The generation of an off-the-shelf in vitro engineered living tissue graft will likely require cryopreservation. However, the efficient addition and removal of cryoprotective agents (CPA) to cells throughout the volume of a three-dimensional (3D) tissue graft remains a significant challenge. In this work, we assessed whether a perfusion bioreactor-based method could be used to improve the viability of cryopreserved mesenchymal stromal cell- (MSC) based tissue constructs as compared to using conventional diffusion-based methods. The bioreactor was first used to saturate 3D constructs with CPA under perfused flow. Following cryopreservation, the bioreactor was then also used for the efficient removal of the CPA from the 3D tissues. We demonstrate that addition and removal of CPA under perfused flow significantly increased the viability of MSC within cryopreserved 3D tissue constructs as compared to conventional diffusion-based methods.

Department of Biomedicine University of Basel University Hospital Basel Basel Switzerland; Department of Biomedical Engineering University of Basel University Hospital Basel Basel Switzerland

Institute for Problems of Cryobiology and Cryomedicine NAS Ukraine Kharkov Ukraine

Institute for Problems of Cryobiology and Cryomedicine NAS Ukraine Kharkov Ukraine; Department of Biomaterials and Biophysical Methods Institute of Experimental Medicine AS CR v v i Prague Czech Republic; Department of Biomedicine University of Basel University Hospital Basel Basel Switzerland

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Hypothermic Storage of 3D Cultured Multipotent Mesenchymal Stromal Cells for Regenerative Medicine Applications