An optimized HEK293T cell expansion protocol using a hollow-fiber bioreactor system

Viral vectors are commonly used to introduce genetic material into cells to modify cell function for a variety of purposes. Manufacture of those modified viruses may use a variety of cell types to generate high titers of viral particles; one of the most common being HEK293 cells. These cells have been modified into different lines aimed at satisfying specific use cases. HEK293T cells, for example, have been modified to include the SV40 large T antigen. Efficient viral particle production by HEK293T cells requires the maintenance of favorable cell culture conditions during expansion and transfection. This protocol describes the use of the Quantum(®) hollow-fiber bioreactor (HFB) system for the automated expansion of HEK293T cells, and the results derived using the protocol described herein were not compared with those from tissue culture flasks or other expansion platforms, as the parameters described are unique to Quantum’s hollow fiber cell expansion environment. The purpose of this protocol is to help users of Quantum to focus on relevant parameters of expansion in the HFB milieu and to provide guidelines for a successful expansion of HEK293T cells in the Quantum system. The steps provided have been optimized to reliably control environmental factors related to glucose, lactate, and pH. Data reflecting this consistency are provided along with procedural time points reflected in text and figure formats.