003 Radiofrequency heating of magnetic nanoparticle cryoprotectant solutions for improved cryopreservation protocols

• Published in: Cryobiology 2013-12, Vol.67 (3), p.398-399
• Main Authors: Etheridge, Michael L., Xu, Yi, Choi, Jeunghwan, Bischof, John C.
• Format: Article
• Language: English
• ISSN: 0011-2240; 1090-2392
• DOI: 10.1016/j.cryobiol.2013.09.009

While cryopreservation through vitrification holds great promise, practical application has been limited to smaller systems (cells and thin tissues) due to diffusive transfer and phase-change limitations, which are typically manifested as devitrification and cracking failures during thaw. Here we describe a new approach for rapidly and uniformly heating cryopreserved biospecimens with radiofrequency (RF) excited magnetic nanoparticles (mNPs). Importantly, heating rates can be increased several fold over conventional boundary heating and are independent of sample size and dielectric properties, overcoming the inhomogeneity associated with microwave thawing. Proof-of-principle experiments in aqueous and cryoprotectant solutions are presented and then modeling is used to further illustrate the potential of this innovative approach. Three embodiments of mNP suspensions are studied – aqueous, 6M glycerol in phosphate buffered saline, and 8.4M VS55 cryoprotectant solution (Mehl, Cryobiology 30 (1993) 509–518). The iron oxide mNPs used are commercially available (Ferrotec, Inc.) and have been previously shown to produce significant heating in aqueous suspensions (Etheridge and Bischof, ABME 41 (2013) 78–88). Nanoparticle concentrations up to 10mg Fe/ml are shown to have negligible impact on the freezing behavior of these solutions, including the glass transition, critical cooling/warming rates, and specific heat, as measured by differential scanning calorimetry. One milliliter mNP suspensions at 5 and 10mg Fe/ml in water and cryoprotectant solution were frozen in liquid nitrogen (at sufficient rates to achieve vitrification in the cryoprotectant samples) and then reheated in an insulated, 370kHz and 20kA/m RF coil. Sample temperatures were monitored with 40-gauge thermocouples embedded prior to cooling. RF fields can produce interference in metallic thermocouples, but this was characterized and found to be negligible for the fine gauge thermocouples used. Heating rates at the glass transition of approximately 200° C/min were achieved in the 10mg Fe/ml samples, avoiding devitrification in the cryoprotectant solutions. Since RF fields at frequencies