Investigation of platelet–neutrophil interactions in whole blood by flow cytometry

Abstract

Evidence is increasing that platelets can initiate and propagate inflammatory processes by interacting with leucocytes and the vascular endothelium. Platelets have been shown to bind to neutrophils, existing as platelet/neutrophil complexes (PNC) within the circulation. We describe a simple flow cytometric method for assessing and investigating platelet interactions with neutrophils in small volumes of whole blood. Twenty-five percent (sd 6%) of circulating neutrophils from healthy adults were associated with platelets. Formation of these platelet–neutrophil complexes was CD62P (P-selectin) and divalent cation dependent. Platelet activation (with ADP or thrombin) caused a rapid and sustained rise in %PNC which differed from the pattern of free platelet activation as assessed by CD62P expression. F–met–leu–phe induced neutrophil activation but did not increase the percentage PNC. Platelet activation also caused increased neutrophil CD11b/CD18 expression which was most marked on neutrophils complexed with platelets. This straightforward technique is simple, reproducible, and allows assessment of platelet–neutrophil interactions and activation of neutrophils. It may also provide a method for estimating platelet activation in whole blood.