Quantitative assessment of platelets, platelet microparticles, and platelet aggregates with flow cytometry

With fluorescent beads it has become possible to determine absolute numbers of cells in a given sample instead of relative percentages on a standard flow cytometer. This study assesses the ability to count platelets, microparticles, and aggregates with a flow cytometer. Whole blood was stimulated with 0.1 U thrombin per milliliter. Platelet and microparticle counts decreased, while the number of aggregates increased. Unactivated whole blood was diluted with buffer and showed a corresponding decrease in the concentration of platelets, microparticles, and aggregates. The platelet count on the flow cytometer was always in good correlation with counts on an automated blood analyzer. Only the cytometer, and not the automated analyzer, was able to detect and count microparticles and aggregates. In highly diluted samples of unactivated whole blood there was a spurious relative increase in CD62p-positive platelets because of a surplus of anti-CD62p antibodies and a relative increase in microparticles. Flow cytometry is a valuable method for counting platelets, aggregates, and microparticles in unstimulated and activated blood samples. If the platelet count changes and drops to less than 50% of the count for which the amount of antibody and the cytometer settings have initially been adjusted, care has to be taken to avoid misinterpretation.