Stabilization of Transfected Cells Expressing Low-Incidence Blood Group Antigens: Novel Methods Facilitating Their Use as Reagent-Cells.
BACKGROUND
The 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.


METHODS
We 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.


RESULTS
TransFix® 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.


CONCLUSIONS
Both 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.
Immunohematology Laboratory, Banc de Sang i Teixits, Barcelona, Spain.
Cecilia
C
González
Immunohematology Laboratory, Banc de Sang i Teixits, Barcelona, Spain.
Rosa
R
Esteban
Immunohematology Laboratory, Banc de Sang i Teixits, Barcelona, Spain.
Carme
C
Canals
Immunohematology Laboratory, Banc de Sang i Teixits, Barcelona, Spain.
Eduardo
E
Muñiz-Díaz
Immunohematology Laboratory, Banc de Sang i Teixits, Barcelona, Spain.
Núria
N
Nogués
D000818:Animals; D001789:Blood Group Antigens / Q000096:biosynthesis / Q000097:blood* / Q000276:immunology*; D001793:Blood Preservation; D016466:CHO Cells; D002460:Cell Line; D058948:Cell Tracking / Q000379:methods; D003412:Cricetulus; D015925:Cryopreservation; D004906:Erythrocyte Count; D004912:Erythrocytes / Q000276:immunology*; D005434:Flow Cytometry; D005612:Freeze Drying; D006801:Humans; D016624:Negative Staining; D014162:Transfection
D001789:Blood Group Antigens