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Background

Background: Platelet-monocyte complex (PMC) formation is a marker of in vivo platelet activation and may be readily measured by flow cytometry. Due to the high frequency of free platelets relative to monocytes and PMCs, false-positive identification through coincidence remains a significant technical problem.To overcome this problem, we evaluated the use of a doublet-discriminator strategy (DDM) to allow faster sample acquisition whilst significantly reducing aberrant coincidence.Methods: Fourteen healthy volunteers and 20 patients with coronary artery disease (CAD) gave arterial and/or peripheral venous blood samples (NaCit). Whole blood was labelled in duplicate with anti-CD61 and anti-CD14 using a standard lyse/wash protocol. One of each paired sample was serially diluted before analysis; the second was analyzed at full concentration but using FL1-width to exclude co-incident platelet and monocyte events. Control experiments were performed with ex vivo thrombin activated samples.Results: With the DDM use PMC frequencies in the peripheral blood of healthy individuals and in CAD patients fell significantly [6.27% 6 1.77 (mean 6 sd) to 2.57% 6 0.99 (P 5 0.02)] and from16.04% (611.26) to 7.66% (65.18) (P < 0.01), respectively. DDM use significantly reduced the percentage of PMCs in the ex vivo thrombin activated samples (P < 0.05).Conclusions: Use of DDM effectively reduces the coincidence and enumerates true PMC in the samples of normal individuals and in patients with CAD and in ex vivo thrombin activated samples.