Observation and analysis of the ultra-rapid cooling effect of cryoprotectant droplets on a sapphire surface

Abstract

Abstract Cryopreservation has emerged as a promising technology to realize semipermanent storage of biomaterials widely used in assisted reproduction and cell therapy. A large portion of successful cryopreservation requires the combination of ultra-rapid cooling and appropriate low-toxicity cryoprotectant agents, which inhibits the formation of ice crystals that damages the cells lethally. Conventional approaches are not competent in accurately capturing high enough cooling rate and more importantly, not able to simultaneously track the instant appearance of the sample at a microscale level during the momentary cooling process. A rapid freezing device for the droplet sample study purpose was designed and developed, which is capable of visualization of the sample on a sapphire surface at a vitrification cooling rate over 104 K/min. The instantaneous temperature changes and the appearance of the cooling samples were recorded at the speed of 10 kHz and 2000 frames. Typical vitrification and crystallization processes were observed and analysed. Low concentrations of the CPA solution inhibited the formation and growth of the crystals, and resulted in the formation of layered ice, punctate ice and scattered small ice. The experimental results could contribute to a better understanding of the ultra-rapid cooling mechanism on cryoprotectant droplets.