NUMERICAL INVESTIGATION OF A NOVEL MICROFLUIDIC SYSTEM AND ITS APPLICATION IN ACHIEVING ULTRA-FAST COOLING/WARMING RATES FOR CELL VITRIFICATION CRYOPRESERVATION

摘要

Background: During conventional cryopreservation, cells are often damaged by ice formation. An alternative approach is to use ultra-rapid cooling rates to achieve vitrification of cell suspension. A variety of systems and methods have been developed, e.g. open pulled straws (OPS), closed pulled straws (CPS) , cryo-loop and micro-droplets, by which cooling/warming rates up to 10~4～10~5 °C/min can be achieved. Although these methods have been well used in some areas, they are still far from perfect because (1) the cooling/warming rates are still not fast enough and thus high-concentration cryoprotectants should be used; (2) the sample size is often seriously limited. Method of approach: The lacks of existing methods are analyzed from the point of view of heat transfer principle as follows. Firstly, the heat transfer coefficient (h) between sample and cooling/warming media is usually insufficient. During the cooling process, the direct plunging high temperature sample into liquid nitrogen results in strong boiling and vaporization of liquid nitrogen around the sample surface (Leidenfrost effect), and forms a "vapor coat" which acts as a heat-insulation layer. As a result, the heat transfer coefficient is badly limited (about 1～2×10~3 W/m~2·K or even lower ). For the rewarming process, the heat transfer coefficient is also insufficient for the conventional water bath method since only simple conduction and limited convection dominate in the process. Secondly, most of the sample carriers are made of plastic, and their large thermal mass (as a consequence of its material and thickness) also limits the heat transfer efficiency. Some carrier free methods avoid this problem but increases the risk of loss or contamination of cells by the direct contact with cooling/warming media. Thirdly, the geometries of the samples in these methods are generally cylindrical or spherical, which limit the ratio of the sample surface versus its volume. As a result, the heat conduction inside the samples significantly lowers the sample cooling/warming rates.