Change of platelet activation markers using flow cytometry in patients with hematology/oncology disorders after transfusion.

In spite of the frequent need of platelet transfusions, there is limited information on the association of platelet activation markers, in transfused patients with hematology/oncology disorders, with platelet function using flow cytometry. The goal of this study was to evaluate the changes of PAC-1 binding and CD62P expression, with or without agonists in patients after transfusions. Twenty-eight whole blood samples were obtained from 24 patients admitted to the department of Hematology & Oncology and transfused with platelets; these samples were compared to 30 healthy controls. Whole blood samples, either with or without agonists, such as 20 microM adenosine diphosphate (ADP) or 100 microM thrombin receptor activating peptide (TRAP), were stained with the fluorescein conjugated monoclonal antibodies PAC-1 or CD62P. Then, the percent expression for each marker was analysed using flow cytometry. ADP and TRAP induced an increased percentage of CD62P expression and PAC-1 binding after platelet transfusions compared to the samples studied before transfusion, and these findings were lower than those of the healthy controls. However, the expression of platelets without the agonists was not significantly changed, despite the transfusions. Therefore, agonist-induced platelet activation markers, studied by flow cytometry, appear to be more useful for the evaluation of platelet function after transfusions than platelet activation markers without agonists.

[Establishment of reference values for platelet activation markers by flow cytometry.].

BACKGROUND: This study was purposed to establish reference values for platelet activation markers and leukocyte-platelet aggregates in the evaluation of the platelet function tests using flow cytometry. METHODS: Whole blood samples were obtained from 30 volunteers of healthy adults. Diluted blood samples, either in the resting state or in activated state by the addition of agonist, 20 microM ADP or 100 microM TRAP, were stained with fluorescent conjugated monoclonal antibody of PAC1 or CD62P. Then, the percentages of expression for each marker were analyzed by flow cytometry. For leukocyte-platelet aggregates, monoclonal antibodies of CD41a, CD14 and CD45 were added simultaneously to undiluted whole blood. RESULTS: Reference values for the percentages of the expression of PAC1 and CD62P, respectively, were 0.1-12.5% and 0.0-4.7% at the resting state, 65.3-92.4% and 39.0-75.7% with the addition of 20 microM ADP, and 68.1-93.1% and 60.5-91.2% with the addition of 100 microM TRAP. Reference values for leukocyte-platelet aggregates, granulocyte-platelet aggregates, and lymphocyte-platelets aggregates were 2.8-23.6%, 5.3-34.2%, and 4.9-21.6%, respectively. CONCLUSIONS: The platelet activation markers at the resting or an activated state with agonists and leukocyte-platelet aggregates could be analyzed using flow cytometry. These reference values should be helpful in interpreting platelet function tests by flow cytometry.