Cryomacroscopy in 3d: a Device Prototype for the Study of Cryopreservation

Light Source INTRODUCTION This study presents a new device prototype for visualization of physical effects associated with large-scale cryopreservation—the preservation of tissues at very low temperatures. Cryopreservation represents the only method for long-term preservation of biomaterials. While techniques for cryopreservation of single cells and small tissue structures are well established, cryopreservation techniques for bulky tissues and organs are still at the developmental stage. Critical to the success of cryopreservation is the control of ice formation—the cornerstone of cryoinjury. One of the most promising techniques for large-scale cryopreservation is known as vitrification, where the crystal phase is suppressed, and the biological material is trapped in a glassy-like state (vitreous in Latin means glassy) [1]. Vitrification is achieved with the addition of a high concentration of cryoprotective agents (CPAs), which experience an exponential increase in viscosity with the decreasing temperature and, thereby, suppress crystallization. Hence, the likelihood of crystallization during vitrification is dependent upon the specific thermal history. Crystallization may initiate during cooling, may continue during rewarming—recrystallization, or even initiate during rewarming— devitrification. Another potentially devastating effect during vitrification is thermo-mechanical stress, caused by the high cooling and rewarming rates required to promote vitrification. When the developing stress exceeds the strength of the material, structural damage follows, with fracture formation as its most dramatic outcome [2]. The current study focuses on developing a device—the cryomacroscope—to visual physical effects along the path-dependent process of vitrification. This device is based on an early prototype developed to visual physical effects in thin vitrified specimens [3]. The innovation in the current device is in the use of a mechanism to scan bulky specimens relevant to large-scale cryopreservation. Design considerations for the new cryomacroscope prototype are derived from its potential versatile use: (i) as a basic research tool, (ii) as a tool to develop cryopreservation solutions and protocols, and (iii) as a quality assurance tool for mass production of cryopreservation products. The current study combines solid mechanics analysis to explain the onset of fracturing in selected cryopreservation experiments. EXPERIMENTAL SETUP Figure 1 displays a general view of the scanning cryomacroscope prototype, designed to be compatible with the commercial controlledrate freezer, Kryo10-16 (Planer, Inc.., UK). Figure 2 displays an experimentation platform developed for basic research studies, used in the current study. The specimen container in this setup is a cuvette, rather than a vial or a bag, to improve the quality of imaging while minimizing optical distortion (Fig. 2). Proceedings of the ASME 2012 Summer Bioengineering Conference SBC2012 June 20-23, Fajardo, Puerto Rico, USA