Two cryopreservation methods, colligative cryoprotection coupled with controlled cooling and vitrification-based, encapsulation-dehydration were validated by five members of the EU Research Infrastructure consortium, COBRA, and two independent external validators. The test strain Chlorella vulgaris SAG 211-11b was successfully cryopreserved using two-step cooling employing passive (Mr Frosty®) and Controlled Rate Freezers (CRF) attaining the desired recovery target within 15% of the median viability level (94%). Significant differences (P<0.05) between cooling regimes were observed where Mr Frosty® was more variable (Inter-Quartile Range being 21.5%, versus 13.0% for CRF samples). Viability assessment using fluorescein diacetate gave significantly (P<0.0001) higher survival than growth in agar with median values being 96% and 89%, respectively. On employing encapsulation-dehydration, greater variability between some validators was observed, with six labs observing recovery in 100% of the beads (84-95% of cells surviving) and one lab observing survival in 80% of the treated beads. Bead disruption followed by algal growth in agar was considered the most reliable and accurate method of assessing cell survival for encapsulation-dehydration.
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Document Type: Research Article
Publication date: 2007-09-01
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CryoLetters is a bimonthly international journal for low temperature sciences, including cryobiology, cryopreservation or vitrification of cells and tissues, chemical and physical aspects of freezing and drying, and studies involving ecology of cold environments, and cold adaptation
The journal publishes original research reports, authoritative reviews, technical developments and commissioned book reviews of studies of the effects produced by low temperatures on a wide variety of scientific and technical processes, or those involving low temperature techniques in the investigation of physical, chemical, biological and ecological problems.