Understanding Temperature Profiles Experienced by Biological Samples During a Hybrid Vitrification Technique

From thermodynamic point of view, vitrification is considered as a superior preservation technique in comparison with the traditional slow-cooling cryopreservation techniques, due to formation of the glassy state in both intra and extracellular environment. While vitrification of biological samples are difficult to achieve, recently a hybrid technique involving partial desiccation of the cell samples prior to cryogenic exposure has been successfully employed to achieve vitrification. In this technique cells in monolayer attached to a substrate was suspended in a trehalose solution and then rapidly and uniformly desiccated to a low moisture content (<0.12 g of water per g of dry weight) using a spin-drying technique. The spin-dried samples were stored in liquid nitrogen (LN₂) at a vitrified state (Fig. 1). It was shown that following re-warming to room temperature and re-hydration with a fully complemented cell culture medium, 51% of the spin-dried and vitrified cells survived and demonstrated normal growth characteristics. The current study further investigates the temperature profiles experienced by the cell samples during partial desiccation and cryogenic exposure to identify possible ways to improve this novel vitrification strategy. Physical Vapor Deposition technique was employed to develop glass substrate having thermocouples (2×2×2 μm) at four radial positions across the substrate to record the thermal history of the cell samples during the entire process. Efforts are undertaken to understand the uncertainties related temperature measurement spatially and with respect to time. These temperature characterization studies are important to optimize the newly developed hybrid vitrification technique for vitrification of cellular samples.