Abstract
BackgroundThe identification of erythrocyte antibodies in the serum of patients rely on panels of human red blood cells (RBCs), which coexpress many antigens and are not easily available for low-incidence blood group phenotypes. These problems have been addressed by generating cell lines expressing unique blood group antigens, which may be used as an alternative to human RBCs. However, the use of cell lines implies several drawbacks, like the requirement of cell culture facilities and the high cost of cryopreservation. The application of cell stabilization methods could facilitate their use as reagent cells in clinical laboratories.MethodsWe generated stably-transfected cells expressing low-incidence blood group antigens (Dia and Lua). High-expresser clones were used to assess the effect of TransFix® treatment and lyophilization as cell preservation methods. Cells were kept at 4°C and cell morphology, membrane permeability and antigenic properties were evaluated at several time-points after treatment.ResultsTransFix® addition to cell suspensions allows cell stabilization and proper antigen detection for at least 120 days, despite an increase in membrane permeability and a reduction in antigen expression levels. Lyophilized cells showed minor morphological changes and antigen expression levels were rather conserved at days 1, 15 and 120, indicating a high stability of the freeze-dried product. These stabilized cells have been proved to react specifically with human sera containing alloantibodies.ConclusionsBoth stabilization methods allow long-term preservation of the transfected cells antigenic properties and may facilitate their distribution and use as reagent-cells expressing low-incidence antigens, overcoming the limited availability of such rare RBCs.
Highlights
Antibodies against blood group antigens can induce clinical conditions such as haemolytic transfusion reactions, haemolytic disease of the fetus and newborn (HDFN) and autoimmune haemolytic anaemia
These problems have been addressed by generating cell lines expressing unique blood group antigens, which may be used as an alternative to human red blood cells (RBCs)
Stabilization of Reagent Cells Expressing Blood Group Antigens funders had no role in study design, data collection and analysis, or preparation of the manuscript
Summary
Antibodies against blood group antigens can induce clinical conditions such as haemolytic transfusion reactions, haemolytic disease of the fetus and newborn (HDFN) and autoimmune haemolytic anaemia. Current antibody identification methods rely on panels of human red blood cells (RBCs) that have a limited viability and may carry biohazard risks These panel RBCs simultaneously express a large number of antigens, which makes the antibody identification method to be based on the lack of reactivity with antigen-negative cells. RBCs expressing low-incidence blood group antigens are not available, which hampers their inclusion in these panels These problems have been addressed by generating cell lines stably expressing a unique RBC membrane protein, which may be used as reagent-cells to identify antibodies in the serum of sensitized patients. The application of cell stabilization methods could facilitate their use as reagent cells in clinical laboratories.
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