143. Optimisation of current Good Manufacturing Practice (cGMP) compliant controlled rate freezing for human embryonic stem cells

Morris, G.J.; Acton, Elizabeth; Collins, Denis M.; Bös-Mikich, Adriana; Sousa, Paul De

doi:10.1016/j.cryobiol.2010.10.147

Cited by 5 publications

(2 citation statements)

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“…31 In this profile (Fig. 3, Profile B) there is an extended holding period in the temperature zone following ice nucleation, which allows more time for osmotic equilibrium to be established between the cells and the external environment at high subzero temperatures.…”

Section: Discussionmentioning

confidence: 99%

GMP Cryopreservation of Large Volumes of Cells for Regenerative Medicine: Active Control of the Freezing Process

MassieIsobel¹,

Selden²,

HodgsonHumphrey³

et al. 2014

Tissue Engineering Part C: Methods

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Cryopreservation protocols are increasingly required in regenerative medicine applications but must deliver functional products at clinical scale and comply with Good Manufacturing Process (GMP). While GMP cryopreservation is achievable on a small scale using a Stirling cryocooler-based controlled rate freezer (CRF) (EF600), successful large-scale GMP cryopreservation is more challenging due to heat transfer issues and control of ice nucleation, both complex events that impact success. We have developed a large-scale cryocoolerbased CRF (VIA Freeze) that can process larger volumes and have evaluated it using alginate-encapsulated liver cell (HepG2) spheroids (ELS). It is anticipated that ELS will comprise the cellular component of a bioartificial liver and will be required in volumes of *2 L for clinical use. Sample temperatures and Stirling cryocooler power consumption was recorded throughout cooling runs for both small (500 mL) and large (200 mL) volume samples. ELS recoveries were assessed using viability (FDA/PI staining with image analysis), cell number (nuclei count), and function (protein secretion), along with cryoscanning electron microscopy and freeze substitution techniques to identify possible injury mechanisms. Slow cooling profiles were successfully applied to samples in both the EF600 and the VIA Freeze, and a number of cooling and warming profiles were evaluated. An optimized cooling protocol with a nonlinear cooling profile from ice nucleation to -60°C was implemented in both the EF600 and VIA Freeze. In the VIA Freeze the nucleation of ice is detected by the control software, allowing both noninvasive detection of the nucleation event for quality control purposes and the potential to modify the cooling profile following ice nucleation in an active manner. When processing 200 mL of ELS in the VIA Freeze-viabilities at 93.4% -7.4%, viable cell numbers at 14.3 -1.7 million nuclei/mL alginate, and protein secretion at 10.5 -1.7 mg/ mL/24 h were obtained which, compared well with control ELS (viability -98.1% -0.9%; viable cell numbers -18.3 -1.0 million nuclei/mL alginate; and protein secretion -18.7 -1.8 mg/mL/24 h). Large volume GMP cryopreservation of ELS is possible with good functional recovery using the VIA Freeze and may also be applied to other regenerative medicine applications.

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Section: Discussionmentioning

confidence: 99%

GMP Cryopreservation of Large Volumes of Cells for Regenerative Medicine: Active Control of the Freezing Process

MassieIsobel¹,

Selden²,

HodgsonHumphrey³

et al. 2014

Tissue Engineering Part C: Methods

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“…Liquid nitrogen-free, CRFs, such as those employing the Stirling Cycle principle, may provide an alternative [112,113]. While such equipment provides a clean room-compatible solution for controlled rate freezing, they should be assessed for their ability to provide the required cooling rates, unit volumes and bank sizes appropriate to the cell lines being banked.…”

Section: Appendix 8: Preservation Technologies and Methodsmentioning

confidence: 99%

Points to Consider in the Development of Seed Stocks of Pluripotent Stem Cells for Clinical Applications: International Stem Cell Banking Initiative (ISCBI)

et al. 2015

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Fundamental Points to Consider in the Cryopreservation and Shipment of Cells for Human Application

Stacey

Healy

Man

et al. 2016

Bioprocessing for Cell Based Therapies

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143. Optimisation of current Good Manufacturing Practice (cGMP) compliant controlled rate freezing for human embryonic stem cells

Cited by 5 publications

References 0 publications

GMP Cryopreservation of Large Volumes of Cells for Regenerative Medicine: Active Control of the Freezing Process

GMP Cryopreservation of Large Volumes of Cells for Regenerative Medicine: Active Control of the Freezing Process

Points to Consider in the Development of Seed Stocks of Pluripotent Stem Cells for Clinical Applications: International Stem Cell Banking Initiative (ISCBI)

Fundamental Points to Consider in the Cryopreservation and Shipment of Cells for Human Application

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