Circulating microparticles are submicron vesicles released from cell membranes in response to activation or apoptosis. Acknowledgment of their role both as markers and pathogenic effectors in thrombosis, inflammation, and the spread of cancer has increased the interest of their measurement in clinical practice. However, assessment of their clinical use is impeded by technological issues. Among the different methodologies available, flow cytometry is the most commonly used technique. This review addresses flow cytometry limitations in microparticle measurement that may be subdivided into three domains: sizing, probing, and counting. This article also covers the various standardization strategies and technological improvements that have been proposed to overcome these limitations. New tools using size-calibrated beads and recent progress in instrumentation have opened new avenues to improve detection of microparticle populations of smaller sizes. Significant optimization in microparticle detection is also expected from the use of new fluorescent dyes and the establishment of practical recommendations. Finally, absolute counting of microparticles will also benefit from adapted bead-based strategies or, alternatively, from the generalized availability of volumetric systems. Overall, recent technological improvements maintain flow cytometry as a highly competitive analytical method to measure microparticles. Challenging these evolutions in pathological situations is a mandatory step to validate their real impact in clinical practice.
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