High Throughput Transient Transfections of Hek293Ebna Cells — Devel Opment of an Integrated Expression Purification Process at 100 ml Scale

This work describes the set-up and optimisation of a simple, robust and cost effective process for the high throughput production of recombinant proteins using HEK293EBNA cells at 100 ml scale. Several transfection reagents were tested and evaluated for simplicity of transfection procedure, cost, reproducibility and expression yield. A purification process was set-up, which allows the fast purification of tagged proteins via a robotized system. The reproducibility and robustness of the whole process was monitored by inclusion of a well-expressed protein into the daily production batches. The statistical analysis of the purification yields of this control protein revealed that the process is operator independent and uncritical to cell density and nutrient conditions within the seed cultures. Over a period of more than a year more than 1,200 transfections were carried out yielding more than 630 different proteins in microgram quantities for screening purposes.