Rapid removal of glycerol from frozen‐thawed red blood cells

Abstract

The storage of red blood cells (RBCs) in a refrigerated state allows a shelf life of a few weeks, whereas RBCs frozen in 40% glycerol have a shelf life of 10 years. Despite the clear logistical advantages of frozen blood, it is not widely used in transfusion medicine. One of the main reasons is that existing post-thaw washing methods to remove glycerol are prohibitively time consuming, requiring about an hour to remove glycerol from a single unit of blood. In this study, we have investigated the potential for more rapid removal of glycerol. Using published biophysical data for human RBCs, we mathematically optimized a three-step deglycerolization process, yielding a procedure that was less than 32 s long. This procedure was found to yield 70% hemolysis, a value that was much higher than expected. Consequently, we systematically evaluated three-step deglycerolization procedures, varying the solution composition and equilibration time in each step. Our best results consisted of less than 20% hemolysis for a deglycerolization time of 3 min, and it is expected that even further improvements could be made with a more thorough optimization and more reliable biophysical data. Our results demonstrate the potential for significantly reducing the deglycerolization time compared with existing methods. © 2013 American Institute of Chemical Engineers Biotechnol. Prog., 29:609-620, 2013.

Comparative effectiveness of Dimethyl Sulphoxide (DMSO) and Glycerol as cryoprotective agent in preserving Vero cells

“…In contrast, Feradis et al (2001) found that glycerol and DMSO at 6% concentration produced equivalent survival rates of cynomolgus epididymal sperm after cryopreservation. Lusianti et al (2013) observed that storage of red blood cells (RBCs) in a refrigerated state allowed a shelf-life of a few weeks, whereas RBCs frozen in 40% glycerol had a shelf-life of 10 years.…”

Foundations of modeling in cryobiology—I: Concentration, Gibbs energy, and chemical potential relationships

“…While not complete in their description of the processes, the predictions of these models have been extremely useful in guiding experimental design, demonstrating feasibility of approaches, etc. [11,12,20,36,44,[48][49][50]69,78,84,122]. Many advances in computational complexity, transport and solution theories [10,28,32,57,97], as well as experimental parameter identification techniques [24][25][26]58,116,117] have been made along the way, facilitating more accurate models and better predictions of intracellular ice formation.…”

Fatigue damage to pig erythrocytes during repeated swelling and shrinkage

“…The supernatant was centrifuged again at 10,000g for 5 min to remove the cell membrane fragments. Finally, the hemoglobin concentration in the supernatant was detected using an automatic biochemistry analyzer, and the mortality of erythrocytes was estimated according to the percentage of hemolysis (mortality of erythrocytes % percentage of hemolysis = hemoglobin concentration in the supernatant  100%/hemoglobin concentration in completely lysed sample [15]). The experiments were repeated 3 times.…”

“…Existing studies have suggested that the volume set point is closely related to cell function, cell proliferation, and cell death [10,[12][13][14]. In practice, the volume excursion of cells should be in the range between the upper and the lower volume tolerance limits (different cell types have different tolerance limits; for human erythrocytes, the upper and lower volume tolerance limits are approximately 1.38 and 0.53 times of the isotonic cell volume, respectively [15]); otherwise, the functional integrity of cells cannot be maintained [8,16]. Therefore, osmotic tolerance limits [8,[16][17][18], volume excursion [10,13] and the volume set point can be considered as indicators to evaluate osmotic damage to cells.…”

“…However, because of its time-consuming, complex operation; lower cell recovery rate; and mechanical stress damage [21,22], scientists are exploring the washing routines of the variety of washing machines [15]. In addition, various alternative approaches have been also proposed, mainly including the dialysis-based method [23][24][25] dilution-filtration method [26][27][28][29], and theoretical work has been performed [30].…”