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Updated: May 3, 2026
Techniques for the Analysis of Extracellular Vesicles Using Flow Cytometry
Published on: March 17, 2015
A flow cytometric method for characterization of circulating cell-derived microparticles in plasma
Morten Hjuler Nielsen1, Henning Beck-Nielsen2, Morten Nørgaard Andersen3
1Danish PhD School of Molecular Metabolism, Faculty of Health Sciences, University of Southern Denmark, Odense, Denmark.
Background And Aim:
Previous studies on circulating microparticles (MPs) indicate that the majority of MPs are of a size below the detection limit of most standard flow cytometers. The objective of the present study was to establish a method to analyze MP subpopulations above the threshold of detection of a new generation BD FACSAria™ III digital flow cytometer.
Methods:
We analyzed MP subpopulations in plasma from 24 healthy individuals (9 males and 15 females). MPs were identified according to their size (<1.0-µm), by Lactadherin-FITC labelling, and by exposure of cell-specific markers. The sensitivity of the flow cytometer was tested against that of a previous-generation instrument FC500. Reproducibility of the FACSAria and our set-up was investigated, and the percentage of phosphatidylserine (PS) exposing MPs binding Lactadherin was determined.
Results:
By using a flow cytometric approach we identified and quantitated MPs derived from platelets, monocytes, erythrocytes and endothelial cells. In addition, levels of tissue factor-positive MPs were determined. The FACSAria demonstrated improved sensitivity and increased MP detection range compared to the FC500 instrument. The reproducibility of PS+PMP and PS+MP measurements was 11.7 and 23.2%, respectively. When expressed as a percentage of total MPs, the PS-positive MP population represented 15.1±5.5%, and PS-positive MPs were significantly increased in men.
Conclusion:
We have established a method to measure MPs above the detection limit of a new generation flow cytometer and derived from a number of cell-types in a healthy population of men and women.
Insights
This study developed a new flow cytometry method to detect small circulating microparticles (MPs) in healthy individuals. The new method, using the BD FACSAria™ III, improved detection sensitivity and identified various MP subpopulations, with higher levels of phosphatidylserine-exposing MPs found in men.
Area of Science:
- Biomedical Engineering
- Flow Cytometry
- Hematology
Background:
- Circulating microparticles (MPs) are crucial biomarkers, but many are smaller than the detection limit of standard flow cytometers.
- Previous flow cytometry instruments had limited sensitivity for analyzing the majority of microparticles.
Purpose of the Study:
- To establish a novel flow cytometry method for analyzing microparticle (MP) subpopulations.
- To leverage a new generation BD FACSAria™ III digital flow cytometer for enhanced MP detection.
Main Methods:
- Plasma samples from 24 healthy individuals were analyzed for MP subpopulations.
- MPs were identified by size (<1.0-µm), Lactadherin-FITC labeling, and cell-specific markers.
- The sensitivity and reproducibility of the BD FACSAria™ III were evaluated against a previous-generation instrument.
Main Results:
- The BD FACSAria™ III demonstrated superior sensitivity and a wider MP detection range compared to the FC500.
- Microparticle subpopulations from platelets, monocytes, erythrocytes, and endothelial cells were identified and quantified.
- Phosphatidylserine (PS)-exposing MPs constituted 15.1±5.5% of total MPs, with significantly higher levels observed in men.
Conclusions:
- A robust method for measuring microparticles (MPs) above the detection limit of advanced flow cytometers was successfully established.
- The method allows for the characterization of diverse MP subpopulations in a healthy population.
- Findings indicate potential sex-based differences in PS-exposing MP levels.
