Strategies for the cryopreservation of microencapsulated cells.
The major challenge in developing cryopreservation protocols for microencapsulated cells is that the relatively large size (300-400 microm) and the fragile semipermeable membrane of microcapsules makes them particularly prone to cryodamage. Rapid-cooling cryopreservation protocols with high DMSO concentrations (3.5M, 25% v/v) resulted in low post-thaw cell viability (<10%), which did not improve with higher concentrations (4.5M, 32% v/v) and longer exposure to DMSO, even though the majority of microcapsules (60-80%) remained intact. Subsequent investigations of slow cooling with a range of DMSO and EG concentrations resulted in a much higher post-thaw cell viability (80-85%), with the majority of the microcapsules remaining intact ( approximately 60%) when DMSO was used at a concentration of 2.8M (20% v/v) and EG at a concentration of 2.7M (15% v/v). The presence of 0.25M sucrose significantly improved post-thaw cell viability upon slow cooling with 2.8M (20% v/v) DMSO, although it had no effect on microcapsule integrity. Multistep exposure and removal of sucrose did not significantly improve either post-thaw cell viability or microcapsule integrity, compared to a single-step protocol. Ficoll 20% (w/v) also did not significantly improve post-thaw cell viability and microcapsule integrity. Hence, the optimal condition for microcapsule cryopreservation developed in this study is slow cooling with 2.8M (20% v/v) DMSO and 0.25M sucrose.
Heng
Boon Chin
BC
Department of Obstetrics & Gynaecology, Faculty of Medicine, National University of Singapore, Lower Kent Ridge Road, Singapore. obgngsc@nus.edu.sg
Yu
Hanry
H
Chye Ng
Soon
S
D000818:Animals; D002460:Cell Line; D002470:Cell Survival / Q000187:drug effects / Q000502:physiology; D002522:Chlorocebus aethiops; D020099:Coated Materials, Biocompatible / Q000737:chemistry; D015925:Cryopreservation / Q000379:methods*; D003451:Cryoprotective Agents / Q000494:pharmacology; D004121:Dimethyl Sulfoxide / Q000737:chemistry / Q000494:pharmacology*; D019855:Ethylene Glycol / Q000737:chemistry / Q000494:pharmacology*; D007668:Kidney / Q000737:chemistry / Q000166:cytology* / Q000187:drug effects / Q000502:physiology*; D008567:Membranes, Artificial; D008863:Microspheres; D010316:Particle Size; D013048:Specimen Handling / Q000379:methods; D013395:Sucrose / Q000737:chemistry / Q000378:metabolism*
D020099:Coated Materials, Biocompatible; D003451:Cryoprotective Agents; D008567:Membranes, Artificial; D013395:Sucrose; D019855:Ethylene Glycol; D004121:Dimethyl Sulfoxide