Purifying stem cell-derived red blood cells: a high-throughput label-free downstream processing strategy based on microfluidic spiral inertial separation and membrane filtration

Guzniczak, E., Otto, O., Whyte, G., Chandra, T., Robertson, N. A., Willoughby, N., Jimenez, M. and Bridle, H. (2020) Purifying stem cell-derived red blood cells: a high-throughput label-free downstream processing strategy based on microfluidic spiral inertial separation and membrane filtration. Biotechnology and Bioengineering, 117(7), pp. 2032-2045. (doi: 10.1002/bit.27319) (PMID:32100873) (PMCID:PMC7383897)

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Abstract

Cell-based therapeutics, such as in vitro manufactured red blood cells (mRBC), are different to traditional biopharmaceutical products (the final product being the cells themselves as opposed to biological molecules such as proteins) and that presents a challenge of developing new robust and economically feasible manufacturing processes, especially for sample purification. Current purification technologies have limited throughput, rely on expensive fluorescent or magnetic immuno-labelling with a significant (up to 70%) cell loss and quality impairment. To address this challenge, previously characterised mechanical properties of umbilical cord blood CD34+ cells undergoing in vitro erythropoiesis were used to develop a mRBC purification strategy. The approach consists of two main stages: (1) a microfluidic separation using inertial focusing for deformability-based sorting of enucleated cells (mRBC) from nuclei and nucleated cells resulting in 70% purity and (2) membrane filtration to enhance the purity to 99%. Herein, we propose a new route for high-throughput (processing millions of cells /min and mls of medium /min) purification process for mRBC, leading to high mRBC purity while maintaining cells integrity and no alterations in their global gene expression profile. Further adaption of this separation approach offers a potential route for processing of a wide range of cellular products.

Item Type:Articles
Additional Information:FACS sorting was performed in the University of Edinburgh FACS Facility by Dr. Martin Waterfall. O. O. gratefully acknowledges funding from the German Federal Ministry of Education and Research (ZIK grant under grant agreement 03Z22CN11) and from the Deutschen Zentrum für Herz‐Kreislaufforschung. MJ would like to thank the Engineering and Physical Sciences Research Council (EPSRC) and the Royal Academy of Engineering for her Personal Research Fellowship (EP/R006482/1 and RF/201718/1741).
Keywords:Purification, deformability, sorting, spiral microchannel, stem cell-derived red blood cells.
Status:Published
Refereed:Yes
Glasgow Author(s) Enlighten ID:Jimenez, Dr Melanie
Authors: Guzniczak, E., Otto, O., Whyte, G., Chandra, T., Robertson, N. A., Willoughby, N., Jimenez, M., and Bridle, H.
College/School:College of Science and Engineering > School of Engineering > Biomedical Engineering
Journal Name:Biotechnology and Bioengineering
Publisher:Wiley
ISSN:0006-3592
ISSN (Online):1097-0290
Published Online:15 March 2020
Copyright Holders:Copyright © 2020 The Authors
First Published:First published in Biotechnology and Bioengineering 117(7): 2032-2045
Publisher Policy:Reproduced under a Creative Commons License

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Project CodeAward NoProject NamePrincipal InvestigatorFunder's NameFunder RefLead Dept
302160New Microsystems for Antimicrobial ResistanceMelanie JimenezEngineering and Physical Sciences Research Council (EPSRC)EP/R006482/1ENG - Biomedical Engineering