Читать книгу Transfusion Medicine - Jeffrey McCullough - Страница 130

RBCs, frozen or deglycerolized, are the cells that have been stored in the frozen state at optimal temperatures in the presence of a cryoprotective agent, which is removed by washing before transfusion [24, 31]. The red cells must be frozen within 6 days after collection, and they can be stored for up to 10 years, although the AABB (formerly American Association of Blood Banks) Standards [24] do not include a standard for storage duration, and acceptable postthaw results have been found after storage at −80°C for 37 years [32–35]. The cryoprotectant commonly used is glycerol, which must be removed before transfusion to avoid osmotic hemolysis when the cells are transfused. The method of freezing and storage must preserve at least 80% of the original red cells, and at least 70% of those cells must survive 24 hours after transfusion.

Freezing of red cells is based on work from more than 50 years ago showing that glycerol protected human red cells from freezing injury [36], and that red cells preserved with glycerol were clinically effective [37–39]. From this work, “high‐” and “low‐concentration” glycerol methods were developed [40, 41]. These methods actually relate to the concentration of glycerol, which determines the nature of the freezing injury to the cells. When freezing is slow, extracellular ice forms, which increases the extracellular osmolarity, causing intracellular water to diffuse out of the cell and resulting in intracellular dehydration and damage [40]. This type of injury is prevented by solutes such as glycerol that penetrate the cell and minimize the dehydration [40]. Because the freezing process is slow, high concentrations of cryoprotectant, usually 40% glycerol, are required. Red cells preserved with this high concentration of glycerol can be stored at about −85°C, a temperature that is achievable by mechanical freezers.

Rapid freezing causes intracellular ice crystals and resulting cell damage [40]. However, because the freezing is faster, lower concentrations of cryoprotectant, usually about 20% glycerol, are effective [42]. This lower concentration of glycerol necessitates storage of red cells at a temperature of about −196°C, achievable only by using liquid nitrogen.

These two methods—with different freezing rates, concentrations of glycerol, storage conditions, and processes for removing the glycerol— involve different technologies [40, 42]. Technology development played a major role in making red cell freezing clinically available. During the 1970s, a disposable plastic bowl and semiautomated washing system were developed that greatly facilitated glycerol removal from high glycerol concentration red cells [43–45]. In the rapid‐freeze method, the concentration of glycerol is low enough that glycerol removal can be done by washing in ordinary blood bags; complex instruments are not required. To summarize, the high‐concentration glycerol method involves more simple freezing and storage but complex deglycerolizing procedures. The low‐concentration glycerol method involves complex freezing and storage but simple deglycerolizing procedures. Frozen deglycerolized red cells are composed of essentially red cells suspended in an electrolyte solution. Most of the plasma, platelets, and leukocytes have been removed either by the freezing, thawing, or washing step necessary to remove the glycerol cryoprotectant. Thus, the deglycerolized red cells have a 24‐hour storage period, which is a major factor in the logistics of their use. Frozen RBCs are used primarily for storage of rare RBC types [46].

Red cells must be frozen within 6 days of collection to provide acceptable posttransfusion survival. Red cells that have been stored longer than 6 days can be frozen if they are “rejuvenated” [47]. Rejuvenation restores metabolic functions after the red cells are incubated with solutions containing pyruvate, inosine, glucose, phosphate, and adenine followed by freezing [47]. This is a helpful strategy to freeze red cells in situations such as: (a) red cells found after 6 days of storage have a rare phenotype, (b) red cells donated for autologous transfusion but the surgery is postponed, and (c) rare phenotype red cells thawed but not used. The rejuvenation and subsequent freezing process is complex, expensive, and not widely used.