(344e) Automated Liquid Handler Advancements for High-Throughput Scale-Down Perfusion Cell Culture Screening

Perfusion bioreactor technology is highly versatile and can provide increases to productivity in addition to enabling the production of unstable or toxic proteins that are traditionally difficult to produce in fed-batch systems. Process development activities such as process optimization, clone selection, and media screening require a large number of experimental conditions and a representative high-throughput small-scale model is desirable. Automated microscale bioreactor tools like the ambr15 offer a useful option to culture cells, but perfusion cell culture faces challenges due in part to the complexity of mimicking cell separation at larger scales. Progress has been made to solve this issue by utilizing cell sedimentation, which has the benefit of being executed entirely within the ambr15 environment, but it subjects the cells to extended periods of hypoxia due to the long settling time and the relatively slow speed of the single channel ambr15 liquid handler. A faster and more representative alternative is provided by centrifugation of the vessels to pellet the cells and harvest the supernatant with an automated multichannel fluid handler, but this has previouslyrequired significant operator involvement. Here, we developed a workflow consisting of a Tecan Fluent with an integrated centrifuge and custom 3D-printed attachments. Vessels are removed from ambr15 and placed in an automated workstation. Once in the Tecan environment, the entire process of decapping vessels, cell separation by centrifugation, media exchange, and initial media addition is automated and requires no operator interaction. This reduces the time for a vessel to be without agitation, oxygen, and pH control from 90 minutes to 20 minutes. The workflow demonstrated comparable cell growth profiles, productivity, and product quality to our bench-scale intensified N-1 perfusion step. Additionally, the fully automated workflow lowers the barrier to entry for operators by automating a previously manually complex process and has reduced contamination rates from the perfusion step from 10% to 0%.