Ultra High Density Perfusion for Enabling Automated Cell Banking

The typical process for manufacturing biologics begins with the thaw of a cell bank and the expansion of inoculum culture. Current technologies for the generation of these cell banks can be labor intensive, and include several steps that introduce a significant risk of contamination or operator error. We describe here the development of an ultra-high-density, controlled cell culture process that mitigates these risks, and can be used for the production of both standard- and high-density cell banks. Alternating tangential flow perfusion technology was implemented and challenges of oxygen delivery and carbon dioxide accumulation were addressed. The final process supported high-viability cultures with ultra-high cell densities. These cultures were subjected to a revised banking protocol, which involved in situ addition of DMSO and direct vialing, eliminating the need for pooling cultures, centrifugation, or re-suspension, and increasing process consistency. Cell banks generated in this manner showed comparable recovery from thaw as traditional cell banks, and were considered suitable replacements for the standard-density banks. High-density, large volume cell banks generated using this process can be thawed directly into a seed train bioreactor, thus avoiding shake flask culture, reducing the time required for inoculum expansion by up to two weeks, and lowering the cost of production.