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Citation: Molecular Therapy — Methods Clinical Development (2014) 1, 14015; doi:10.1038/mtm.2014.15 © 2014 The American Society of Gene Cell Therapy All rights reserved 2329-0501/14 /mtm ARTICle Optimizing the production of suspension cells using the G-Rex “M” series 1 1 2 2 1 3 3 4 Pradip Bajgain , Roopa Mucharla , John Wilson , Dan Welch , Usanarat Anurathapan , Bitao Liang , Xiaohua Lu , Kyle Ripple , John M Centanni4 5 5 5 1 1 1 1 , Christine Hall , David Hsu , Larry A Couture , Shubhranshu Gupta , Adrian P Gee , Helen E Heslop , Ann M Leen , Cliona M Rooney1 1 and Juan F Vera Broader implementation of cell-based therapies has been hindered by the logistics associated with the expansion of clinically relevant cell numbers ex vivo. To overcome this limitation, Wilson Wolf Manufacturing developed the G-Rex, a cell culture flask with a gas-permeable membrane at the base that supports large media volumes without compromising gas exchange. Although this culture platform has recently gained traction with the scientific community due to its superior performance when compared with traditional culture systems, the limits of this technology have yet to be explored. In this study, we investigated multiple variables including optimal seeding density and media volume, as well as maximum cell output per unit of surface area. Additionally, we have identified a novel means of estimating culture growth kinetics. All of these parameters wer