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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 expressin...

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Autores principales: González, Cecilia, Esteban, Rosa, Canals, Carme, Muñiz-Díaz, Eduardo, Nogués, Núria
Formato: Online Artículo Texto
Lenguaje:English
Publicado: Public Library of Science 2016
Materias:
Acceso en línea:https://www.ncbi.nlm.nih.gov/pmc/articles/PMC5014343/
https://www.ncbi.nlm.nih.gov/pubmed/27603310
http://dx.doi.org/10.1371/journal.pone.0161968
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author González, Cecilia
Esteban, Rosa
Canals, Carme
Muñiz-Díaz, Eduardo
Nogués, Núria
author_facet González, Cecilia
Esteban, Rosa
Canals, Carme
Muñiz-Díaz, Eduardo
Nogués, Núria
author_sort González, Cecilia
collection PubMed
description 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 (Di(a) and Lu(a)). 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.
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spelling pubmed-50143432016-09-27 Stabilization of Transfected Cells Expressing Low-Incidence Blood Group Antigens: Novel Methods Facilitating Their Use as Reagent-Cells González, Cecilia Esteban, Rosa Canals, Carme Muñiz-Díaz, Eduardo Nogués, Núria PLoS One Research Article 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 (Di(a) and Lu(a)). 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. Public Library of Science 2016-09-07 /pmc/articles/PMC5014343/ /pubmed/27603310 http://dx.doi.org/10.1371/journal.pone.0161968 Text en © 2016 González et al http://creativecommons.org/licenses/by/4.0/ This is an open access article distributed under the terms of the Creative Commons Attribution License (http://creativecommons.org/licenses/by/4.0/) , which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited.
spellingShingle Research Article
González, Cecilia
Esteban, Rosa
Canals, Carme
Muñiz-Díaz, Eduardo
Nogués, Núria
Stabilization of Transfected Cells Expressing Low-Incidence Blood Group Antigens: Novel Methods Facilitating Their Use as Reagent-Cells
title Stabilization of Transfected Cells Expressing Low-Incidence Blood Group Antigens: Novel Methods Facilitating Their Use as Reagent-Cells
title_full Stabilization of Transfected Cells Expressing Low-Incidence Blood Group Antigens: Novel Methods Facilitating Their Use as Reagent-Cells
title_fullStr Stabilization of Transfected Cells Expressing Low-Incidence Blood Group Antigens: Novel Methods Facilitating Their Use as Reagent-Cells
title_full_unstemmed Stabilization of Transfected Cells Expressing Low-Incidence Blood Group Antigens: Novel Methods Facilitating Their Use as Reagent-Cells
title_short Stabilization of Transfected Cells Expressing Low-Incidence Blood Group Antigens: Novel Methods Facilitating Their Use as Reagent-Cells
title_sort stabilization of transfected cells expressing low-incidence blood group antigens: novel methods facilitating their use as reagent-cells
topic Research Article
url https://www.ncbi.nlm.nih.gov/pmc/articles/PMC5014343/
https://www.ncbi.nlm.nih.gov/pubmed/27603310
http://dx.doi.org/10.1371/journal.pone.0161968
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