title: Spatial considerations during cryopreservation of a large volume sample.
abstract: There have been relatively few studies on the implications of the physical conditions experienced by cells during large volume (litres) cryopreservation - most studies have focused on the problem of cryopreservation of smaller volumes, typically up to 2 ml. This study explores the effects of ice growth by progressive solidification, generally seen during larger scale cryopreservation, on encapsulated liver hepatocyte spheroids, and it develops a method to reliably sample different regions across the frozen cores of samples experiencing progressive solidification. These issues are examined in the context of a Bioartificial Liver Device which requires cryopreservation of a 2 L volume in a strict cylindrical geometry for optimal clinical delivery. Progressive solidification cannot be avoided in this arrangement. In such a system optimal cryoprotectant concentrations and cooling rates are known. However, applying these parameters to a large volume is challenging due to the thermal mass and subsequent thermal lag. The specific impact of this to the cryopreservation outcome is required. Under conditions of progressive solidification, the spatial location of Encapsulated Liver Spheroids had a strong impact on post-thaw recovery. Cells in areas first and last to solidify demonstrated significantly impaired post-thaw function, whereas areas solidifying through the majority of the process exhibited higher post-thaw outcome. It was also found that samples where the ice thawed more rapidly had greater post-thaw viability 24 h post-thaw (75.7 ± 3.9% and 62.0 ± 7.2% respectively). These findings have implications for the cryopreservation of large volumes with a rigid shape and for the cryopreservation of a Bioartificial Liver Device.
authors.affiliation: Institute for Liver and Digestive Health, Royal Free Hospital Campus, UCL, London, NW3 2PF, UK; Asymptote Ltd. St. John's Innovation Centre, Cowley Road, Cambridge, CB4 0WS, UK. Electronic address: peter.kilbride.11@ucl.ac.uk.
authors.forename: Peter
authors.initials: P
authors.lastname: Kilbride
authors.affiliation: Asymptote Ltd. St. John's Innovation Centre, Cowley Road, Cambridge, CB4 0WS, UK.
authors.forename: Stephen
authors.initials: S
authors.lastname: Lamb
authors.affiliation: Asymptote Ltd. St. John's Innovation Centre, Cowley Road, Cambridge, CB4 0WS, UK.
authors.forename: Stuart
authors.initials: S
authors.lastname: Milne
authors.affiliation: Institute for Liver and Digestive Health, Royal Free Hospital Campus, UCL, London, NW3 2PF, UK.
authors.forename: Stephanie
authors.initials: S
authors.lastname: Gibbons
authors.affiliation: Institute for Liver and Digestive Health, Royal Free Hospital Campus, UCL, London, NW3 2PF, UK.
authors.forename: Eloy
authors.initials: E
authors.lastname: Erro
authors.affiliation: Institute for Liver and Digestive Health, Royal Free Hospital Campus, UCL, London, NW3 2PF, UK.
authors.forename: James
authors.initials: J
authors.lastname: Bundy
authors.affiliation: Institute for Liver and Digestive Health, Royal Free Hospital Campus, UCL, London, NW3 2PF, UK.
authors.forename: Clare
authors.initials: C
authors.lastname: Selden
authors.affiliation: Department of Surgery, Royal Free Hospital Campus, UCL, London, NW3 2PF, UK.
authors.forename: Barry
authors.initials: B
authors.lastname: Fuller
authors.affiliation: Asymptote Ltd. St. John's Innovation Centre, Cowley Road, Cambridge, CB4 0WS, UK.
authors.forename: John
authors.initials: J
authors.lastname: Morris
mesh_terms: D000818:Animals; D015925:Cryopreservation / Q000379:methods*; D003451:Cryoprotective Agents / Q000494:pharmacology; D005615:Freezing; D022781:Hepatocytes / Q000166:cytology; D006801:Humans; D019164:Liver, Artificial; D008297:Male; D018874:Spheroids, Cellular / Q000166:cytology
chemical_list: D003451:Cryoprotective Agents
keywords: Bioartificial liver; HepG2; Large volume cryopreservation; Progressive solidification