficiency was improved. The
transfection efficiency of PEI was highly dependent on the transfection conditions
and directly related to the level of recombinant protein. Several factors were required
to optimize the transient transfection process; these factors included the media type
(which is compatible with low or high cell density transfection), the preculture CHOK1
suspension cell density, and the pDNAto PEI level. Based on design of experiment
(DoE) analyses, the optimal transfection conditions for 10 × 106 cells/mL in the
CHOMACS CD medium achieved 73% transfection efficiency and a cell viability of
over 80%. These results were confirmed for the production of transforming growth
factor-beta 1 (TGF-??1) in a shake flask. The purified TGF-??1 protein concentration
from 60 mL supernatant was 27 µg/mL, and the protein was biologically active.