The cryopreservation of cells has been in routine use for decades. However, despite the extensive research in the field, cryopreservation of large tissues and organs is still experimental. The present review highlights the major studies of directional freezing and vitrification of large tissues and whole organs and describes the different parameters that impact the success rate of large tissue and organ cryopreservation. Key factors, such as mass and heat transfer, cryoprotectant toxicity, nucleation, crystal growth, and chilling injury, which all have a significant influence on whole-organ cryopreservation outcomes, are reviewed. In addition, an overview of the principles of directional freezing and vitrification is given and the manners in which cryopreservation impacts large tissues and organs are described in detail.

Cryopreservation of ovarian tissue has been considered experimental for many years, but very recently the American Society of Reproductive Medicine is reviewing the process and perhaps soon will remove the label of “experimental” and recognize it as an established method for preserving female fertility when gonadotoxic treatments cannot be delayed or in patients before puberty or when there is desire to cryopreserve more than just few oocytes. This article discusses in detail the 3 methodologies used for cryopreservation: (a) slow freezing, (b) directional freezing, and (c) vitrification.

Red blood cell (RBC) units are administered routinely into patients expressing a wide range of acute and chronic conditions (e.g., anemia, traumatic bleeding, chronic diseases, and surgery). The modern blood banking system has been designed to answer this need and assure a continuous, high quality blood supply to patients. However, RBCs units can be stored under hypothermic conditions for only up to 42 days, which leads to periodic shortages. Cryopreservation can solve these shortages, but current freezing methods employ high glycerol concentrations, which need to be removed and the cells washed prior to transfusion, resulting in a long (more than 1 hour) and cumbersome washing step. Thus, frozen RBCs have limited use in acute and trauma situations. In addition, transportation of frozen samples is complicated and costly. Freeze drying (lyophilization) of RBCs has been suggested as a solution for these problems, since it will allow for a low weight sample to be stored at room temperature, but reaching this goal is not a simple task. We studied the effect of different solutions (IMT2 and IMT3) containing trehalose and antioxidants or trehalose and human serum albumin, respectively, on freezing/thawing and freeze drying of RBCs. In addition, we evaluated the effect of cells concentrations and cooling rates on the post thaw and post rehydration recoveries of the RBCs. Finally, we developed a new radio frequency (RF) lyophilization device for a more rapid and homogeneous sublimation process of the frozen RBCs samples. Recovery and free Hb were measured as well as oxygen association/dissociation and cell's deformability. We found that IMT3 (0.3 M trehalose and 10% HSA) solution that was directionally frozen at a rapid interface velocity of 1 mm/sec (resulting in a cooling rate of 150°C/min) yielded the best results (better than IMT2 solution and slow interface velocity). Freeze thawing gave 100% survival, while freeze drying followed by rehydration with 20% dextran-40kDa solution resulted in 75% survival. However, recovery following freeze drying was possible only when 20% Dextran-40 solution was used as the rehydration medium. The rehydrated cells were not stable upon an eight-fold dilution. The RF lyophilization system increased the sublimation rate more than twice compared to conventional drying and maintained a high survival rate of the RBCs after partial drying.

Currently vitrification is the method of choice for low-temperature preservation of oocytes and embryos. However, that was not the case until about 10 years ago when freezing methods were used relatively successfully for embryos and investigated (unsuccessfully) for oocyte preservation. In this paper we will review the history of oocyte and embryo cryopreservation and look into ways and methods for overcoming and improving the vitrification method since it suffers from inherent disadvantages since it is a cumbersome, time-consuming and costly procedure.