Tracking the Evolution of Transiently Transfected Individual Cells in a Microfluidic Platform

Transient gene expression (TGE) technology enables the rapid production of large amount of recombinant proteins, without the need of fastidious screening of the producing cells required for stable transfection (ST). However, several barriers must be overcome before reaching the production yields using ST. For optimizing the production yields from suspended cells using TGE, a better understanding of the transfection conditions at the single cell level are required. In this study, a universal droplet microfluidic platform was used to assess the heterogeneities of CHO-S population transiently transfected with cationic liposomes (CL) (lipoplexes) complexed with GFP-coding plasmid DNA (pDNA). A single cell analysis of GFP production kinetics revealed the presence of a subpopulation producing higher levels of GFP compared with the main population. The size of high producing (HP) cells, their relative abundance, and their specific productivity were dependent on the charge and the pDNA content of the different lipoplexes: HPs showed increased cell size in comparison to the average population, lipoplexes with positive charge produced more HPs, and lipoplexes carrying a larger amount of pDNA yielded a higher specific productivity of HPs. This study demonstrates the potential for time-resolved single-cell measurements to explain population dynamics from a microscopic point of view.