Effect of red blood cell preservation by droplet freezing with non‐permeable cryoprotective agents in blood group antigen reactivity

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Red blood cell (RBC) transfusion is important in the treatment of patients with diseases such as haemoglobinopathies, myelodysplastic syndromes, chronic renal disease and others. However, a relevant complication of RBC transfusion is the production of alloantibodies, which can increase the risk of haemolytic transfusion reactions, reduce the efficacy of transfusion therapy and, in extreme cases, threaten the life of the transfusion recipient (Hendrickson & Tormey, 2016; Nickel et al., 2016).
The accurate screening and identification of RBC antibodies is necessary to identify alloimmunized patients and to find compatible blood units (Sood et al., 2013). The classical method for antibody screening and identification is haemagglutination using the indirect antiglobulin test (IAT), which is commonly performed with a red cell panel (AABB). However, erythrocytes of commercial panels do not always present the necessary phenotypes, and it is sometimes unfeasible to identify antibodies, especially in the cases of patients with rare phenotypes or in the presence of multiple antibodies (Garozzo et al., 2007). In these cases, it is necessary to make use of additional selected red cells with different antigen combinations.
Frequently, reference immunohaematology laboratories use non‐commercial RBCs to confirm or rule out irregular antibodies. These RBCs can be obtained from phenotyped blood donors and may be preserved by freezing in small volumes for future use. In RBC cryopreservation protocols, cryoprotective agents (CPAs) are used to prevent RBC injury during freezing/thawing (Schmid et al., 2011; Henkelman et al., 2015), allowing the storage of erythrocytes for use in immunohaematological tests for several years.
Previous studies have shown that RBCs maintain their rheological properties, antigenic pattern and usability for serologic testing after freezing (Bronson & McGinniss, 1962; Pegg et al., 1982; Henkelman et al., 2010). Although the freezing of RBCs in glycerol is widely used by blood group reference laboratories (Smith, 1950; Högman et al., 1986), the preservation in liquid nitrogen by droplet freezing, using non‐permeable CPAs such as polyvinylpyrrolidone (PVP) and sucrose–dextrose solution (S + D), has emerged as an effective alternative for the cryopreservation of small aliquots of erythrocytes for posterior use in serological tests (Schmid et al., 2011).
Schmid et al. (2011) compared RBC preservation by droplet freezing with PVP or S + D and by bulk freezing with glycerol (n = 14), and showed that RBC preservation with PVP is a very effective method, providing 85% of RBC recovery. However, the stability of RBC antigens was not evaluated. In this study, we compared the antigenic pattern of RBCs preserved by droplet freezing in liquid nitrogen with two CPAs – S + D and PVP in the context of a blood bank.
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