BACKGROUND AND OBJECTIVES: Athletes may illegally enhance endurance performance by transfusing homologous red blood cells (RBCs) and thereby increasing the oxygen carrying capacity of their blood. Detecting this dangerous practice is difficult by currently used methods. The aim of this work was to develop tests capable of detecting a mixed red cell population by flow cytometry, utilizing the likelihood of differences in minor blood group antigens. DESIGN AND METHODS: Twelve antisera directed against blood group antigens, derived from donor plasma, were used in conjunction with a secondary antibody directly conjugated with fluorescein to label IgG-coated RBCs. Optimal concentrations of RBCs and antibodies were determined on panel cells used in blood banking for the identification of specific antibodies. Blood samples from 25 patients purportedly transfused with 1-3 units of RBCs were screened for evidence of transfusion, and the percentages of antigen-positive and antigen-negative red cells were automatically calculated by the software installed in the flow cytometer after setting gates around these populations on histograms of fluorescence. RESULTS: Mixed RBC populations were identified in 22 of 25 patients tested. The three patients with antigenically homogeneous populations of RBCs were subsequently found not to have received their scheduled transfusions. INTERPRETATION AND CONCLUSIONS: This technique can detect small (<5%) populations of cells that are antigenically distinct from an individual's own RBCs. These results show the potential for flow cytometry to identify illicit homologous blood transfusion in athletes, and suggest the risk of false positives may be low.
BACKGROUND AND OBJECTIVES: Athletes may illegally enhance endurance performance by transfusing homologous red blood cells (RBCs) and thereby increasing the oxygen carrying capacity of their blood. Detecting this dangerous practice is difficult by currently used methods. The aim of this work was to develop tests capable of detecting a mixed red cell population by flow cytometry, utilizing the likelihood of differences in minor blood group antigens. DESIGN AND METHODS: Twelve antisera directed against blood group antigens, derived from donor plasma, were used in conjunction with a secondary antibody directly conjugated with fluorescein to label IgG-coated RBCs. Optimal concentrations of RBCs and antibodies were determined on panel cells used in blood banking for the identification of specific antibodies. Blood samples from 25 patients purportedly transfused with 1-3 units of RBCs were screened for evidence of transfusion, and the percentages of antigen-positive and antigen-negative red cells were automatically calculated by the software installed in the flow cytometer after setting gates around these populations on histograms of fluorescence. RESULTS: Mixed RBC populations were identified in 22 of 25 patients tested. The three patients with antigenically homogeneous populations of RBCs were subsequently found not to have received their scheduled transfusions. INTERPRETATION AND CONCLUSIONS: This technique can detect small (<5%) populations of cells that are antigenically distinct from an individual's own RBCs. These results show the potential for flow cytometry to identify illicit homologous blood transfusion in athletes, and suggest the risk of false positives may be low.
Authors: Francesco Donati; Xavier de la Torre; Sarajane Pagliarosi; Daniela Pirri; Giuliana Prevete; Francesco Botrè Journal: Front Sports Act Living Date: 2022-02-10