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Background These last decades have seen the emergence and development ofcell-based therapies, notably those based on mesenchymal stromal cells (MSCs).The advancement of these promising treatments requires increasing thethroughput of processed cell for industrialization in order to reduceproduction costs. Among the various bioproduction challenges, downstreamprocessing (DSP), including medium exchange, cell washing, cell harvesting andvolume reduction, remains a critical step on which improvements are needed.Typically, these processes are performed by centrifugation. However, thisapproach limits the automation, especially in small batch productions where itis performed manually in open system. Methods An acoustophoresis-based systemwas developed for cell washing. The cells were transferred from one stream toanother via the acoustic forces and were collected in a different medium. Theoptimal flow rates of the different streams were assessed using Red Blood Cells(RBCs) suspended in an albumin solution. Finally, the impact of acousticwashing on adipose tissue-derived MSCs (AD-MSCs) transcriptome was investigatedby RNA-sequencing. Results With a single passage through the acoustic device atinput flow rate of 45 mL/h, the albumin removal was up to 90% while recovering99% of RBCs. To further increase the protein removal, a loop washing in twosteps was performed and has allowed an albumin removal $\ge$99% and aRBCs/AD-MSCs recovery of 99%. After loop washing of AD-MSCs, only 2 genes, HES4and MIR-3648-1, were differently expressed compared to the input. Conclusion Inthis study, we developed a continuous cell washing system based onacoustophoresis. The process allows a high cell throughput while inducinglittle gene expression changes. These results highly suggest that cell washingbased on acoustophoresis is a relevant and promising solution for numerousapplications in cell manufacturing.

Feasibility of an Acoustophoresis-based System for a High-Throughput Cell Washing: Application to Bioproduction