Abstract
Quantification of the amount of cryoprotective agent (CPA) in a tissue is an essential step in the design of successful cryopreservation protocols. This chapter details two inexpensive methods to measure cryoprotective agent permeation into tissues as functions of time. One of the methods to measure the CPA permeation is to permeate a series of tissue samples from a surrounding solution at a specified concentration of CPA, each sample for a different amount of time, and then to quantitate the amount of CPA that was taken up in the tissue during that time period. The quantification is performed by equilibrating the permeated tissue with a surrounding solution and then measuring the osmolality of the solution to determine the amounts of CPAs that have come out of each tissue sample corresponding to each permeation time. An alternative method to measuring the CPA permeation as a function of time, which requires fewer tissue samples, is to measure the CPA efflux as a function of time. In the efflux method, a CPA-permeated tissue sample is placed in a surrounding solution, and solution samples are taken at different time points throughout the efflux to quantitate how much CPA has left the tissue by each time point.
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Acknowledgments
The articular cartilage research projects referred to in this manuscript were funded by the Edmonton Orthopaedic Research Committee and the Canadian Institutes of Health Research. Janet A. W. Elliott holds a Canada Research Chair in Thermodynamics. Kezhou Wu is funded by the Li Ka Shing Sino-Canadian Exchange Program between the University of Alberta and Shantou University. Nadia Shardt acknowledges funding from the Natural Sciences and Engineering Research Council of Canada (NSERC), Alberta Innovates and Alberta Advanced Education, the Government of Alberta, and the University of Alberta.
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Wu, K., Laouar, L., Shardt, N., Elliott, J.A.W., Jomha, N.M. (2021). Osmometric Measurements of Cryoprotective Agent Permeation into Tissues. In: Wolkers, W.F., Oldenhof, H. (eds) Cryopreservation and Freeze-Drying Protocols. Methods in Molecular Biology, vol 2180. Humana, New York, NY. https://doi.org/10.1007/978-1-0716-0783-1_11
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DOI: https://doi.org/10.1007/978-1-0716-0783-1_11
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