Abstract
We describe a method for polyethyleneimine (PEI) mediated transient transfection of suspension-adapted Chinese hamster ovary (CHO-DG44) cells for protein expression applicable at scales from 2mL to 2L. The method involves transfection at a high cell density (5×106 cells/mL) by direct addition of plasmid DNA (pDNA) and PEI to the culture and subsequent incubation at 31°C with agitation by orbital shaking. This method requires 0.3mg/L of coding pDNA, 2.7mg/L of nonspecific (filler) DNA and 15mg/L of PEI. The production phase is performed at 31°C in the presence of 0.25% N,N-dimethylacetamide (DMA). We also provide information on culture vessel options, recommended working volumes, and recommended shaking speeds for transfections at scales from 2mL to 2L.
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