Introduction

This chapter will review the use of flow cytometry for the detection of circulating activated platelets and the analysis of many other aspects of platelet function. Flow cytometry rapidly measures the specific characteristics of a large number of individual cells. Before flow cytometric analysis, cells in suspension are fluorescently labelled, typically with a fluorescently conjugated monoclonal antibody. In the flow cytometer, the suspended cells pass through a flow chamber and, at a rate of 1000–10000 cells per minute, through the focused beam of a laser. After fluorescent activation of the fluorophore at the excitation wavelength, a detector processes the emitted fluorescence and light scattering properties of each cell. (Ref. provides a very readable overview of the principles of flow cytometry.)

In the absence of an added exogenous platelet agonist, whole blood flow cytometry can determine the activation state of circulating platelets, as judged by the binding of an activation-dependent monoclonal antibody. In addition to this assessment of platelet function in vivo, inclusion of an exogenous agonist in the assay enables analysis of the reactivity of circulating platelets in vitro. In the latter application, whole blood flow cytometry is a physiological assay of platelet function in that an agonist results in a specific functional response by the platelets: a change in the surface expression of a physiological receptor (or other antigen or bound ligand), as determined by a change in the binding of a monoclonal antibody.