Flow cytometric analysis and quantification of MPs in both ex vivo LPS-treated whole blood and in vivo endotoxin volunteers. Determination of forward (FSC) and side-scatter (SSC) characteristics of light by 1.0-μm latex beads in buffer was used to establish the MP (or R1) gate (A). The R2 (or bead) gate includes 7.2-μm latex beads used for enumeration of MPs as described in “Materials and methods.” Detection of phosphatidyl-serine (PS)-positive MPs by annexin V-Cy5 labeling on the y-axis in relation to light scatter (SSC) on the x-axis (B). Determination of the limit for negative fluorescence, performed in the presence of EDTA, was performed as a negative control for annexin V (C). MPs isolated from whole blood following ex vivo stimulation by LPS are shown in panels D and E. In the remaining panels (F-I), MPs isolated from whole blood of volunteers exposed to endotoxin are shown. In all cases, MPs were triple-labeled for annexin V (not shown) with anti-CD14-PE (y-axis) and anti-TF-FITC (x-axis) (D,F), or anti-CD144-PE (y axis) and anti-TF-FITC (x-axis) (H) or relevant isotype control IgG-PE (y-axis) and IgG-FITC (x-axis) (E,G,I).